Post on 11-Feb-2022
www.heatox.org Project no. 506820
HEATOX
Heat-generated food toxicants: identification, characterisation and risk minimisation
Final report Instrument: Specific Targeted Research Project Thematic Priority: 5. Food Quality and Safety Period covered: From 1 November 2003 to 28 February 2007 Date of preparation: 12 April 2007 Start date of project: 1 November 2003 Duration: 40 months Project coordinator name: Kerstin Skog Project coordinator organisation name: Lunds Universitet
FOOD QUALITY AND SAFETY
Page 2 of 77
Content 1. Summary ....................................................................................................... 3 1.1 Detailed objectives..........................................................................................5
1.2. HEATOX Partners.................................................................................................................6 1.3. PhD’s ....................................................................................................................................7 1.4. External Panel......................................................................................................................8 2. Formation and Processing.............................................................................. 9
2.1. Chemical mechanisms and kinetics of formation of hazardous compounds.......................9 2.2. New and improved processing technologies to minimise the formation of hazardous compounds.....................................................................................................................................13
2.2.1. Potato....................................................................................................... 13 2.2.2. Bread ....................................................................................................... 19
3. Analysis ....................................................................................................... 21 3.1. Method development.............................................................................................................21 3.2. Analytical results on heat-induced toxicants levels in foods .............................................23 3.3. Analytical method for biomarkers ........................................................................................25
3.3.1. Identification of DNA-adducts in vivo............................................................. 25 3.3.2. Acrylamide metabolites as biomarkers of exposure.......................................... 26
4. Hazard characterisation............................................................................... 30 4.1. In-vivo genotoxicity and carcinogenicity .............................................................................30
4.1.1. Data on the relationship between internal dose and estimated intake of acrylamide in blood samples from a biobank............................................................................ 32
4.2. Tumorigenicity .......................................................................................................................34 4.2.1. Reproductive toxicity .................................................................................. 34 4.2.2. Neurotoxicity ............................................................................................. 35
5. Exposure Assessment .................................................................................. 37 5.1. Monte Carlo Risk Assessment...............................................................................................37
5.1.1. Dietary exposure assessment....................................................................... 37 5.1.2. Scenario-analysis of effect of new processing techniques.................................. 37 5.1.3. Brand loyalty ............................................................................................. 38 5.1.4. Home cooking............................................................................................ 38 5.1.5. Intake in British and Dutch infants ................................................................ 39
5.2. Training on probabilistic exposure assessment ..................................................................39 5.3. Estimation of breast cancer risk from human exposure to heat-induced toxicants.........39 5.4. Measurement of internal doses of acrylamide and glycidamide – a tool for exposure assessment and risk assessment.................................................................................................41
5.4.1. Haemoglobin adducts as a measure of internal dose........................................ 41 5.4.2. Bioavailability of acrylamide in food .............................................................. 41 5.4.3. Endogenous formation of acrylamide............................................................. 41 5.4.4. Inter- and intra-variation of internal doses in humans...................................... 41 5.4.5. Relationship between internal dose and estimated acrylamide intake from food ... 42
6. Risk assessment and risk communication .................................................... 43 6.1. Risk assessment ....................................................................................................................43
6.1.1. Models for extrapolation of cancer risk........................................................... 43 6.1.2. Final Risk Characterisation........................................................................... 43
6.2. Risk communication ..............................................................................................................45 6.2.1. Risk Communication Strategy....................................................................... 46
Acknowledgement………………………………………………………………………………….49 Publishable results of the Final plan for using and disseminating the knowledge……..………… 50 Dissemination of Knowledge……………...…………………………………………………........54
Page 3 of 77
Final report – Contract FOOD-CT-2003-506820
Heat-generated food toxicants: Identification, Characterisation and Risk Minimisation (HEATOX)
1. Summary
Heating food gives many advantages – it adds taste, colour and texture and minimises harmful germs. Flavour and aroma compounds are produced via the Maillard reaction but at the same time various health hazardous compounds may form. The main route for acrylamide formation goes via the Maillard reaction between reducing sugars (glucose and fructose) but also sucrose, and the amino acid asparagine. The HEATOX project involved 24 partners in 14 countries. The focus of the HEATOX project was health risks associated with hazardous compounds, for example acrylamide, in heat-treated carbohydrate-rich foods. The main objectives were to estimate health risks that may be associated with hazardous compounds in heat-treated food and to find cooking/processing methods which minimise the amounts of these compounds, thereby providing safe, nutritious and high-quality foodstuffs. Among the important results are recommendations to consumers, restaurants and the food industry on how to minimise the amounts of heat-generated toxicants in foods, while ensuring product quality from a nutritional and sensory point of view.
The HEATOX project started on the 1st November 2003 and had a duration of 40 months. Project web-site: www.heatox.org Co-ordinator: Kerstin Skog, Lund University, Sweden Department of Food Technology, Engineering and Nutrition e-mail: heatox@inl.lth.se Scientific Officer: Jürgen Lucas, e-mail: jurgen.lucas@ec.europe.eu The HEATOX project has dealt with questions regarding heat-generated food toxicants e.g.
• In which foods are these compounds mainly found? • How are they formed? • Do they constitute a health risk? • How can we measure/control the amounts produced? • How much is consumed? • What are the effects on the human body? • How can they be avoided? • Is there a cooking method to be recommended?
This European Research Project is supported within the European Commission's 6th Framework Programme on Research, Priority 5 on Food Quality and Safety (Contract n° Food-CT-2003-506820 Specific Targeted Project). This publication reflects the author’s views and not necessarily those of the EC. The information in this document is provided as is and no guarantee or warranty is given that the information is fit for any particular purpose.
Page 4 of 77
Modern science has showed that heating of meat and other protein rich foods can generate various kinds of potentially hazardous compounds, some of which are genotoxic and carcinogenic. The focus of the HEATOX project was health risks associated with hazardous compounds in heat treated carbohydrate-rich foods where substantial amounts of acrylamide and similar compounds can be formed. HEATOX explored their mechanisms of formation, impact of raw material composition, inhibiting factors, cooking and processing methods in industry and households with the to aim control and minimise the formation of hazardous compounds. Acrylamide was given particular emphasis in the HEATOX project. Other compounds representing potential health hazards, such furans, are also formed during heating, thus it is important that efforts in reducing acrylamide exposure at the same time do not increase the formation of other hazardous compounds. Validated methods for food analysis and exposure biomarkers have been developed. Different hazards have been explored and characterised in various toxicological models, for example genotoxicity, carcinogenicity, neuro-developmental and reproductive toxicity. Molecular characterisation by toxicogenomics has been performed. Experimental animal and cell systems and also humans were studied. Particularly the relevance of low dose exposure, bioavailability and extrapolation from experimental systems to humans was addressed using biomarkers of exposure and effect. The exposure assessment and data on hazard characterisation, including data generated outside HEATOX, were combined in a risk characterisation of intake of heat-treated carbohydrate rich foods. To support and strengthen the project profile, a risk communication strategy was developed early in the project. A tailored action plan was implemented successively during the whole project period. The ultimate aim was to ensure that project deliverables were as useful and focused as possible and that the dialogue within HEATOX and between HEATOX and the outside world was effective. The HEATOX project co-operated with other research activities carried out by other groups around the world, e.g. the project was linked with the COST 927 action. Dissemination of the project results was a key issue, which involved for example publications, conferences, and the HEATOX workshops.
Formation Exposure Assessment Hazard characterisation Analysis
Risk Assessment
Management, Communication, Dissemination and Training
Formation
Industrial Processing
Household Cooking
External Exposure
Internal Exposure
DNA damage
Non- genetic damage
Mutation, Cancer
Fertility
Neurological effects
Other Health
Effects
Project overview
Page 5 of 77
1.1 Detailed objectives: I. To elucidate the chemical reaction mechanisms involved and to develop and validate new/improved production methods and technological minimisation strategies ♦ Effects of variation in the composition of the raw material ♦ Reaction pathways and mechanisms, inhibiting/enhancing compounds, and kinetics ♦ Cooking and processing methods for the food industry, catering facilities and households Expected results: Recommendations on how to limit the formation of heat-induced toxicants in order to minimise the formation of heat-induced toxicants, while ensuring product quality from safety, nutritional and sensory points of view. II. To provide validated analytical methods (i.e. screening and confirmatory) and quality-controlled analytical data needed for the execution of the other tasks • Quantitative analytical methods for biomarkers and for toxicants in foods • Identification methods and data on new toxicants or known toxicants in new food matrices • Quality-controlled data on toxicant levels in foods Expected results: Validated analytical methods for a wide range of heat-induced toxicants and for biomarkers of exposure. III. To perform hazard characterisation ♦ Neurodevelopmental toxicity, mechanisms of tumourgenicity, and mechanisms of reproductive
toxicity ♦ Bioavailability in animals and humans determined by different biomarkers for internal dose and
effect ♦ Dose-response relationships in the low-dose region Expected results: Characterisation of toxic effects including their mechanisms and dose-response relationships. IV. To assess the exposure of heat-induced toxicants ♦ Improved estimation of the intake of heat-induced toxicants based on analysis of cooked foods and
food intake data, especially concerning variability and risk groups ♦ Comparison of dietary intake data regarding heat-induced toxicants with data on the internal dose
(heat-induced toxicants adduct levels, e.g. DNA-adduct levels) ♦ Elucidation of the role of possible endogenous formation of heat-induced toxicants ♦ Assessment of the possibility of using statistics for conclusive cancer epidemiology studies Expected results: Validated data on heat-induced toxicants exposure and intra- and inter-individual variation in exposure. V. To perform risk assessment and communicate the results of the project ♦ Assessment of the health risk associated with the current exposure to known heat-induced
toxicants (risk characterisation) and possibly new compounds, also taking into account existing information on hazards from sources outside the project
♦ Comparative risk/benefit assessments of heat-treated foods by linking results from animal and human studies to human health risk
♦ Targeted recommendations and guidelines to all important user groups ♦ Broad information to the general public, the food industry and its different relevant
associations/federations, national food agencies, consumer groups/associations, EFSA, different Commission services (e.g. DG Research, DG Health and Consumer Protection, DG Agriculture, DG Enterprise, etc.) on the project, its results and their implications including also future aspects.
Expected results: Increased knowledge and harmonised agreement on the possible risks of heat-induced food toxicants and recommendations on how the toxicant levels can be controlled.
Page 6 of 77
1.2 HEATOX Partners
P 1 Lund University (Sweden) • Kerstin Skog, Co-ordinator P 2 Graz University of Technology (Austria) • Michael Murkovic P 3 The University of Reading (UK) • Don Mottram P 4 Swedish University of Agricultural Sciences (Sweden) • Per Åman P 5 University of Bologna (Italy) • Marco Dalla Rosa P 6 Swedish Institute for Food and Biotechnology (Sweden) • Hans Lingnert P 7 Wageningen University (The Netherlands) • Tiny van Boekel P 8 Central Science Laboratory, York (UK) • Laurence Castle P 9 Swedish National Food Administration (Sweden) • Karl-Erik Hellenäs P10 Institute of Chemical Technology Prague (Czech Republic) • Jana Hajslova P11 Agrotechnology and Food Innovations (The Netherlands) • Charon Zondervan P12 University of Barcelona (Spain) • Maria Teresa Galceran P13 TÜBÝTAK-Marmara Research Center (Turkey) • Hülya Ölmez P14 Stockholm University (Sweden) • Margareta Törnqvist P15 National Food Institute (Denmark) • Henrik Frandsen P16 Norwegian Institute of Public Health (Norway) • Jan Alexander P17 RIKILT Institute of Food Safety (The Netherlands) • Jacob van Klaveren P18 German Institute for Human Nutrition (Germany) • Hansruedi Glatt P19 University of Leeds (UK) • Bronek Wedzicha P20 BEUC, European Consumers' Organisation (Belgium) • Barbara Gallani P21 National Veterinary Institute (Norway) • Helga Odden Reksnes P22 University of Zürich (Switzerland) • Hanspeter Naegeli P23 University of Chile (Chile) • Lilia Masson P24 Queens’ University Belfast (UK) • Chris Elliott
Page 7 of 77
1.3 PhD’s Exchange of young scientist has been encouraged within the HEATOX project; several young researchers have visited other HEATOX partners for 1-4 weeks to learn new methods, plan collaborative work, discuss results etc. The successful integration of the young researchers with senior researchers and cooperation between the different groups has contributed to interdisciplinary relations and building of networks. Several partners participate in Cost Action 927 “Thermally Processed Foods: Possible Health Implications”, co-ordinated by V Fogliano (University of Naples, Italy), which has provided additional opportunities for student exchange. Special emphasis has been put on communication training for PhDs and young researchers. At the semi annual meetings, seminars have been arranged on the topic of risk communication and dissemination in relation to their HEATOX work, for example short oral presentations, posters and also participation in the HEATOX workshop. PhD students/Young scientists working in the HEATOX project: German Institute of Human Nutrition (Germany) • Yasmine Sommer Graz University of Technology (Austria) • Kristina Bagdonaite Institute of Chemical Technology Prague (Czech Republic) • Lenka Dunovska Leeds University • Jonas Mojica Lazaro Lund University (Sweden) • Peter Viberg, Gunilla Viklund National Food Institute (Denmark) • Pelle T Olesen Norwegian Institute of Public Health (Norway) • Thomas Bjellaas, Siri Helland Hansen, Hege Benedikte Ölstörn RIKILT Institute of Food Safety (The Netherlands) • Anika de Mul Stockholm University (Sweden) • Anna Vikström Swedish National Food Administration (Sweden) • Louise Durling, Erik Petersson Swedish University of Agricultural Sciences (Sweden) • Arwa Mustafa The University of Reading (UK) • Mei Yin Low University of Barcelona (Spain) • Soubhi Altaki, Erika Teixido University of Bologna (Spain) • Alessandro Angioloni, Mara Bacchiocca, Andrea Gasparri, Pietro Rocculi University of Zürich (Switzerland) • Flurina Clement, Ramiro Dip Wageningen University (The Netherlands) • Jeroen Knol Queen’s University Belfast (UK) • Andrew Preston
Page 8 of 77
1.4 External Panel The External Panel constituted the core risk communication network and provided valuable discussion with HEATOX’ stakeholders. In charge: Karl-Erik Hellenäs. Members: Consumer • Barbara Gallani (from June 2005)/Beate Kettlitz (until March 2005), BEUC, BE
Industry • Beate Kettlitz (from March 2005)/Domenique Teaymans (until March 2005), CIAA, BE • Sam Lalljie, ILSI, BE • Julia Gelbert, BLL/ZUTECH, GE
Research • David Lineback, JIFSAN, USA • Erland Bråthen (until 2005), MATFORSK, NO • Vincenzo Fogliano, COST-927, IT • Elke Anklam (until July 2006), JRC irmm, BE
Authorities • Almut Bitterhof (from spring 2006)/Martin Slayne (until spring 2006), DG Sanco, BE • Claudia Heppner, EFSA, Contam panel, IT • Lutz Dehne, BfR, DE • Wendy Matthews (from 2004)/Karen Goonan (until 2004), FSA, UK • Sara Henry, FDA, USA • Lauren Jackson, FDA, USA Management & Information Flow
Management Formation, Processing
Analysis Hazard characterisation
Exposure Risk assessment, communication
Scientific Committee
Coordinator
EC- DG Research
External consulting panel
(One representative per partner incl. Coordinator)
(Theme leaders and coordinator)
Technical Project Assistant
WP 1 WP 2.1, 2.2 WP 3.1, 3.2, 3.3
WP 4.1, 4.2, 4.3
WP 5.1, 5.2, 5.3
WP 6.1, 6.2
Board of Managers
Page 9 of 77
2. Formation and Processing Wageningen University (The Netherlands), University of Leeds (United Kingdom),
The University of Reading (United Kingdom), Graz University of Technology (Austria), Lund University (Sweden), Agrotechnology & Food Innovations B.V. (The Netherlands), University of Chile, University of Bologna (Italy),
Swedish Institute for Food and Biotechnology (Sweden), Swedish University of Agricultural Sciences (Sweden)
2.1. Chemical mechanisms and kinetics of formation of hazardous compounds The Maillard reaction between amino acids and sugars is essential for the development of desirable colour and flavour in baked and fried foods. However, the Maillard reaction has been shown to generate acrylamide at the same time, through the reaction of the amino acid asparagine with reducing sugars. Thus, acrylamide is formed by the thermal reaction of natural food components, when moisture levels become low. A proposed mechanism for acrylamide formation (see Figure 1) involves the formation of a Schiff base from the reaction of a carbonyl compound with asparagine. Decarboxylation of the Schiff base, in a Strecker-type reaction, gives an unstable intermediate that can hydrolyse to 3-aminopropamide, which on elimination of ammonia yields acrylamide. Alternatively the decarboxylated Schiff base could form acrylamide via elimination of an imine.
Figure 1. Proposed mechanism for the formation of acrylamide in heat-treated foods.
O NH2
NH2 COOH
O NR
NH2OHO
O
O NR
NH2
H OO N
R
O
NH2
O
NH2
O NH2
NH2
O O
NH2
RO
H2O H2O
- H2O
- NH3
+
ASPARAGINE REACTIVE CARBONYL FROM
MAILLARD REACTIONSCHIFF BASE
STRECKERALDEHYDE3-AMINOPROPANAMIDE ACRYLAMIDE
Page 10 of 77
The formation of acrylamide in food is under kinetic control; this means that the amount which is formed depends on the rate of reaction and the time for which it has been allowed to proceed. Reaction kinetics relates to the rate limiting steps in chemical reactions; these are the control points. For this reason, an understanding of the factors which control the rate limiting steps in acrylamide formation offers opportunity to control the extent of its production. Chemical mechanisms and kinetics of formation were studied, based on reactions between sugars and amino acids via the Maillard reaction. Equimolar solutions of glucose and asparagine (0.1 and 0.2 M) were prepared in phosphate buffer (0.1 M) with different pH (4.5, 5.5, 6.0 and 6.8). Samples were heated at 120-200°C for 0-128 min. A mechanistic kinetic model has been derived for the aqueous model systems from the proposed reaction network (Figure 2). For each reaction step a differential equation was set up, and translated into a mathematical model.
Figure 2. Proposed reaction network for the formation of acrylamide from asparagine and glucose/ fructose through early Maillard reaction. Product X: product(s) formed from acrylamide breakdown.
Multiresponse modelling using non-linear regression with the determinant criterion was used to estimate model parameters, i.e. the rate constants and activation energies based on the experimental data obtained from the experiments with the aqueous model systems (Table 1).
Table 1. Estimates of rate constants (k) at 120, 140, 160, 180 and 200 ºC and activation energies (Ea) ± 95% Highest Posterior Density (HPD) interval as found by kinetic modelling for the proposed kinetic model for pH 6.8.
k (10-3 min-1 or·10-3 mol·min-1) 120 ºC 140 ºC 160 ºC 180 ºC 200 ºC Ea (kJ/mol)
k1 0.131 ± 0.025 0.308 ± 0.049 0.668 ± 0.13 1.35 ± 0.36 2.58 ± 0.89 57.6 ± 8.0
k2 4.98 ± 1.2 16.7 ± 3.0 50.1 ± 9.6 136 ± 35 341 ± 114 81.7 ± 8.6
k3 0.0819 ± 0.045 0.369 ± 0.14 1.45 ± 0.44 5.04 ± 1.6 15.8 ± 6.4 102 ± 14
k4 0.176 ± 0.081 0.712 ± 0.23 2.53 ± 0.51 8.05 ± 1.2 23.2 ± 4.3 94.4 ± 11
k5 15.7 ± 3.5 28.4 ± 4.6 48.7 ± 5.9 79.6 ± 8.9 125 ± 16 40.1 ± 5.0
k6 7.96 ± 5.1 28.1 ± 13 88.1 ± 25 250 ± 45 650 ± 136 85.1 ± 14
Page 11 of 77
Figure 3 shows how the model fits the experimental data for the formation of acrylamide in the aqueous reaction system at pH 6.8. The proposed model gives a reasonable estimation for the formation of acrylamide in an aqueous model system. The mechanistic kinetic model derived in this study is an important step into the realization of a tool that can be used to predict how acrylamide reduction in foods containing asparagine and reducing sugars can be accomplished.
Figure 3. Model fit (lines) to experimental data (symbols) of acrylamide concentration for glucose/ asparagine (0.2 mol/l, pH 6.8) aqueous reaction system heated at 120 (blue), 140 (red), 160 (purple), 180 (green) and 200 °C (light blue).
Another kinetic model for the formation of acrylamide in foods (potato, wheat and rye products) involves steps which lead to the destruction, binding or physical ‘loss’ of acrylamide. Our ability to formulate correctly the kinetic model for acrylamide levels in foods thus depends critically on us being able to formulate the kinetics of these subsequent processes. This is particularly important with regard to the control (i.e., reduction) of acrylamide levels because such ‘losses’ could be maximised through adjustment of the reaction conditions. Furthermore, there are views that perhaps much more acrylamide is actually formed in foods than is measured, and an understanding of the fate of acrylamide is important so that we might be able to identify the chemical nature of any ‘bound’ acrylamide and the stability of any reaction products which are formed. If acrylamide is being formed in much greater yield than so far observed, there is also a danger that a change in the reaction conditions could alter the balance between the rates of formation and of loss, so that much higher yields are obtained. We believe, therefore, that consideration of the kinetics of acrylamide loss is fundamental to the derivation of the kinetic model. The kinetics of the reaction of acrylamide with glycine and proline were determined in aqueous systems as a function of pH and temperature, and the relative reactivities of all 20 amino acids measured at pH 5.5. Maltodextrin gels were prepared containing acrylamide and the amino acids, and were freeze-dried before being heated to 180 °C. The experiments in aqueous systems have shown the relevance of Michael addition type reactions of amino acids with acrylamide. Acrylamide reacted more readily with glycine than with Maillard precursors in solution and this reaction presented second order kinetics, i.e.,
]lyk[acryl][gdt
d[acryl]=−
However, glycine was not the most reactive amino acid towards acrylamide; proline and cysteine showed much higher rates of reaction in comparison with other amino acids. pH was a significant factor in the reaction; glycine and proline react by a common mechanism in which there exists an acid-base catalytic effect by a species with pKa=5.2-5.5.
])glyNH[(][A 21-
0−+= kkk
where k is the apparent first order rate constant and A- is the suggested catalyst.
Page 12 of 77
The maltodextrin dry solid system demonstrated reactivity of amino acids towards acrylamide when heated in the absence of water at 180 ºC. The rate of loss of acrylamide in this system was very similar to that in starchy foods. In this model, most of the amino acids showed similar reactivity towards acrylamide, with the exception of lysine, which was higher. Lysine is also one of the most reactive amino acids in the Maillard reaction. The effect of temperature on the amino acid-acrylamide reaction followed the Arrhenius relationship, showing the highest Ea (83 kJ mol-1) for the reaction of acrylamide alone in the matrix. The reaction of glycine and proline with acrylamide showed lower but similar Ea of approximately 70 kJ mol-1. Our data indicates that acrylamide is lost from food by reaction with amino acids. This augments the already established effect of amino acids (e.g., glycine) on acrylamide formation by competing with asparagine for a key precursor of acrylamide, and explains published reports describing the effects of amino-containing food ingredients on acrylamide residues. Maltodextrin proved to be a good model system in which to study reactions of acrylamide with amino acids in the dry state, as evidenced by rates of reaction of acrylamide which are similar to those observed in foods. The relationships between the formation (and loss) of flavour compounds and acrylamide were measured in potato and rye. Simple cakes were prepared from flour and water and were heated at 180 C, from 10 to 60 min. The flavour compounds studied were Strecker aldehydes, which are formed relatively early in the Maillard reaction and may react further, and alkylpyrazines, which may be considered as end-products. The data obtained was used to create a kinetic model, which was then used to predict the effect of glycine on flavour and acrylamide formation in potato cakes. The relationships between the formation (and loss) of flavour compounds and acrylamide were different in potato and rye. In both potato and rye, Strecker aldehydes formed and decomposed in a similar way. However, in rye the rate of formation of the Strecker aldehydes was similar to that for acrylamide, whereas, in potato, acrylamide formed at a slower rate than in rye. Pyrazine formation in rye and potato mirrored acrylamide formation. However, acrylamide levels decreased on prolonged heating whilst pyrazine levels were maintained in both rye and potato. It appeared that moisture levels below 5% were needed for pyrazines and acrylamide to form in both foods. Potato cakes contained higher initial moisture levels and hence formation of both acrylamide and pyrazines occurred later in the cooking process for potato than for rye. The Maillard reaction could be responsible for potential toxicants other than acrylamide. Two databases, each containing chemical and toxicity data, have been compiled in Microsoft Access for Maillard compounds and for lipid-derived compounds. The Maillard database lists approximately 570 volatile compounds, while approximately 200 volatile compounds are listed as products of heated lipid systems. A molecular modelling approach has been used to assess the potential toxicity of each compound. Both databases are fully searchable and contain the following information: • Chemical name, Chemical structure, CAS number • Other names • Foods in which the compound is present • Foods in which the compound is present at levels above 1 ppm • Reaction mixtures in which the compound is present • EU Approved Flavourings Register FL No. • Predicted carcinogenicity probability • Predicted mutagenicity probability • Predicted rat oral LD50 (mg/kg) • Predicted chronic lowest observed adverse effect level (LOAEL) (mg/kg) • Predicted skin sensitisation probability The databases provide a useful source of information for chemists and toxicologists. They are an important starting point for future research into toxic compounds formed during cooking. The data bases can be found at www.heatox.org, Achievements.
Page 13 of 77
The formation of acrylamide during the roasting process of coffee was investigated in detail in lab scale using a roasting device that employs convection roasting. The model system was a mixture of asparagine with glucose or sucrose using a heating device that allowed the constant heating of small samples (max 300 mg) up to 300 °C with an efficient heat transfer. Studies using 13C-labelled asparagine showed that this amino acid is directly converted to acrylamide. Intermediates were synthesised (N-glucosyl-asparagine and the asparagine Amadori compound) that were converted to acrylamide by heating. At high temperatures – which are used during roasting of coffee – these two compounds could be identified as intermediate reaction products. A statistical analysis suggests that asparagine is the limiting compound in reactions leading to acrylamide in coffee. Sucrose is present in green coffee in concentrations up to 9%. A statistical analysis suggests that asparagine is the limiting compound in reactions leading to acrylamide in coffee. In coffee shorter roasting times and lower temperatures are leading to higher acrylamide concentrations. This means that darker roasted coffee – higher degree of roasting – has lower acrylamide concentrations. HMF (hydroxymethylfurfural) is formed in coffee very quickly which means that within 2 min a maximum concentration is achieved. After this time a quick degradation occurs to levels significantly lower (ca 100 µg/g). This means that HMF is not heat stable and reacts further during the roasting process to substances that are not known now. HMF is formed from sucrose with and without the participation of amino acids.
2.2. New and improved processing technologies to minimise the formation of hazardous compounds 2.2.1. Potato Potato has been a staple food for many decades and mainly consumed as boiled or fried potatoes. Our dietary habits have changed and today we are used to many different potato dishes and there are a number of potato products on the market. Potatoes are rich in starch, and the aim of heating potato is to make them palatable and to gelatinise the starch to make it available for digestion. At the beginning of the project a mindmap was made, se Figure 4 below. Such a mindmap serves to find relationships between the numerous variables that – in this case – potentially influence various quality parameters of French fries. With a special focus on acrylamide, the mindmap gave clusters of variables that were somehow related. These clusters of variables formed the basis of our three-year research programme.
Page
14 of
77
(loca
l)A
cryl
amid
eco
ncen
trat
ion
Form
atio
n ra
te o
fA
A
Con
vect
ive
tran
spor
t of A
A
Red
uctio
nrat
eof
AA
Diff
usio
n co
effic
ient
AA
Asp
arag
ine (A
SN)
conc
entra
tion
Acr
olein
Glyc
erid
es
reac
tion
cons
tant
k1 (v
ia ac
rolei
n)
reac
tion
cons
tant
k2 (v
ia A
SN)
gluco
se
fruct
ose
malt
ose
and/
orsu
cros
e co
ncen
tratio
n
Prod
uct
tem
pera
ture
(loca
l)
pH p
rodu
ctlo
cal
aw p
rodu
ct(lo
cal)
?
Hea
t pro
duct
ion
due
to 'c
hem
ical' r
eact
ions
Hea
t con
duct
ion
coef
ficien
tH
eat e
xcha
nge
coef
ficien
tm
edium
to p
rodu
ct
loca
l moi
sture
cont
ent
ion
conc
entra
tion
type
of
ions
redo
x po
tent
ial
type
of
heat
ingm
edium
(eg
oil)
Fatty
acid
con
tent
of th
e m
edium
Surfa
cero
ughn
ess o
fpr
oduc
t
type
of
varie
ty(p
otat
o)
cook
ing
prop
ertie
sof
the
pota
to
(sto
rage
-) a
ge p
otat
o
Cut
ting
tech
nolo
gy
tem
pera
ture
of t
hem
edium
Spec
ific h
eat o
fth
e pr
oduc
t
Prod
uct d
ensit
y
visco
sity
of th
ehe
ating
med
ium
turb
ulenc
e of
the
med
ium n
ear t
hepr
oduc
t sur
face
Form
atio
n of
lard
/ stre
cker
reac
tion
prod
ucts
Cru
st
Evap
orat
ion
ofw
ater
diffu
sion
coef
ficien
tof
wat
er in
side
the
prod
uct
Pota
to p
iece
(disk
,re
ctan
gle, s
pher
e)
Diam
eter
pota
to p
iece
Thick
ness
pota
to p
iece
Hea
t exc
hang
ingsu
rface
Vol
ume
pota
topi
ece
Ave
rage
AA
conc
entr
atio
n
Stat
us o
f the
starc
h
wat
erbi
nding
cap
acity
of th
e pr
oduc
t
(bio
)che
mica
lre
actio
n ki
netic
s
Prop
ertie
s of t
he h
eatin
gm
edium
and
use
d te
chno
logy
(e.g
. hot
air,
oil o
r bak
ing p
an)
effe
ct o
f loc
alco
nditio
ns
Effe
ct o
f cut
ting,
tech
nolo
gy a
ndsh
ape
of th
e pr
oduc
t
Prop
ertie
s of t
hepr
oduc
t
Star
t to
desc
ribe
the
prod
uct
quali
ty p
rope
rties
in m
ore
deta
il
Col
our
Tast
e an
dFl
avou
r
Diff
usio
nco
effic
ient o
fsu
gars
Glyc
erol
Hyd
rolys
e
?
Dry
mat
erial
con
tent
of th
e po
tato
Ripe
ness
of t
hepo
tato
es
Gro
wth
pat
tern
of th
e cr
op
Phys
iolo
gical
age
ofro
ot a
nd sp
out
Wat
er su
pply,
light
inten
sity,
day
leng
th, s
oil
cond
ition N
itrog
en su
pply
Calc
ium su
pply
Chlo
ride
supp
ly
Phos
phat
e su
pply
wat
er- a
nd m
inera
lint
ake
by th
e cr
op
Blan
ching
tem
pera
ture
Blan
ching
tim
eeffe
ctive
ness
of
blan
ching
Initia
l sug
arco
ncen
tratio
n of
the
pota
to
Suga
r con
cent
ratio
npo
tato
piec
e af
ter
blan
ching
Suga
r con
cent
ratio
n
Prop
ertie
s of c
rop
and
harv
ersti
ng
Diff
eren
ce b
etw
een
loca
lan
d av
erag
e A
A
Blan
ching
Initia
l am
ino a
cids
conc
entra
tion
ofpo
tato
Free
am
ino a
cidco
ncen
tratio
n of
pot
ato
piec
e af
ter b
lanch
ing
Fig
ure
4.
Min
dm
ap f
or
pota
to p
roce
ssin
g a
nd a
cryl
amid
e fo
rmat
ion
Page 15 of 77
2.2.1.1. Heating model: Fry Simulator – French fries A computational model was made to estimate the impact of various processing parameters on the formation of acrylamide in French fries. The model uses a cross section of a French fry of infinite length and is made up from a 40*40 grid. The model was validated by measuring temperature and water profiles in time and at various positions inside the French fries. At a later stage also acrylamide measurement were performed on parts of the fries. The latter results showed that although the trends are correctly predicted in the Fry Simulator, its value in absolute sense is limited. Moreover, since the model only calculates acrylamide in the middle, it leaves out the acrylamide that is formed on both ends of the fries. From experimental data it is well known that the highest levels are found at both ends. 2.2.1.2. French fries Experiments were performed with potatoes grown in the Netherlands. Batches of three varieties of 1000 kg each (Asterix, Bintje, Lady Olympia) were supplied by a potato processor, shortly after the harvest and were stored at 8°C until the end of June. In the varieties Asterix and Bintje the sugar concentrations increased during a 6 month storage period. The sugar contents of the Lady Olympia potatoes decreased during the storage. We also found that the acrylamide levels were positively correlated to the levels of reducing sugars. We found that acidic blanching prior to industrial frying is an efficient way to reduce acrylamide during final product preparation at home or in restaurants and caterers. When using 0.5% of citric or lactic acid, acrylamide levels are reduced some 80% while expert panels do not detect any acidic aftertaste. Other methods of introducing small amounts of acid onto the surface have been considered, such as spraying acid solutions on frozen fries. Such methods were expected to be no more efficient than acidic blanching but much more difficult to implement. 2.2.1.3. Potato crisps Panda Potatoes variety, crops 2004, 2005, 2006 were used for experiments and were pre-treated before frying at 120, 150 and 180 ºC: washing with distilled water, blanching at 90 ºC per 1 minute and immersion in dilute citric acid solution 1-0.25% for 1 hour. Experiments were performed in vacuum and at atmospheric pressure. Vacuum frying with and without pre-treatments was shown to produce low level acrylamide crisps, see Table 2. It is possible to improve their quality characteristics. The pre-treatments represent a practical industrial alternative to reduce acrylamide level in potatoes crisps.
Table 2: Effect of pre-treatments, frying methods and frying temperatures on acrylamide content and acceptability score (1-7) of crisps.
Pre-treatment Frying method
Blanching
Immersion citric acid
0,25% Atmospheric Vacuum
(50 mm Hg) Acrylamide
Acceptability
90ºC/1 min 1h Temp.ºC Temp.ºC ppb Score*
+ + 150 - 6 4,3 + + 160 - 5 4,3 + + 170 - 30 4,4 - - 150 - 87 2,1 - - 160 - 233 2,9 - - 170 - 760 2,9 + + - 150 ND 3,3 + + - 160 ND 3,5 + + - 170 4 3,5 - - - 150 80 4,6 - - - 160 218 5,3 - - - 170 267 5,1
*Score 1-7, with 7 as the best ND = Not Detected
Page 16 of 77
Blanching and change of pH by immersion of sliced raw potatoes in citric acid solution (0.25%), had a strong influence for reducing the acrylamide content in crisps fried at atmospheric pressure and in vacuum. Acrylamide content of potatoes crisps with pre-treatments fried in vacuum were very low between non detected levels up to 4 ppb. At atmospheric pressure with pre-treatments, the acrylamide content was between 6 up to 30 ppb. Without pretreatments, the acrylamide content of the crisps varied between 80 – 267 ppb for vacuum frying and at between 87 – 760 ppb for frying at atmospheric pressure. The sensory evaluation (trained and consumer panel) obtained as acceptability score ( 1 – 7), was better for the crisps with pretreatments fried at atmospheric pressure than in vacuum, 4.3 compared with 3.5. A commercial crisp sample sensory evaluated at the same time scored 4.8 with an acrylamide content of 536 ppb. The sensory panel concluded that the best crisps were obtained in the samples corresponding to control assays, submitted to vacuum frying processing at 160ºC and 170 C. Considering the assays with pre-treatments, the vacuum fried crisps got a better evaluation than the crisps fried at atmospheric pressure. Conclusion: Vacuum frying is a processing method not commonly used in the industry, however, it can be a very good alternative in the near future (acrylamide free crisps)
Raw Potatoes, Var Panda Blanching Citric Acid Immersion
Slices potatoes
Vacuum Fryer
Potato Crisps
Photos: Lilia Masson
Page 17 of 77
2.2.1.4. Raw materials selection
Potato field in the south of Sweden. Photo: K. Olsson, Svalöf Weibull
Well characterised Swedish-grown potatoes, including two breeding clones that are resistant to low-temperature sweetening were studied and batches were obtained from three consecutive years. The content of free amino acids and sugars in potato were studied in potatoes from three growing seasons. In some potatoes, the levels of reducing sugars were practically the same the first two years, but almost doubled the third year, while in another variety; the level of reducing sugars decreased to half the second year and increased the third year. The concentrations of asparagine were generally not so much affected by the growing season. These observations make it difficult to draw conclusions on which variety processing conditions that are best for crisp production because this may vary with year. Frying conditions were discussed with crisp producers and producers of frying equipment to establish laboratory conditions that would resemble industrial conditions. For industrial crisp production, potato slices are usually passed on a belt through a continuous fryer with a starting temperature around 180-190 C and an end temperature of around 150 – 175°C. Quality parameters such as crispiness and colour have to be taken in account. A water content below 2% is considered to give a good crispiness and shelf-life. Using genotypes with a wide range of precursor concentrations, we found that the asparagine levels affect the formation of acrylamide to a higher degree than earlier studies have shown. Crisps made from potatoes with the lowest asparagine content, generally had the lowest concentration of acrylamide regardless of storage temperature. In agreement with work by others, we found that storage at 4°C enhanced the concentration of sugars in he tubers and thus the acrylamide content compared with storage at 8°C. Blanching before frying was shown to efficiently reduce the acrylamide content in crisps. For potatoes stored at 4 C, the acrylamide content was generally reduced by at least 50%. The mass transport of precursors during heating may be important for acrylamide formation, but this requires further investigation.
Page 18 of 77
2.2.1.5. Home cooking Much work has been devoted to the mitigation of acrylamide in potato crisps and French fries, but literature data on acrylamide in home-cooked potato dishes is scare. Several home-cooking experiments were performed. Roasted potato wedges and pan fried boiled potato dices (a common Swedish dish) had an appetising appearance; the wedges were soft with a golden yellow surface. Acrylamide was found in all cooked samples. Blanching before oven roasting made it possible to reduce the heating time in the oven and yet obtain potato wedges with similar eating quality. Blanching combined with a shorter roasting time is an easy and efficient way of reducing the acrylamide content in oven roasted potato wedges.
The effect of frying in a commercial available domestic fryer was studied in relation to acrylamide formation in French fries. It was found that the frying temperature drops drastically and slowly reached its original temperature. The drop in temperature depends of the product/oil ratio and makes it difficult to define an initial oil temperature as a measure to reduce acrylamide.
The results showed that higher initial oil temperature causes a faster recover of oil temperature during frying, as well as a greater acrylamide formation rate. The choice of a lower initial frying temperature lowered the acrylamide content of the product. At the optimal cooking time in terms of culinary quality, the lower the potato quantity the lower acrylamide content was found. Our results underline that the kind of frying equipment (heating power), frying time, oil temperature, amount of potato immersed in the hot oil (product/oil ratio) have to be taken into account for optimisation of product quality (colour, texture, water and oil content) and acrylamide content in French fries. Since par-fried French fries do not contain acrylamide when leaving the factory, it is important to instruct users (both consumers and the out-of-home channel) to cook the products in a proper way. Time and temperature instructions should be clearly stated on the package, preferably at somewhat lower temperatures. Instructions should also consider the volume of product to be cooked. For frying the ratio of oil-to-product should be clear. For oven cooking, the electrical power-to-product ratio should be indicated. Secondly, we found that power of household equipment (ovens and deep fryers) varies and should be taken into account as well. Producers of consumer appliances could improve their product by using better time and temperature controls. Most important perhaps is that many consumers overcook French fries, thereby increasing acrylamide levels substantially, simply because of specific liking (darker and dryer end product) or routine behaviour or sometimes negligence.
Domestic fryer. (Photo: University of Bologna)
Page 19 of 77
2.2.2. Bread Studies have shown that asparagine is the limiting factor for formation of acrylamide in bread, and that asparagine is mainly concentrated in cereal bran. This means that high fibre flour will result in higher acrylamide content than sifted flour. New fermentation techniques could help in break down of asparagine and thus lowering acrylamide. The effects of asparagine and fructose, added to the dough, were studied in an experiment with a full factorial design. More than 99% of the acrylamide was found in the crust. Added asparagine dramatically increased the content of acrylamide in crusts dry matter (from about 80 µg/Kg to between 600 and 6000 µg/Kg) while added fructose did not influence this content. Both time and temperature increased the acrylamide content in crust dry matter, and a significant interaction was found between these two factors. When baked at different conditions with the same ingredients, a highly significant relationship between colour and acrylamide content was found. Added asparagine increased acrylamide content in yeast-leavened breads while added glycine decreased the content. The more asparagine in the dough, the stronger was the reducing effect of glycine. When glycine was applied on the surface of the fermented dough, also a significant reduction of acrylamide content in the bread was found. Addition of glycine but not of asparagine caused an increased browning reaction during baking. Fermentation of doughs made with different milling fractions showed that most of the asparagine was used up after 2 hr of fermentation with bakers yeast. Sourdough, on the other hand, did not reduce the content of free asparagine as efficiently but had a strong negative impact on asparagine utilization by the yeast. This indicates that this type of fermentation may result in bread with higher acrylamide content than breads fermented with yeast only. Short fermentation time compared to longer fermentation reduced acrylamide content in bread made with whole grain wheat by 87%. For bread made with rye bran, the corresponding reduction was 77%. Baking time and temperature increased acrylamide content in rye crisp bread. A significant interaction between time and temperature of baking was found. Added asparagine had a significant effect on the formation of acrylamide but fructose had not. There was a correlation between acrylamide content and colour of the milled bread in the time-temperature experiment but this correlation was not observed in the experiment with added precursors. Added oat-bran concentrate with high content of mixed-linkage β-glucan did not influence the acrylamide content in the breads. 2.2.2.1. New baking techniques New baking techniques such as air jet impingement and infrared radiation baking has focussed on wheat based bread. Various baking technologies have been studied: Traditional baking; Steam baking; Jet air impingement baking, and Infrared radiation (IR) baking.
Baking by infrared radiation. (Photo: Swedish Institute for Food and Biotechnology)
Page 20 of 77
First the influence of baking parameters on acrylamide formation and bread characteristics at conventional baking was evaluated. We could show that the highest acrylamide levels normally were found in the outer part of the crust and that the levels increased with increased baking temperature (200-260 C) and prolonged baking time (10-25 min). However, at the highest temperature-time combination (260°C; 20 min) the acrylamide level decreased and was lower in the outer crust than in the inner crust. From this knowledge base, we worked along two lines: the application of steam to traditional baking and alternative heating techniques, such as infrared radiation and air jet impingement. Steam baking We studied the effects of using steam during the final part of baking. The reference baking parameters were 200 C baking temperature for 20 minutes, and this was compared to baking when steam was introduced after 5, 10, or 15 minutes of baking. The steam also lowered the temperature of the bread, so for comparison baking experiments were performed where similar temperature profiles were obtained by temperature adjustments in the oven. Both the steam baking and the “falling temperature” baking substantially lowered the acrylamide levels in the final bread crust. However the crust colour was more influenced in the falling temperature breads than in the steam baked breads. With steam baking it was possible to find conditions that gave bread of the same colour level as that achieved by traditional baking, but with considerably lower acrylamide levels. With falling temperature baking, however, though acrylamide levels were lowered, the bread colour was lighter. Using sensory analysis, we could show that it was possible to bake bread (using steam the final 5 minutes) that was not significantly different from the control regarding odour, appearance, texture, and flavour, but having 40% lower acrylamide level. Alternative baking techniques The effect of new baking techniques, such as air jet impingement and infrared radiation baking, on bread crust formation and characteristics was initially studied. From the knowledge of such experiments baking conditions were defined to reach similar crust colour with the various baking techniques. It was obvious that the acrylamide content in the bread crust could be reduced by using alternative baking technologies. Reduced levels were obtained with infrared radiation baking as well as with air jet impingement baking. Preliminary experiments were also performed to study the flavour formation in bread baked by these new technologies. It turned out to be a close relationship between a large number of volatiles and the formation of acrylamide for the baking methods studied, especially those volatile and odorous compounds that are known to be formed via browning reactions in the bread crust, e.g. Strecker aldehydes and alkyl pyrazines. Obviously, a baking method that resulted in low acrylamide content in these experiments also tended to result in low contents of important flavour substances. We then further focussed on IR baking to evaluate how the IR baking can be optimized with regard to acrylamide minimisation and sensory product quality. Two different IR wavelength regions (one in the shorter range, IRS; and one in the medium range, IRM) and two effect levels (100% and 50%) were compared, using traditional baking as a reference. Flat bread cakes were used for the study. We followed the development of crust colour, acrylamide, flavour compounds and sensory characteristics with baking time at the various baking conditions. One important finding from these experiments was that with IR baking it is possible to obtain a sensory profile almost identical to that of the conventionally baked bread, but with considerably lower acrylamide content. A reduction of the acrylamide content by 60% could be shown. The main conclusion is that it is possible to reduce the acrylamide content while retaining the sensory quality both by introducing steam in traditional baking and by using new alternative baking techniques. In this work it was possible to reduce the acrylamide content by 40% in white bread, by applying steam during the final five minutes of baking, with no significant changes of the sensory quality. Using infrared radiation heating it was possible to reduce the acrylamide level in flat bread cakes by 60% with retained sensory properties.
Page 21 of 77
2. Analysis
Graz University of Technology (Austria), Central Science Laboratory, York (United Kingdom), Queens’ University Belfast (United Kingdom), TÜBÝTAK-Marmara Research Center (Turkey),
Swedish Food Administration (Sweden), Universitat de Barcelona (Spain), Institute of Chemical Technology Prague (Czech Republic), Universitat de Barcelona (Spain), National Food Institute (Denmark),
Norwegian Institute of Public Health (Norway), Stockholm University (Sweden), Lund University (Sweden)
3.1. Method development Several experiments have been performed with the aim to optimize and validate analytical methods for heat-induced toxicants in different types of foods, of high accuracy and precision and suitable for use by producers and enforcement laboratories. Methodology for acrylamide analysis is now mature. The available LC-MS and GC-MS methods are capable of testing all the food types of interest down to the concentrations of interest. Accuracy is acceptable although precision could be improved. Method development and validation has been assisted by an understanding of the potential interferences in particular food types, and how to avoid them. To provide analytical data for intake estimates, the detection limit of the bromination-GC-MS method was improved to give a low (single-figure) µg/kg limit of detection. The improved method was applied to the analysis of UK total diet samples. The estimated dietary exposure to acrylamide was within, and at the low end of, estimates made in other countries. This Br-GC-MS method was found to be more variable than wanted in between-lab trials and an LC-MS/MS method was developed and validated. The development work included systematic studies on extraction conditions, optimised extract clean-up by SPE, and chromatographic column separation. The method was subjected to inter-laboratory validation under the auspice of JRC-irmm. Repeatability (r) figures were 4-9% with potato crisp samples and 3-8% with bakery products. The reproducibility (R) values were 9-13% and 5-12%, respectively. For evaluation of trueness, the study included a spiked potato powder (500 µg/kg) and a candidate reference material from crisp bread (980±90 µg/kg). The mean results for these were 485 µg/kg (RSDR=9%) and 950 µg/kg (RSDR=5%), respectively, indicating that the method bias is low or nonexistent. A variation of the method was developed for testing low acrylamide levels in baby food. The multimode step was omitted and proteins were precipitated by adding KAl(SO4)2 prior to SPE on ENV+. An LOQ of 0.5 µg/L was obtained. A new GC/HRTOF-MS analytical procedure enabling direct determination of acrylamide (no derivatisation) in various food matrices was validated. Also, a new LC-MS/MS method employing a novel sample preparation strategy was implemented. For quantification, either D3-acrylamide (GC/HRTOF-MS) or 13C3-acrylamide (LC-MS/MS) are employed. The accuracy of these methods is checked regularly via proficiency test scheme FAPAS®. Satisfactory z-scores were obtained: -0.3 (Baby Rusk Test Material, series 30 Round 9); 0.2 (Potato Crisps; 30:11), 0.4 (Breakfast Cereals; 30:14) and 1.5 (Crisp bread; 30:15). Performance characteristics are as follows:
Page 22 of 77
GC/HRTOF-MS LC-MS/MS LoD = 10-30 µg/kg, depending on matrix repeatability (RSD) = 2% (chips, ca 500 µg/kg)
LoD = 10-25 µg/kg depending on matrix repeatability (RSD) = 3% (chips, ca. 700 µg/kg)
Figure 5: SPME-GC-ITD/MS and SPME-GCxGC-TOF/MS methods for determination of heat-generated volatile compounds.
A method for the analysis of acrylamide by LC-MS/MS using an ion trap system with atmospheric pressure chemical ionization (APCI) was developed. A micro-emulsion electro-kinetic chromatography (MEEKC) method has been established suitable for acrylamide analysis at high levels in certain food types. Besides the direct analysis of acrylamide using LC-MS, a derivatisation method using 2-mercaptobenzoic acid was adopted that results in a substance with a higher molecular weight, which is more selective than the low molecular weight of acrylamide itself at 72. This method was used for the analysis of acrylamide in coffee and in model systems. The same derivatisation with mercaptobenzoic acid was used prior to a CZE analysis using FASI (field-amplified electrokinetic sample injection) as a pre-concentration on-line technique to increase sensitivity. Additionally a method for an intermediate compound was developed namely 3-amino-propionamide. This substance could be identified in a model system as an intermediate but it was not found during the roasting of coffee. A method for the determination of HMF (hydroxymethylfurfural) by GC-ITMS using N,O-bis(trimethylsylil) trifluoroacetamide as derivatising reagent has been developed and also an LC-MS method using an ion trap system with ESI as ionisation source. A method for the analysis of a metabolite HMFA (5-hydroxymethyl-furanoic acid) was developed. This metabolite is excreted via the kidney to the urine. The method was applied to the analysis of urine to estimate the exposure. The results suggest that the exposure is higher than calculated from the exposure data (food analysis). Therefore, it is necessary to analyse other foods and identify any additional source(s) of exposure. As part of method development for various heat-induced toxicants, different extraction techniques including solid phase extraction (SPE), solid phase micro-extraction (SPME) and pressurized solvent extraction have been explored to good effect. In response to the report on "masked" acrylamide, the pilot study of its formation in model system was initiated and possible precursors of “masked” acrylamide were identified.
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
Akrylamidm/z 71,036Hmotnostní okno: 0,02 Da
D3-m/z 74,056
í okno: 0,02 Da
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
m/z 71,036Hmotnostní okno: 0,02 Da
D3-Am/z 74,056
í Da
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
m/z 71,036Hmotnostní okno: 0,02 Da
D3-m/z 74,056
í Da
TIC
Acrylamide
m/z 71.037
Mass window: 0.02 Da
-
m/z 74.056
Mass window: 0.02 Da
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
/
D3-/z 74,056
í Da
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
D -/z 74,056
TIC
Acrylamide
m/z: 71.036
Mass winow: 0.02 Da
D3-Acrylamide
m/z: 74.056
Mass window: 0.02 Da
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
Akrylamidm/z 71,036Hmotnostní okno: 0,02 Da
D3-m/z 74,056
í okno: 0,02 Da
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
m/z 71,036Hmotnostní okno: 0,02 Da
D3-Am/z 74,056
í Da
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
m/z 71,036Hmotnostní okno: 0,02 Da
D3-m/z 74,056
í Da
TIC
Acrylamide
m/z 71.037
Mass window: 0.02 Da
-
m/z 74.056
Mass window: 0.02 Da
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
/
D3-/z 74,056
í Da
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
D -/z 74,056
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e3
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e3
7.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e5
7.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
7.336.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
Akrylamidm/z 71,036Hmotnostní okno: 0,02 Da
D3-m/z 74,056
í okno: 0,02 Da
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
% 8.21
8.448.37
Akrylamidm/z 71,036Hmotnostní okno: 0,02 Da
D3-m/z 74,056
í okno: 0,02 Da
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e3
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e3
7.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e5
7.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05
7.336.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
m/z 71,036Hmotnostní okno: 0,02 Da
D3-Am/z 74,056
í Da
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time
6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
m/z 71,036Hmotnostní okno: 0,02 Da
D3-Am/z 74,056
í Da
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e3
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.49
7.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e3
6.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e5
7.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time
7.336.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e3
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e3
7.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e5
7.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
m/z 71,036Hmotnostní okno: 0,02 Da
D3-m/z 74,056
í Da
TIC
Acrylamide
m/z 71.037
Mass window: 0.02 Da
-
m/z 74.056
Mass window: 0.02 Da
7.336.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
m/z 71,036Hmotnostní okno: 0,02 Da
D3-m/z 74,056
í Da
TIC
Acrylamide
m/z 71.037
Mass window: 0.02 Da
-
m/z 74.056
Mass window: 0.02 Da
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e3
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e3
7.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e5
7.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
/
D3-/z 74,056
í Da
TIC
7.336.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
/
D3-/z 74,056
í Da
TIC
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e3
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e3
7.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e5
7.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
7.336.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
6.20 6.40 6.60 6.80 7.00 7.20 7.40 7.60 7.80 8.00 8.20 8.40Time0
100
%
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e3
0
100
%
0
100
%
2: TOF MS EI+ 71.036 0.02Da
9.91e37.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e3
7.12
6.20 6.496.26 6.667.28 7.687.73 8.04 8.338.29
2: TOF MS EI+ 74.056 0.02Da
5.25e37.12
2: TOF MS EI+ TIC
2.05e5
7.12
2: TOF MS EI+ TIC
2.05e57.33
6.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
D -/z 74,056
TIC
7.336.65
6.23
6.05 6.16
6.50
6.32
6.546.83
6.726.78
6.87
7.257.00
6.97 7.12
7.43
7.37
7.50 7.66
7.547.60
8.168.09
7.72 8.027.857.79 7.98
8.21
8.448.37
D -/z 74,056
TIC
Acrylamide
m/z: 71.036
Mass winow: 0.02 Da
D3-Acrylamide
m/z: 74.056
Mass window: 0.02 Da
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00 5.50 6.00Time0
100
%
0
100
%
crisps MRM of 4 Channels ES+ 75.05 > 58.35
2.41e52.66
crisps MRM of 4 Channels ES+ 72.05 > 55.35
1.27e52.66
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00 5.50 6.00Time0
100
%
0.50 1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00 5.50 6.00Time0
100
%
0
100
%
crisps MRM of 4 Channels ES+ 75.05 > 58.35
2.41e5
0
100
%
crisps MRM of 4 Channels ES+ 75.05 > 58.35
2.41e52.66
crisps MRM of 4 Channels ES+ 72.05 > 55.35
1.27e5
2.66
crisps MRM of 4 Channels ES+ 72.05 > 55.35
1.27e52.66
13C3 – Acrylamidetransition m/z 75 → 58level: 1000 µg/kg
Acrylamidetransition m/z 72 → 55level: 650 µg/kg
13C3 – Acrylamidetransition m/z 75 → 58level: 1000 µg/kg
Acrylamidetransition m/z 72 → 55level: 650 µg/kg
GC/HRTOF-MS LC/MS-MS
Page 23 of 77
3.2. Analytical results on heat-induced toxicants levels in foods The acrylamide levels in breast milk and Swedish baby food products were tested. The average levels for gruel, porridge and canned baby food, all ready to eat, were 1.4, 26, and 7.8 µg kg-1 respectively. For all breast milk samples except one, acrylamide was below the limit of quantification (0.5 µg kg-1). The mean intake between seven and twelve months of age was estimated to be about 0.5 µg/kg body weight/day. The acrylamide level in toasted bread was investigated in order to aid the estimation of intake from home-prepared foods. The amounts of acrylamide produced in the different breads by toasting were strongly related to the concentrations before toasting. There was hardly any acrylamide increase in the light toasted breads. The levels for medium toasting were about 2 to 5-times higher than the levels before toasting.
Table 3. Acrylamide concentrations in toasted bread (corrected for weight loss).
Weight loss Acrylamide Weight loss Acrylamide Weight loss Acrylamide Weight loss Acrylamide (%) (µg/kg) (%) (µg/kg) (%) (µg/kg) (%) (µg/kg)
no toasting 0 3,0 0 8,4 0 28,8 0 41,6
1 11 3,9 10 8,7 11 29,7 11 43,9
2 14 4,2 12 9,6 12 28,3 13 60,5
3 17 15,9 15 21,6 14 57,7 15 84,6
4 20 31,0 19 57,8 18 113,5 17 118,0
wheat Toasting
time
wheat wheat + rye wheat + rye
Analysis of acrylamide in foodstuffs (n=98) from Spanish markets and homemade processed foods has been performed. Similarly, a total of 227 data were collected for acrylamide contents in foods from the Turkish market, and 40 data for furan content. The survey for acrylamide included traditional Turkish foods and home-cooked foods. Moreover, French fries collected from fast food restaurants and French fries that were prepared by using frozen potato were also analysed. The French fries from fast food restaurants had the higher acrylamide content. Compared to other food groups, the traditional Turkish foods and bakery products contained lower levels of acrylamide. The survey was performed over two years. The same brands of products were monitored during this period. Although there is not enough data for proving it statistically, there were indications that acrylamide levels in some specific products, including baby biscuits, toasted bread and potato chips, decreased during this period. Typical results are given in Table 4.
Table 4. Data for acrylamide content in some food products sold in Turkey. Examples of Foods Acrylamide
(µg/kg) Examples of Foods Acrylamide
(µg/kg) Roasted almond 207-313 Fried vegetables (eggplant, pepper,
squash) <10
Pop corn 170 Pickled olives <10-139 Baby biscuits 36-613 Corn bread <10 Corn chips 198-835 Turkish coffee 227-300 Potato chips 59-2336 Instant coffee* 336 Bread (wheat) <10-79 Fried potato – home made 60-66 Toasted bread 58-205 Fried potato – frozen potato 72-76 Pilav (ready to eat rice) <10 Fried potato – fast food 375-428
Page 24 of 77
The carcinogenic substance furan ’emerged’ as a heat-induced toxicant in foods during the planning and conduct of the HEATOX project. The issue originated from work by the US-FDA. Our studies made an important contribution in helping to understand aspects of the analysis of foods for furan – crucially the need to avoid strong heating during headspace analysis to avoid misleadingly high results. This provided acceptance criteria and so gave confidence in the analytical data on furan occurrence and concentrations that is being complied by EFSA. The headspace GC-MS method of testing for furan is straightforward and accurate. It can be used both by producers and official laboratories. Studies were then conducted to understand the chemical precursors, the mechanisms of formation, the nature of the foods affected by formation of furan, and the effect of normal consumer practice of heating and cooking before consumption. This work confirmed that there are different precursors and mechanisms of formation. On other furan compounds, HMF (hydroxymethylfurfural) was found at high levels in foods containing high amounts of carbohydrates that are stored for a long time or heated strongly. These included dried fruits, coffee, Madeira wine, caramel, balsamic vinegar, non alcoholic and alcoholic beverages. Comprehensive databases on (i) the reaction products from Maillard browning, and (ii) lipid oxidation products, were set-up and populated with occurrence data and with hazard indications derived from QSAR (quantitative structure-activity relationship) considerations. Similarly, a database for specified heat-induced toxicants (acrylamide, ethylcarbamate, furan, heterocyclic amines, hydroxymethylfurfural, polycyclic aromatic hydrocarbons and nitrosoamines) has been designed. Concentration levels of heat-induced toxicants in cooked food samples have been introduced in the database. Sample food and cooking data together with analytical information (sample treatment, identification and determination details) have been included in the database. As an example, in Table 5, there is the information about acrylamide in baby food products Table 5. Baby foods
Nº Samples
Nº NDSamples MIN MEDIAN MAX Units Determination
TechniqueExtraction and
clea-up procedures Ref.
Breast milk 4 0 0,25 0 0,65 µg/kg LC-MS/MS Precipitation of macromolecules (KAl (SO4)2)
49
Cereals 12 4 1 10 24 µg/kg LC-MS/MS S-L (H2O), L-L (AcOEt), SPE 49,80,125Infant formulas 67 64 - 10 - µg/kg LC-MS/MS S-L (H2O:MeOH 95:5), SPE 123,125Purées (Fruits, potatoes, vegetables, meat)
22 3 1 22 121 µg/kg LC-MS/MS S-L (H2O), SPE 49,125
Gruels and porridges 10 0 0,25 2 34 µg/kg LC-MS/MS S-L (H2O) 49Others (Biscuits, cakes, crackers, toasts)
10 0 19,9 118 1568,9 µg/kg LC-MS, LC-MS/MS S-L (H2O), L-L (AcOEt), Carrez, SPE
27,70,80,125
Page 25 of 77
3.3. Analytical method for biomarkers
Figure 6. Biomarkers of AA for determination of short-term and long-term exposure
(Thomas Bjellaas - Doctoral Thesis)…
3.3.1. Identification of DNA-adducts in vivo An analytical method including the use of LC/MS has been developed for the detection of genotoxic compounds in a rodent bioassay. 2-methylfuran was detected as a possible genotoxic Maillard reaction product in this bioassay. More unknown genotoxins may be identified using this procedure. Mixtures of single amino acids and carbohydrates were suspended in phosphate buffer and heated to various temperatures for the production of Maillard reaction products as visualized by the colour formation. The Maillard reaction has been upscaled in order to produce sufficient material for rat feeding studies and a method for a fairly homogeneous incorporation of the Maillard reaction product into rat feed pellets has been developed. A study of the palatability in rats showed that two grams of Maillard product in 15 grams of rat feed was well tolerated after a short adaptation period.
Page 26 of 77
Rats were given feed with reaction products from different amino acids and after 5 days DNA was isolated from different organs. The DNA was hydrolysed and analysed by HPLC/MS. Chromatographic data of DNA from dosed animals were compared with data from control animals and differences, suggesting the presence of DNA-adducts, were detected by use of ”metabolite detect” computer software.
Figure 7. Extracted ion chromatograms m/z=232 of hydrolyzed intestinal DNA (upper) and a synthesised standard (lower).
At least two possible DNA adducts with mass 232 were detected by HPLC/MS analysis of hydrolysed DNA extracted from rodents following feeding with Maillard reaction products (Figure 7, upper panel). Analysis of the adducts by high resolution TOF-MS gave exact masses of 232.08345 for both adducts which is in accordance with the molecular formula C10H10N5O2. The molecular formula suggested a guanine adduct to which C5H6O2 had been added followed by elimination of water. A possible candidate to one of the adducts 4-oxo-2-pentenal was synthesised by oxidation of the known Maillard reaction product, 2-methylfuran with m-chloro-perbenzoic acid. Reaction of 4-oxo-2-pentenal with deoxyguanosine gave a product which after acid hydrolysis had identical retention time and mass as the first eluting adduct found in rodent DNA (Figure 7, lower panel). 3.3.2. Acrylamide metabolites as biomarkers of exposure Exposure to acrylamide from food has been estimated using records of food consumption combined with knowledge on acrylamide content of various foods. However, these intake estimates needs to be validated against biological markers of exposure. The concentration of haemoglobin adducts of acrylamide reflects the internal dose of acrylamide during the last 3-4 months, while we had no markers for short term intake of acrylamide. The bioavailability of acrylamide from food is an important issue which was addressed in experiments on mice. In one study, 53 persons answered a food frequency questionnaire about the daily food intake and also had answered a 24 h food recall. Blood and 24h urine samples were collected. In another study, mice were kept in metabolic cages and were exposed to crisp bread with different concentrations of acrylamide, subcutaneous injections of corresponding doses of acrylamide, including 14C-labeled acrylamide.
0 9 - 0 1 1 1 3 . D : E I C 2 3 2 + A l l
0 . 0 0
0 . 2 5
0 . 5 0
0 . 7 5
1 . 0 0
1 . 2 5
7x 1 0I n t e n s .
2 . 5 5 . 0 7 . 5 1 0 . 0 1 2 . 5 1 5 . 0 1 7 . 5 2 0 . 0 T i m e [ m i n ]
0 9 - 0 1 0 1 0 . D : E I C 2 3 2 + A l l
0
2
4
6
6x 1 0
2 . 5 5 . 0 7 . 5 1 0 . 0 1 2 . 5 1 5 . 0 1 7 . 5 2 0 . 0 T i m e [ m i n ]
Page 27 of 77
Acrylamide and its oxidation product glycidamide are conjugated to glutathione and these conjugates are stepwise converted into N-mercapturic acids which are excreted into urine fairly rapidly. The mercapturic acid metabolites including 2H-labelled standards as well as an external standard were synthesised and characterised (Figure 8).
Figure 8. Structures of standards used in the study.
An analytical method of determination was developed based on solid-phase extraction of urine followed by LC-MS/MS separation and quantification (Figure 9). The method of analysis was applied in a pilot study of six persons. Fasting caused a 50 percent reduction in excretion of mercapturic acid metabolites clearly showing that food is an important source. Smoking was associated with a high excretion of mercapturic acids.
Figure 9. HPLC-MS/MS chromatogram of (A) a spiked urine sample at 40 µg/L for MA-AA and MA-GA3, (B) urinary metabolites from a non-smoker and (C) urinary metabolites from a smoker. Peak intensities are set relative to the spiked urinary sample. The grey shaded peak at 8.2 min might possibly be the sulfoxide analogue of MA-AA.
Page 28 of 77
It was shown that excretion of urinary mercapturic acid metabolites was related to food intake, the biomarker responded quickly to food with a high content of acrylamide, e.g. potato crisps (Figure 10).
Figure 10. The total amount of AA found as urinary metabolites in consecutive urine samples in a participant who reported intake of potato crisps during the evening.
On the assumption that 55% of the acrylamide dose is excreted as mercapturic acids, the average dose in the 53 persons was 30 µg/day. Excretion correlated with estimated recent intake, but not with the total estimated amount during the last 24 hrs. Multiple regression analysis showed that the intake of aspartic acid (including also asparagine), starch and coffee were significant predictors of AA metabolite excretion. The method of urinary excretion could also be used for the identification of particular foods contributing to acrylamide exposure. Acrylamide haemoglobin (Hb) adducts were determined in the same study subjects. The level decreased with age, possibly explained by a lower intake of foods with high acrylamide content, such as potato chips and snacks, in middle aged and older persons. A 3.5-fold higher adduct concentration was found in smokers in comparison with non-smokers. No correlation was found between Hb adduct levels and total dietary acrylamide intake estimated from reported food consumptions and levels of acrylamide in foods. After adjusting for age significant predictors for Hb adduct formation were intake of chips, snacks and crisp bread. Haemoglobin adduct concentrations did not correlate with urinary excretion of mercapturic acid derivatives in the same subjects.
Page 29 of 77
In mice exposed systemically to 14C-acrylamide, about 92% of the radioactivity was excreted into urine during the following 48 hrs. There were linear dose response curves both for acrylamide from crisp bread and systemically given (11). The bioavailability of AA from crisp bread was about 100%.
Figure 11. A) The linear relationship between the amount of acrylamide (AA) ingested from crisp bread and the amount of AA recovered as urinary metabolites. B) The percentage of dose recovered in urine for total and the different urinary biomarkers MA-AA, MA-GA3, MA-GA2 and GA
Page 30 of 77
The frequency of MNPCE after oral injection of acrylamide
0
0,2
0,4
0,6
0,8
1
1,2
1,4
0 10 20 30 40 50 60 70
dose, AA mg/kg b.w.
f MN
PCE
(pb,
‰)
3. Hazard characterisation Swedish Food Administration (Sweden), Lund University (Sweden), Stockholm University (Sweden), German Institute for Human Nutrition (Germany), Norwegian Institute of Public Health (Norway),
University of Zürich (Switzerland)
4.1. In-vivo genotoxicity and carcinogenicity In absence of data on the effects of a chemical on humans it is often necessary to extrapolate the effect from high doses given to animals to lower dose regions for exposed humans. Therefore when the effect of very low experimental doses can be determined the extrapolation is more reliable and makes the risk quantification more reliable. The work has been concentrated around the uptake and the effect in the low dose region of acrylamide and other chemicals produced when heating foodstuffs. To clarify the shape of the dose response curve in the low dose region the biomarkers for DNA adducts, haemoglobin adducts for acrylamide and glycidamide, and the micronucleus assay (two-lazer-beam flow cytometer) were used. The mice (35) were given by gavages 14 different doses (control included) of acrylamide. The doses were ranged from 0 to 60.0 mg/kg body weight. Data from the measurement of haemoglobin adducts, Figure 12, and from the micronucleus (MN) assay, Figure 13, are given below. It is clearly shown that there is a dose related increase of the frequency of MN (fMNPCE). The fMNPCE levels for the different mice are about the same as after intraperitoneal injection. The dose response is linear. Also the analysis of haemoglobin adducts showed a linear dose related increase. However, the lowest level of the quantification of haemoglobin adducts was about 1 mg/kg.
Figure 12. Measurement of hemoglobine adducts Figure 13. Micronucleus assay
Results from studies using accelerated mass spectrometry (AMS) show that glycidamide binds to DNA in mice at a dose of about 1.5 µg/kg body weight. In a human study where twenty-four healthy persons eating either a) boiled (max 100 ºC) and fresh food or b) strongly heated (fried) and fresh food, there was a significant difference between the groups regarding the frequency of micronucleated young erythrocytes. From each of the subjects blood samples were drawn twice, before and after an “eating period” of four days. Blood samples were analysed for both the frequency of micronucleus (MN) in young erythrocytes and the haemoglobin adduct levels of acrylamide and glycidamide.
0
5
10
15
20
25
30
35
0 10 20 30 40 50 60 70
Dose, AA mg/kg b.w.
Add
uct l
evel
(nm
ol/g
)
AA
GA
Page 31 of 77
-1
-0,8
-0,6
-0,4
-0,2
0
0,2
0,4
0,6
0,8
fMN
(‰),
diffe
renc
e
a
b
The estimated intake of acrylamide in the two groups was about 25 µg and 3000 µg, respectively. This difference in calculated acrylamide intake was verified by a significant difference (p<0.001) in the level of haemoglobin adducts between the two groups. After the eating period the difference in the mean fMN for the persons belonging to the high-heated-food group (b) was +0.17‰. The difference for the persons belonging to the low-heated-food group (a) was –0.15‰. These two groups are significant separated from each other, p<0.005 (one tailed, paired student t-test), 14.
Figure 14. frequency of micronucleated young erythrocytes in twenty-four healthy persons eating either a) boiled (max 100ºC) and fresh food or b) strongly heated (fried) and fresh food …
There are different possibilities to increase the sensitivity of the micronucleus assay in human erythrocytes; one way is to e.g. discriminate between clastogenic and aneugenic effects. This can be done by a fluorescent in situ hybridisation (FISH) of the DNA in the micronuclei. Another way (without FISH) is to optimize the fixation method, in order to better define the cell population (very young erythrocytes) when analysing the cells in the flow cytometer. This latter option was used as a new refined method when the human study was done. Also other chemicals than acrylamide have been studied upon the potency to increase the frequency of micronuclei, for example furan, metylfuran, HMF, its metabolite SMF, phytosterol oxides, metylacrylamide and also extracts of potato chips. Among these chemicals only SMF showed a positive effect in the in vivo micronucleus assay in mice. A comparison of the genotoxicity of acrylamide and three heterocyclic amines (PhIP, IQ, and MeIQx) was made. In the case of PhIP, we found a significant dose-response relationship, while MeIQx and IQ did not give an increased fMPCE level. A comparison between the heterocyclic amines and acrylamide revealed that the slope of the dose-response curve is about ten times steeper for PhIP than acrylamide. In spite of this, acrylamide probably constitutes a higher human risk than heterocyclic amines since the intake of acrylamide is about a hundred to a thousand fold higher. In conclusion, the results points to an increased risk already at the lowest doses. In the human study the acrylamide dose was about a thousand times lower than the lowest effective doses in earlier studies in mice. It is likely that the explanation of the increase of chromosome instability (micronuclei) is something else than acrylamide and it is possible that the high heating temperature of foodstuffs is responsible for the production of chemicals other than acrylamide inducing a higher frequency of micronuclei.
Page 32 of 77
4.1.1. Data on the relationship between internal dose and estimated intake of acrylamide in blood samples from a biobank Associations between estimated acrylamide-intake, based on self-reported data on usual food consumption combined with data on published acrylamide content in foods, and Hb adducts were examined with linear regression and correlation analysis. A sample from the Malmö Diet and Cancer cohort was selected to obtain a large variation in acrylamide-intake. The estimated acrylamide intakes ranged from 3.5 to 190 µg/day, with a median of 45 µg/day. In randomly selected individuals the estimated intake was 28 µg/day. The estimated intake thus varied with a factor of ca 50. In contrast to this, the internal doses as measured by acrylamide adduct levels in Hb varied with a factor of 5 in the non-smoking group (see 15). Hb adduct levels are expected to give a more exact estimate of the intake than the estimates from questionnaires. Positive partial correlations between dietary acrylamide-estimates and Hb adducts were seen in non-smoking men but not in non-smoking women. In conclusions, two results from this study cast doubt on the validity of dietary intake estimates used in cancer epidemiology: a) The lack of correlations between adduct levels and intake estimates in non-smoking women, b) The much lower variation in internal doses compared to estimated intakes from food frequency questionnaires.
Non-smokers
0,00
0,02
0,04
0,06
0,08
0,10
0 50 100 150 200
Estim. AA-intake (µg /day)
AA
-add
uct l
evel
(nm
ol/g
)
Figure 15. Relation between estimated dietary AA-intakes from foods (µg/day) and Hb-adducts from acrylamide (nmol/g) in non-smoking men and women in a sub-sample from the Malmö Diet and Cancer cohort. Filled circles represent men and unfilled circles represent women.
We studied some non-acrylamide compounds that form when carbohydrates are heated, in particular 5-hydroxymethylfurfural (HMF) and related compounds. These compounds are ingested at extremely high levels (1000 times higher than acrylamide) and it has been reported that rodent sulfotransferases (SULTs) can covert HMF in vitro to a mutagen, 5-sulphooxymethylfurfural (SMF). We wanted to find out whether human SULTs can mediate this reaction in vitro and in vivo and to study resulting adverse effects . The objectives included the development of biomarkers for this activation pathway in animals and humans. Our results may be summarised as follows: A major human SULT form (SULT1A1) was very efficient in converting HMF to SMF. In rat and mouse, only minor SULT forms showed high sulfation activity towards HMF. Therefore, mice and rats may be poor models for humans when studying SULT-mediated toxicity of HMF and congeners. SMF was identified as a urinary HMF metabolite in a mouse line engineered for expression of human SULT1A1, but not in wild-type mice, corroborating that human SULT1A1 may be a risk factor in exposures to HMF.
Page 33 of 77
SMF, unlike HMF, formed DNA adducts in cell-free systems, was mutagenic in conventional bacterial and mammalian cell test systems, and was strongly nephrotoxic in mice (Figure 16). HMF and other hydroxymethylfuran congeners present in foods were mutagenic in bacteria and mammalian cells engineered for expression of human SULT1A1. Whereas HMF was only mutagenic when SULT1A1 was expressed, some analogues were also mutagenic in control V79 cells (but not in control bacteria) suggesting the existence of additional bioactivation mechanisms in mammalian cells (e.g. mediated by unidentified kinases). Externally added furfuryl sulphate (unlike SMF) was only cytotoxic, but not mutagenic, to bacterial and mammalian cells. However, when furfuryl sulphate was generated by SULT from its alcohol within target cells, it was strongly mutagenic. This difference in biological activity between internal and external furfuryl sulphate might be due to its charge and short half-life time preventing permeation of the cell membranes. Adenine was identified as the major nucleobase forming DNA adducts with SMF in cell-free systems. This and other adducts were not detected in target cells and tissues of SMF despite increased mutant frequencies and massive histopathological alterations. Unfortunately we did not succeed developing sufficiently powerful enrichment methods for adducts, as required for their demonstration in cells and tissues. Moreover, the presence of the carbonyl group in 2-postion of HMF/SMF, expected to be still present in the adducts, complicated the analysis, as it might form cross-links with DNA and/or other cellular constituents. We have demonstrated that major human SULTs, unlike major rat and mouse SULTs, efficiently convert HMF to SMF, and that SMF forms DNA adducts in cell-free systems, is mutagenic in bacterial and mammalian cells in vitro, and is strongly nephrotoxic in mice. The negative result of HMF in conventional in vitro systems is due to a inappropriate metabolic situation, as HMF is positive in the same tests after expression of human SULT1A1 at levels occurring in human tissues. Likewise, the low biological activity of HMF in laboratory animals should be regarded with caution, as these models do not truly reflect the human sulfation capacity. A similar situation was found with other hydroxymethylfuran congeners present at high levels in food, such as furfuryl alcohol.
Figure 16. Kidney of an SMF-treated mouse. Note that the proximal tubules were completely destroyed, whereas the glomeruli were damaged much less. The right panel shows abundant proteinaceous casts.
50 50
Page 34 of 77
4.2. Tumorigenicity Two experiments on intestinal tumourigenesis of acrylamide and glycidamide in Min/+ mice and their wt litter mates have been completed. In the first experiment the animals were exposed to acrylamide or glycidamide (10 or 50 mg/kg bw) week 1 and 2 after birth, and in the second experiment they were exposed to acrylamide or glycidamide one week before birth to the dam, alone or in combination with exposure of the pups at week 1 and 2 after birth (50 mg/kg bw). Min/+ mice and wild type mice have been treated postnatal with subcutaneous injections of 5-hydroxy-methyl-furfural (HMF) or its reactive metabolite 5-sulfoxy-methyl-furfural (SMF). Eight weeks old female Min/+ and wt mice were exposed to glycidamide at the dose 50 mg/kg dissolved in 0.9% NaCl or to NaCl-solution alone for 6 and 24 hours. RNA was isolated from the epithelium and cDNA was made by in vitro transcription of total RNA, and Affymetrix GeneChipswere used for the expression analysis. A weak tumourigenic activity in the intestine of Min/+ mice and their wt litter mates was observed after exposure to acrylamide or glycidamide. The weak tumourigenicity observed was probably related to the perinatal exposure since such effects never previously have been observed in rodents exposed later in life. SMF significantly increased the number of adenomas in the small intestine and the number ACF in the colon of Min/+ mice. HMF did also show a tendency to increase the number of lesions. Gene expression profiles showed a high degree of variability and only one gene was significantly up-regulated by glycidamide in wt mice exposed for 6 h (metallothionein-2, 3.27-fold, adjusted p<0.05). No other genes were significantly regulated in each of the treatment. We investigated differences in gene expression between 6 and 24 hours, expecting different expression of genes involved in circadian exercises. We had from 75 to 1790 significantly regulated genes (adjusted p < 0.05) in each of the four data sets, and indeed, genes involved in circadian rhythms came out as a major functional cluster. 4.2.1. Reproductive toxicity Isolated mouse testicular cells and lymphocytes (primary cells) from BigBlueTM C57BL/6 male mice, either mOgg1+/+ or mOgg1-/- (a transgenic mouse strain deficient in oxidated lesion repair) were used. Peripherial blood lymphocytes from humans were also used. The alkaline single cell gel electrophoresis (comet assay) has been used to measure levels of DNA lesions as single strand breaks and alkali labile sites (baseless sites) after in vitro exposure of acrylamide (AA) and glycidamide (GA) in mouse and human cells (see above). To further study and characterise the DNA lesions formed, a modified comet assay was used. The modified assay includes a treatment step in which cellular DNA is subjected to an endogenous DNA repair enzyme that recognises and cleaves certain DNA lesions thereby converting them to single strand breaks, so that they are measurable in the comet assay. The CYP2E1 AA metabolite glycidamide was shown to be genotoxic in mouse testicular cells and lymphocytes in vitro, at concentrations similar to those producing genotoxicity in human lymphocytes. The parent compound AA was as expected very inactive. The DNA lesions induced by GA exposure are recognised by repair enzyme which dramatically increases the lesion levels, implicating that human testicular cells might be particularly susceptible to the active metabolite of acrylamide.
Page 35 of 77
4.2.2. Neurotoxicity Neonatal mice were exposed to different low doses of acrylamide at an age of 10 days. This study indicates that neonatal exposure to acrylamide causes developmental neurotoxic effects that are persistent and also get worse with age. These disturbances are both dose-response and time-response related. The spontaneous motor behavior data showed a disruption of habituation in animals exposed to acrylamide. Habituation, here defined as a decrease in the locomotion, rearing, and total activity variables in response to the diminishing novelty of the test chamber over a 60 min. period, was displayed in the control animals, but the animals exposed to acrylamide were clearly hypoactive during the beginning of the 60 min. period, while towards the end of the test period they were hyperactive. Furthermore, the lowest dose not causing any developmental neurotoxic effects (NOEL) on spontaneous behaviour is 0.1 mg acrylamide/kg body weight. Acrylamide is tumorigenic at high doses in rodents and has been classified as a probable human carcinogen. However, cancer risk projections in the general population remain problematic because the molecular pathogenesis of acrylamide at the low level of dietary uptake is not understood. In particular, the question of how non-genotoxic (epigenetic) responses such as changes of gene expression may modulate the known genotoxicity of acrylamide has never been examined. To close this gap of knowledge, we used high-density DNA microarrays and real-time PCR validations to determine genome-wide transcriptional profiles induced by acrylamide and glycidamide, the oxidative metabolite of acrylamide. Human MCF7 cells (derived from the mammary epithelium) and human CaCo-2 cells (derived from the intestinal epithelium) were exposed to acrylamide or glycidamide. After 24-h exposures, the cells were collected, RNA was extracted and processed for genome-wide expression profiling on Affymetrix microarrays that display the sequences of 47’000 human transcripts. Glycidamide induced gene expression changes of much higher amplitude than acrylamide itself. We identified > 100 human transcripts, coding for proteins with known biological functions that are significantly up- or down-regulated by exposure to glycidamide. The expression profiles resulting from glycidamide treatment are characterized by the dose-dependent up-regulation of cytoprotective factors including the glutathione system, enzymes that reduce toxic carbonyl compounds as well as diverse antioxidants. Low-dose experiments indicate that the transcript coding for epoxide hydrolase-1 represents the most sensitive biomarker for glycidamide exposure. At higher concentrations, glycidamide induces typical markers of tumour progression such as steroid hormone activators, positive regulators of nuclear factor-kB, growth stimulators and apoptosis inhibitors. Concomitantly, transcripts coding for growth suppressors and cell adhesion molecules are down-regulated. Many of these transcriptional changes have been validated by quantitative real-time reverse-transcriptase PCR assays in both MCF7 and CaCo-2 cells. In the case of epoxide hydrolase-1, a significant transcriptional induction was observed at a glycidamide concentration of as low as 0.1 µM. This is a glycidamide level that may be reached transiently in the blood or tissue of humans after dietary uptake of acrylamide. The micromolar doses of glycidamide that induce many other transcriptional responses might seem too high to be relevant for human exposure. However, it cannot be excluded that the sensitivity of target cells depends on the capacity of their detoxification systems and, considering that the uptake of acrylamide is normally accompanied by other toxic food constituents that may saturate these cytoprotective systems, adverse effects may occur at lower concentrations than those expected from threshold experiments performed with a single compound. This functional genomic study revealed that acrylamide and, in particular, glycidamide exert a plethora of transcriptional effects that should be included in risk assessment studies. We propose to implement a threshold level that prevents the induction of potentially hazardous transcriptional responses mediated by acrylamide or glycidamide in target tissues such as the mammary gland or the intestinal epithelium.
Page 36 of 77
The results of the in vitro study suggest that low-dose responses, primarily the induction of epoxide hydrolase-1, exert beneficial effects by promoting inactivation of the toxicant. Other detoxification processes observed at the low-dose glycidamide level of 0.1 µM include its inactivation through conjugation with glutathione (see Figure 17). Adverse effects, such as for example the down-regulation of cell-adhesion molecules (CDH12, ANXA1, ITGB6), the up-regulation of factors that lead to dysregulation of steroid hormone synthesis (AKR1C2, AKR1C3), or factors that promote cell survival and cell cycle progression (NF-kB, RRM2, Cyclin-1, MAPK12) are detected at concentrations above the range that appears to be relevant for human populations. An induction of the DNA repair enzyme EME1, indicative of the formation of DNA damage, is also observed in the upper level of the tested dose range. Thus, the main implication of these findings for risk assessment is that transcriptional signatures associated with DNA damage or tumour cell progression may be expected only at doses that exceed the range of ordinary dietary exposure.
Figure 17. Summary of the main findings from the transcriptomic profiling of glycidamide effects in human cells. Beneficial responses are observed at the low-dose level, potentially hazardous effects are detected at high glycidamide concentrations. Abbreviations: EPXH1, epoxide hydrolase-1; GSH, glutathione; FHO3, formin homology-3; CDH12, cadherin-12; NF-kB, nuclear factor-kB; RRM2, ribonucleotide reductase polypeptide-M2; AKR1C2, aldo-keto reductase-C2; AKR1C3, aldo-keto reductase-C3; EME1, essential meiotic endonuclease-1; ANXA1, annexin-A1; ITGB6, integrin beta-6; MAPK12, mitogen-activated protein kinase-12.
Consequences for risk assessment: these findings indicate that the standard procedure of extrapolating from high-dose effects in rodents or other experimental systems to the very low-dose level of human exposure would lead to an overestimation of the toxicological risk, thus exaggerating the health hazards resulting from the presence of acrylamide in food and its metabolic conversion to glycidamide in the human body.
Page 37 of 77
4. Exposure Assessment RIKILT Institute of Food Safety (The Netherlands), Swedish Food Administration (Sweden),
Stockholm University (Sweden), National Food Institute (Denmark)
5.1. Monte Carlo Risk Assessment For the work on the exposure assessment the Monte Carlo Risk Assessment (MCRA) software has been used. MCRA was developed in other projects, but adapted slightly for HEATOX purposes e.g. an additional option has been developed to study the effect of brand loyalty on exposure. 5.1.1. Dietary exposure assessment The long-term exposure of the Dutch population has been calculated, using the EU database on acrylamide concentrations. Exposure of the total population ranged from 0.4-1.8 µg/kg bw/day (P50-P99), exposure of children aged 1-6 years ranged from 1.0-2.4 µg.kg bw/day. Also a preliminary calculation of the exposure to furan has been made. With the data available from US FDA in 2004, the exposure to furan of the Dutch population was estimated at 0.25-0.87 µg/kg bw/day (P50-P97.5). With the purpose to validate the MCRA software duplicate diets of infants have been collected and analysed. Since only one of the diets contained a quantifiable amount of acrylamide it was not possible to use this study for validation.
Figure 18. Contribution of food groups to acrylamide exposure in different countries for different age groups.
Page 38 of 77
5.1.2. Scenario-analysis of effect of new processing techniques Within the HEATOX-project research is performed to develop new or adapted processing techniques to reduce acrylamide (AA) levels in the following foods; bread, crispbread, coffee, crisps and French fries. These reduced acrylamide levels were applied in scenario analyses with the aim to give insight in the theoretical reduction in AA exposure. The scenario-analysis showed that the reduction of the AA levels in single food groups had small effects on the exposure. When reductions of all food groups are applied in one scenario ‘total’ this resulted in a reduction of the exposure of the total Dutch population by approximately 40%, to 0.3 µg/kg bw/ day (P50) and to 1.2 µg/kg bw/ day (P99.9 ).For children the exposure decreased to 0.6 µg/kg bw/ day. Figure 19 shows the exposure expressed as Margin of Exposure per scenario for the upper 50% of the population. It should be noted that the new processing techniques which were applied in the scenarios were developed on lab scale and are not yet applied on industrial scale; thus the shown reduction is only potential. 5.1.3. Brand loyalty With the developed brand loyalty model the issue of brand loyalty in the scenario-analysis of acrylamide exposure assessment. The model was applied on the case of potato crisps, for which data on levels of different brands was used from a German data source (www.foodwatch.de), and market shares and brand loyalty were simulated. These calculations showed that brand loyalty has influence on the higher exposure levels and that brands with high acrylamide levels can increase exposure levels depending on the market share.
Figure 19. Distribution of Margin of Exposure (MoE) of the acrylamide reduction scenarios over the percentiles.
5.1.4. Home cooking Only limited data was available on the levels in home-cooked foods and no information was available on the amount of home-cooked foods in the diet. The levels reported for home-cooked foods are mostly within the range of levels in the EU-database, thus the intake from home-cooked foods is at least partly covered by intake calculated with the EU-database. However, from the first estimates we may conclude that home-cooking can have (a probably small) influence on the acrylamide exposure, both in increasing and decreasing exposure, therefore it is important that consumers follow the advice not to overcook their food and follow the available cooking guidelines. From the experiments known today it appears that by following the cooking guidelines acrylamide intake form home-cooked foods can be kept as low as possible.
0
200
400
600
800
1000
1200
1400
50 60 70 80 90 100
percentiles
MoE
without mitigation
fries
crisp
bread
coffee
total
Page 39 of 77
5.1.5. Intake in British and Dutch infants A comparison has been made between the acrylamide exposure of Dutch and UK infants using the same exposure assessments methodology. It appears that UK infants have a two times higher intake based on the difference in their diet of acrylamide containing food items. A standardized international methodology for exposure assessment is recommendable.
5.2. Training on probabilistic exposure assessment A training on probabilistic modelling of exposure to acrylamide was organised for European National Food Safety Authorities, including EFSA. The training was aimed at those working at the authorities and who are involved in risk assessment of acrylamide or other heat-toxicant research. In the training participants learned about the probabilistic approach and learned to work with the Monte Carlo Risk Assessment software in relation to acrylamide. Specific acrylamide issues were discussed and studied through exercises, like the grouping of foods, variability in the acrylamide levels, effect of mitigation measures and the possibility to study and how to quantify these aspects with MCRA. The training was very well received, and all participants were able to use the Monte Carlo Risk Assessment (MCRA) software for probabilistic exposure calculations. The calculation performed on acrylamide exposure gave a good estimate of the exposure of the Dutch population. Work on brand loyalty and home-processing showed that these issues may influence exposure, especially the exposure of the high-end consumers may be increased. The effectiveness of different mitigation possibilities to reduce acrylamide exposure levels has been studied using probabilistic models. Some of the mitigation measures showed a significant reduction on a particular food item, but overall the effect on exposure levels was limited. A significant reduction in exposure can only be received when mitigation measures are applied in a variety of products. Comparing acrylamide levels of different countries has been done, and the methodology of how to perform probabilistic intake calculations and scenario studies regarding the effectiveness of different mitigation options has been trained to different European National Food Authorities. This was received very well, and has resulted in new proposals for future cooperation.
5.3. Estimation of breast cancer risk from exposure to heat-induced toxicants An LC/MS/MS method for the analysis of haemoglobin adducts of acrylamide (AA) and glycidamide (GA) was established. The procedure involves isolation of reed blood cells, lysis and isolation of haemoglobin. Normal and adducted N-terminal valine amino acids are liberated by a modified Edman degradation followed by column purification. Normal and adducted valine residues are separated by gradient HPLC with MS/MS detection. The LC/MS/MS method has been optimised and the use of deutorated internal standards (Figure 20) has been included in the analytical procedure. Standard curves for AA-Val-PTH are linear from 0.3 pmol/ml to 28 pmol/ml and for GA-Val-PTH (two diasteriomers) standard curves are linear from 0.8 pmol/ml to 48 pmol/ml.
Page 40 of 77
Different sample purification methods were evaluated, including solid phase extraction (SPE) with different columns, columns with diatomaceous earth and liquid/liquid extractions. A useful and reproducible method using SPE was established.
Figure 20. Chromatograms showing GA-Val-PTH and AA-Val-PTH (upper) as well as the deutorated internal standards (lower)
From 1993 to 1997 blood samples were collected from approx 30.000 women in Denmark. Since then about 430 postmenopausal women had been diagnosed with breast cancer. Globin was purified from the 430 blood samples from the cancer patients as well from the same number of controls. The globin samples were analysed for content of AA-ValPTH and GA-Val-PTH by the LC/MS/MS method. Results of the unadjusted statistical analysis showed that neither acrylamide [IRR (95% CI) 1.01 (0.65-1.57)] nor glycidamide [IRR (95% CI) 0.79 (0.45-1.38)] concentrations were found to be associated with breast cancer incidence. Inclusion of potential confounding factors in the model did not have clear effects on the association between glycidamide and breast cancer, whereas the association between acrylamide and breast cancer was strengthened after inclusion of information on smoking at blood sampling [IRR (95% CI) 1.43 (0.75-2.71)], further inclusion of former smoking, duration of smoking and amount of tobacco currently smoked [IRR (95% CI) 1.70 (0.85-3.38)], as well as additional inclusion of baseline risk factors for breast cancer [IRR (95% CI) 2.04 (1.00-4.18)]. When the concentrations of glycidamide and acrylamide were evaluated with estrogen receptor (ER) specific breast cancer as the outcome, all associations was found to be strongest with regard to ER+ breast cancer. In the fully adjusted model, women with the highest concentrations of acrylamide adducts were found to be at almost 3-times increased risk of ER+ breast cancer when compared to women with the lowest concentrations (p=0.01).
5 6 7 8 9 10 11 12 13 14 Time [min]0.0
0.5
1.0
1.5
0.0
0.2
0.4
0.6
0.8
1.0
5x10Intens.
1.2
N H 2
OS
NN
O
N H 2
OS
NN
O
O H
AA-Val-PTHGA-Val-PTH
D
DD
N H 2
OS
NN
O
OH D
DD
N H 2
OS
NN
O
[D 3 ]GA-Val-PTH[D 3 ]AA-Val-PTH
m/z 304 233
m/z 307 233
m/z 320 233 302+
m/z 323 233 304+
Page 41 of 77
5.4. Measurement of internal doses of acrylamide and glycidamide – a tool for exposure assessment and risk assessment 5.4.1. Haemoglobin adducts as a measure of internal dose Doses in blood of reactive compounds, like acrylamide and its metabolite glycidamide, could be inferred from levels of their respective adducts to haemoglobin (Hb). Hb adduct levels from acrylamide and glycidamide were quantified as a measure of internal doses in samples from humans and animals in different studies. Preparation of samples and analysis of adducts were performed according to an established method (the N-alkyl Edman procedure). 5.4.2. Bioavailability of acrylamide in food The bioavailability of acrylamide from two different kinds of diets, one containing potato and one containing fibre-rich bread, has been studied in mice. No difference in the bioavailability of acrylamide between the different types of diets was observed. Furthermore, it was shown that the “extra acrylamide”, obtained during alkaline extraction of the fibre-rich diet, does not correspond to bioavailable acrylamide. 5.4.3. Endogenous formation of acrylamide Internal dose of acrylamide in blood were measured in free-living animals with assumed no dietary intake of acrylamide; common hare, red deer, roe deer, red fox and wild boar. In all these species the adduct levels were shown to be very low (below or close to the limit of quantification). The conclusion is that there is no or very low endogenous production of acrylamide in mammals, and that possible endogenous formation gives insignificant contribution in humans compared to intake through diet. 5.4.4. Inter- and intra-variation of internal doses in humans Inter-individual variation in acrylamide- and glycidamide doses was studied in blood samples from 142 smoking and non-smoking individuals in the general population. The acrylamide adduct levels varied between 0.02 to 0.1 nmol/g globin (a factor of 5) in non-smokers. In smokers the range of the acrylamide-adduct level was 0.03 - 0.43 nmol/g globin. The variation in glycidamide doses was slightly higher. The results indicate a non-linearity in the internal doses of glycidamide at low exposure doses of acrylamide. This observation has to be more carefully investigated.
Non-smokers
0
0,02
0,04
0,06
0,08
0,1
0 20 40 60 80
Sample identity
Acry
lam
ide-
addu
ct le
vel (
nmol
/g)
Figure 21. Variation in acrylamide-adduct levels in Hb between non-smoking individuals (n=70).
The intra-individual variation of adduct levels from acrylamide was studied in blood samples collected from 15 non-smokers every second month during 1.5 years. There was an intra-individual variation in the acrylamide adduct levels in Hb over time, assumed to reflect a variation in background exposure. This variation was, however, lower than the inter-individual variation.
Page 42 of 77
5.4.5. Relationship between internal dose and estimated acrylamide intake from food The average acrylamide-intake, estimated from food frequency questionnaires (FFQ), varied nearly with a factor of 50 between the donors of the blood bank samples mentioned above. This means nearly 10 times larger individual variation compared to the measured internal doses (vary with a factor of 5). This illustrates the difficulties with estimation of intakes from FFQ. In two human studies volunteers were given controlled intake of food with known acrylamide content. Levels of Hb-adducts from acrylamide were compared with estimated intakes. In each study, about 20 individuals, smokers and non-smokers were having a 28- respectively 5-days period of controlled intake of acrylamide-rich diet (in average 3 and 11 µg acrylamide/kg and day, respectively). In blood samples, collected before and after the exposure periods, Hb-adducts from acrylamide was measured. Both studies clearly showed increased Hb-adduct levels corresponding to the acrylamide intake in both non-smokers and smokers.
Page 43 of 77
6 Risk assessment and risk communication Swedish Food Administration (Sweden), Norwegian Institute of Public Health (Norway), Stockholm University (Sweden),
National Food Institute (Denmark), RIKILT Institute of Food Safety (The Netherlands), BEUC - European Consumers' Organisation (Belgium), National Veterinary Institute (Norway), Lund University (Sweden)
6.1. Risk assessment 6.1.1. Models for extrapolation of cancer risk According to a relative cancer risk model (applied/developed by the partner) the species extrapolation from cancer tests in rodents should be based on internal doses of the ultimate genotoxic agent in the test species and in humans. Internal doses are obtained from Hb-adduct levels.
Figure 22. Important corner-stones in the approach for cancer risk estimation.
Rats were exposed to acrylamide in drinking water in an experiment simulating the conditions used in published cancer tests, and after one week blood samples were collected. In other studies blood samples from humans with background exposure to acrylamide (samples from blood banks), or from humans with increased exposure to acrylamide (experiments with dietary exposure), were studied. Hb-adduct levels from acrylamide and glycidamide were measured. Complementary experiments have been done to allow calculations of internal doses from the measured adduct levels in rodents and humans. Results with non-linearity of the internal doses of glycidamide at low exposure doses of acrylamide were strongly indicated in the human experimental studies. Taken together, the results indicate a relatively lower dose of glycidamide at low acrylamide doses. However, as these low exposure doses are associated with a larger measurement error, the indicated results of non-linearity has to be further strengthened. The result, if true, considerably complicates the cancer risk estimation. Therefore, further verifying measurements are required. 6.1.2. Final Risk Characterisation The work performed in HEATOX together with results obtained outside the project are summarised and commented below. For non-carcinogenic effects of acrylamide, the most sensitive endpoint is neurotoxicity with a NOAEL (No Observable Adverse Effect Level) in experimental animals of 0.2 mg/kg bw/day. The margins of safety relative the NOAEL for the average and high exposure were 200 and 50, respectively. The safety margins for other effects were larger. Comparing estimated NOAEL for neurotoxicity in humans of 10-40 µg/kg bw/day with exposure from food resulted in a safety margin in the range of 2.5 to 40. Given these margins any risks of neurotoxic and other non-carcinogenic effects following dietary acrylamide exposure are likely to be very small.
Adduct level in blood
Internal dose and uptake
Cancer risk
ReactivityPharmaco-kinetics
Genotoxic potencyCancer risk model
Adduct level in blood
Internal dose and uptake
Cancer risk
ReactivityPharmaco-kinetics
Genotoxic potencyCancer risk model
Page 44 of 77
There are obvious problems with epidemiological studies, particularly those addressing low level exposure through food in the general population using Food Frequency Questionnaires (FFQ). First of all none of the studies published so far were designed for investigating the carcinogenic risk of acrylamide. There is no validation of the exposure assessments, and in some of them all dietary sources of acrylamide had not been taken into account. A comparison of intake estimates based on FFQ with that of acrylamide haemoglobin adducts showed large overlaps; a major problem is the correct classification according to exposure. Misclassifications would strongly weaken a hypothetical association between acrylamide exposure and cancer. The next limitation is the ability of the dietary studies, and even the occupational cohort study, to detect the small increases in risk expected from the lowest to the highest exposure groups. Indeed, neither the epidemiologic study nor the studies on dietary exposure to acrylamide did have the power to detect small increases in cancer risk and should be regarded as non-positive and not negative studies. Another issue is the mechanism of cancer induction by acrylamide. There are examples of chemicals that can induce heritable epigenetic modifications to DNA and thereby change gene expression without altering DNA sequence. So far there are no data to prove that epigenetic changes are important in acrylamide carcinogenesis although, for example, tumour location in animals might indicate a hormonal component in the aetiology. All data reported support a genotoxic mechanism, mediated by glycidamide, for acrylamide carcinogenesis. However, this needs further investigations. JECFA compared exposure estimates of 1 and 4 µg/kg bw/day with the BMDL10 of 30 mg/kg bw/day (BMDL10=Bench Mark Dose Lower confidence limit for 10% increased incidence) and obtained margins of exposure (MOEs) of 300 and 75. JECFA considered these exposure margins to be low for a compound being both carcinogenic and genotoxic and that they indicate a human health concern (WHO, 2005). The Scientific Committee of EFSA (EFSA, 2005) also endorsing a MOE approach for compounds that are both carcinogenic and genotoxic is of the view that a MOE of 10,000 or larger would be of low concern from a public health point of view and might be considered as a low priority for risk management actions. The MOEs for acrylamide are clearly below 10,000, but the Scientific Committee of EFSA gives no guidance for risk managers how to interpret a MOE less than 10,000. However, if a MOE lower than 10,000 had no meaning, the entire exercise would be worthless. A reasonable conclusion is that the MOE for acrylamide gives rise for concern and merits risk management action. Although open for criticism, using the life time risk estimates based on extrapolation and inclusion of the scaling factor the average and high exposure estimates is associated with life time risks of 1.4 to 6.4•10-3, a risk that would be considered a public health concern. The theoretical risk estimates given above are compatible with the non-positive outcome for cancer risk of acrylamide in the epidemiological studies based on FFQ. The cancer risk associated with food borne acrylamide exposure is probably low, and can at most only explain a small fraction of cancers associated with diet. However, for other chemicals a risk level to consumers comparable with that of acrylamide would not be ignored. Actions to reduce the risk by reducing acrylamide in foods and thereby food borne exposure are therefore warranted and generally agreed upon. Lastly, as it is known that a great number of proven or suspected carcinogens, e.g. other Maillard reaction products, such as heterocyclic amines, furfural and furan derivatives together with lipid peroxidation products can be formed during heat treatment of food, it is important to further study the health risk associated with heat-generated food toxicants foods. Acrylamide is not the sole problem.
Page 45 of 77
Acrylamide is not the sole problem.
To summarize, the main conclusions are: • Acrylamide has been classified by WHO, World Health organisation. This conclusion is
strengthened by the project. Following exposure to acrylamide via food the primary concern is the possible risk of cancer.
• Compared with many regulated food carcinogens, the exposure of acrylamide poses a higher estimated risk to European consumers.
• Risk assessments and recommendations to minimize exposure to acrylamide made by WHO are still valid.
• Other compounds formed during cooking of food, for example HMF, Furan, and a variety of Maillard reactants and lipid oxidation products may also constitute an increased cancer risk for consumers. Approximately 50 substances that would require risk assessment have been theoretically identified within the project.
• Current knowledge does not allow for a risk-benefit assessment of cooking with respect to acrylamide or other heat-induced toxicants.
Acrylamide Furan HMF
52 “suspect” compounds
Page 46 of 77
6.2. Risk communication Risk communication strategy and actions have been integrated with research activities in the project. Different kinds of strategic communication tools and channels have been applied.
Society
Authority
Consumer
Industry
Research Inte
llig
en
ce Authority
Consumer
Industry
Research
Dis
sem
inati
on
HEA
TO
X I
D..............
.
........
.
...
© Reksnes
Figure 23. The HEATOX ID was created early in the project to illustrate the main strategic grips and to help researchers focus on the communication tasks.
6.2.1. Risk Communication Strategy To support and strengthen the project profile a Risk Communication Strategy was developed. The strategy was based on the overall HEATOX objectives and an initial analysis of communication challenges based on interviews with key persons within the project and representatives from interested parties. The strategy framework was set up in 2004 and offered as a guideline for risk communication within the project and between the project and the outside world. Main channels and a visual profile for the project was outlined and developed early in the project. Based on own observations and comments in Midterm Review Report, efforts were taken to strengthen risk communication and dissemination activities within HEATOX. During summer 2005, a dialogue was initiated with HEATOX External Panel. The aim being to ensure that end deliverables were as useful and focused as possible and that dissemination from HEATOX to the outside world was as effective as possible. Special risk communication training sessions have been organised targeting the PhDs and young researchers in the project. Considerable attention was given to the designing and organising of the main risk communication event of HEATOX, the Workshop in June 2006. The conclusions from the HEATOX workshop have given clear indications as to how some of the end deliverables should be constructed to meet end user’s needs and expectations.
Page 47 of 77
Did risk communication play the intended role in the HEATOX project? What lessons can be learned? Can HEATOX experiences contribute to making Guidelines for good risk communication practice related to heat-induced toxicants? At the end of the project, evaluations have been organised to try to answer these questions and give further insight into the nature of practical risk communication. To assess the risk communication efforts within HEATOX, two limited web-based surveys were conducted during January 2007. To assess the risk communication efforts between HEATOX and the outside world, short personal interviews with members of the External Panel were conducted in February 2007. The interviews focused on possible beneficiaries from HEATOX work, to what extent HEATOX has succeeded in networking with the outside world. The Panel members were also asked to give their personal reflections related to being members of the Panel. 6.2.1.1. HEATOX Advice to Industry and Catering Industry and catering represent important food chain levels where effective risk minimisation initiatives could contribute to reducing the intake of acrylamide. HEATOX research covers many aspects related to the three main food sectors; • Potato products; from growing conditions to crisp or French fry processing • Cereal products; mainly focused on bread and baking • Coffee; roasting process and influence of raw material parameters. There has been active and close contact between CIAA (the Confederation of the Food and Drink Industries of the EU) and HEATOX during the entire project period. The first version of the CIAA Acrylamide Toolbox was evaluated internally within HEATOX by relevant research groups prior to the decision to build further on this basis rather than developing an alternative HEATOX system. Following advice from the HEATOX External Panel and the outcome of the HEATOX Workshop in 2006, HEATOX results relevant to industry and catering sector have been fed into the CIAA Toolbox. 6.2.1.2. HEATOX Advice to Authorities and Consumer Organisations on Home Cooking and Consumption Acrylamide formation and minimisation strategies have been quite extensively studied in the laboratory and in industrial environments where the processing/heating conditions are better controlled than in home cooking. HEATOX researchers have conducted a number of experiments related to deep-fried French fries, crisps, oven-roasted potato wedges and toasted bread. The general intake of acrylamide for adults is quite similar across Europe. Due to the fact that a large number of foods contributing to acrylamide intake are industrially produced the contribution from home cooking is probably quite small in the general population. The HEATOX Workshop in Graz 2006 identified significant differences in home cooking methods and availability of ingredients within countries and certainly across the different European regions and member states. The national differences in dietary habits and cooking practices, as well as the different availability of ingredients, need to be taken into account by national authorities and consumer organisations when developing material like brochures, web pages, and presentations for consumers. The HEATOX advice is aimed as a tool for National Authorities and Consumer organisations for handling the acrylamide issue in relation to home-cooking. This might include giving cooking advice directly to consumers as well as influencing providers of raw materials, pre-fried products and frying equipment for domestic use.
Page 48 of 77
Home cooking National authorities should highlight the following: Potatoes low in sugar • Low sugar potato varieties • Maintenance of suitable storage temperature during the supply chain • Low sugar levels in prefabricated potato products for domestic frying Best frying temperature • Frying temperature in the range 145 to 170°C for deep frying potatoes • Clear and accurate cooking instruction on the package of pre-fried products • Clear and accurate instruction for fryers for domestic use Golden, not brown! • French fries and roast potatoes cooked to a golden-yellow rather than golden-brown colour • Bread toasted to the lightest colour acceptable Consumption Balance the diet as proposed in national dietary recommendations and integrate acrylamide considerations into the “normal” dietary recommendations. 6.2.1.3. Guidelines for Good Risk Communication Practice related to heat-induced toxicants Objectives related to integrating risk communication activities into HEATOX form the basis for one deliverable: Guidelines to Good Risk Communication Practice related to heat-induced toxicants. Risk communication within HEATOX has had positive effects on the outcome of the project. On a short term basis, the results from the project will be more clearly disseminated towards the targeted audiences. On a long term basis, the HEATOX experience has probably motivated most of the younger researchers and many of the senior researchers to engage in risk communication activities and thus bridging the gap between science and the end users of scientific results. The multidisciplinary and holistic approach has been recognised as important and this will probably influence the final project results. Both senior scientists in capacity of theme and workpackage leaders and the PhDs have taken actively part in risk communication within HEATOX. The planned actions taken towards different stakeholders throughout the project at specific occasions have probably influenced the outcome in a positive way. Many HEATOX scientists have taken actively part in numerous risk communication activities directed towards other stakeholders. The HEATOX network, with the External Panel and Scientific Committee as core has probably had some, but limited, effects on the outcome of the project. Several of the members have been engaged in the scientific dialogue at the HEATOX meetings. The Panel played an important role in recruiting stakeholders to the HEATOX workshop and several of them also played important roles during the actual workshop. The External Panel has contributed to bringing outside knowledge and experiences to the HEATOX table and to some extent also disseminated results on behalf of HEATOX. Some have definitely functioned as HEATOX ambassadors while others have played a less significant role in the HEATOX network. The Guidelines to Good Risk Communication Practice related to heat-induced toxicants have been constructed on the basis of experiences from earlier and ongoing risk communication projects and practices, updated risk communication research and experiences from the HEATOX project. If communicated properly, it is anticipated that these guidelines could have crossover effect on other similar project, enhancing the integration of risk communication into risk-related research even further.
Page 49 of 77
Acknowledgement
The HEATOX project has been carried out as a wide international collaboration with integration of different and complementary research fields, creating a big, interesting and useful network. The success of the HEATOX project has not only been the completion of the tasks and deliverables, but the wide dissemination of results for the benefit of industry, authority and consumers. Dissemination has been achieved (and will continue) as scientific papers for refereed journals; totally around 60 papers so far, or by attendance at national and international meetings and workshops with posters and paper presentations; papers in other journals and presentations in radio and TV. Totally around 20 PhD students or young researchers have been involved in the project, and contributed with enthusiasm, curiosity and thus helped in creating a warm and stimulating atmosphere. Some have already got their Doctoral degree and several Thesis will be presented during the coming years, and I am very proud of the thought that the HEATOX project has been a part of this research. I wish to express my sincere gratitude to all HEATOX partners and members of the External panel for all work and discussions, and to our scientific officers, Achim Boenke who was in charge at the start of the HEATOX project, and Jürgen Lucas who has been our guide throughout most of the project time. It has been a pleasure to work with you and I am looking forward to seeing you again. Lund, April 2007 Life’s truest happiness lies in the friendship we make on the way…
Page
50 of
77
Pu
bli
shab
le r
esu
lts
of
the F
inal
pla
n f
or
usi
ng
an
d d
isse
min
ati
ng
th
e k
no
wle
dg
e
Exp
loit
able
K
now
ledg
e (d
escr
ipti
on)
Exp
loit
able
pr
oduc
t(s)
or
m
easu
re(s
)
Sect
or(s
) of
ap
plic
atio
n
Tim
etab
le
for
com
mer
cial
us
e
Pat
ents
or
othe
r IP
R
prot
ecti
on
Ow
ner
&
Oth
er
Par
tner
(s)
invo
lved
Com
men
ts
P
ubli
cati
ons
Scie
ntif
ic
com
mun
ity
2007
and
fo
rwar
d
See
Par
tner
lis
t, p
age
6.
Mos
t pa
rtne
rs w
ill p
rese
nt t
he r
esul
ts i
n sc
ient
ific
jo
urna
ls a
nd a
t in
tern
atio
nal c
onfe
renc
es
Kno
wle
dge
on th
e ro
le o
f pre
curs
ors
and
cook
ing
cond
itio
ns fo
r th
e fo
rmat
ion
of
acry
lam
ide
in
pota
to p
rodu
cts
Po
tato
indu
stry
Sc
ient
ific
com
mun
ity
P1
The
kno
wle
dge
will
be
acti
vely
dis
sem
inat
ed a
nd u
sed
as a
bas
is in
futu
re p
roje
cts
and
colla
bora
tion
s.
Mea
sure
men
t of
14
C-l
abel
led
acry
lam
ide
(AA
) in
bi
olog
ical
sam
ples
.
T
oxic
olog
ical
st
udie
s
P1
The
kno
wle
dge
will
be
acti
vely
dis
sem
inat
ed a
nd u
sed
as a
bas
is in
futu
re p
roje
cts
and
colla
bora
tion
s.
Form
atio
n of
HM
F du
ring
cof
fee
roas
ting
Red
ucti
on o
f H
MF
form
atio
n
1. I
ndus
try
P2
Dat
abas
e co
ntai
ning
po
tent
ial t
oxic
ants
fo
und
in h
eate
d fo
ods
T
oxic
olog
ical
st
udie
s C
hem
ical
co
mpo
siti
on
P3 a
nd P
8 T
he d
atab
ase
lists
800
vol
atile
pro
duct
s of
the
M
ailla
rd r
eact
ion
and
lipid
oxi
dati
on to
geth
er w
ith
pred
icte
d to
xico
logi
cal d
ata.
In
the
futu
re, t
he
data
base
will
be
conv
erte
d in
to a
sui
tabl
e fo
rm fo
r vi
ewin
g on
the
CSL
web
sit
e w
here
it w
ill b
e m
aint
aine
d an
d up
date
d, a
s ap
prop
riat
e A
sses
smen
t of
opti
mal
set
-up
of
fryi
ng e
nd-p
oint
Ele
ctro
nic
devi
ce a
nd
appr
opri
ate
algo
rith
m
Equ
ipm
ents
for
hom
e co
okin
g an
d ca
teri
ng
fryi
ng
2008
Y
es
Part
ner
5
Page
51 of
77
Exp
loit
able
K
now
ledg
e (d
escr
ipti
on)
Exp
loit
able
pr
oduc
t(s)
or
m
easu
re(s
)
Sect
or(s
) of
ap
plic
atio
n
Tim
etab
le
for
com
mer
cial
us
e
Pat
ents
or
othe
r IP
R
prot
ecti
on
Ow
ner
&
Oth
er
Par
tner
(s)
invo
lved
Com
men
ts
Indi
cati
ons
or
sugg
esti
ons
to b
e ad
dres
sed
to t
he
equi
pmen
t use
r
Gui
delin
es to
be
pri
nted
on
equi
pmen
t m
anua
ls
Equ
ipm
ents
for
hom
e co
okin
g an
d ca
teri
ng
fryi
ng
2007
-200
8 N
o Pa
rtne
r 5
New
kno
wle
dge
on
the
appl
icat
ion
of
stea
m d
urin
g ba
king
.
B
akin
g in
dust
ry
May
be
intr
oduc
ed b
y co
mpa
nies
qu
ite
soon
P6
T
he k
now
ledg
e w
ill b
e ac
tive
ly d
isse
min
ated
. The
n ea
ch c
ompa
ny h
as to
test
, eva
luat
e an
d op
tim
ise
the
tech
niqu
e fo
r ea
ch s
peci
fic a
pplic
atio
n (t
akin
g ac
ryla
mid
e re
duct
ion
and
othe
r qu
alit
y as
pect
s in
to
cons
ider
atio
n). S
IK is
pre
pare
d to
sup
port
on
a co
nsul
tant
bas
is.
New
kno
wle
dge
on
the
appl
icat
ion
of
IR te
chno
logy
in
baki
ng
B
akin
g in
dust
ry
Inve
stm
ents
in
new
type
s of
bak
ing
oven
s ne
eded
be
fore
in
dust
rial
use
P6
A
s ab
ove.
Mit
igat
ion
stra
tegi
es
that
are
hel
pful
to
redu
ce A
A in
take
re
late
d to
hom
e pr
epar
atio
n of
Fr
ench
frie
s
Leaf
let
Con
sum
ers
2007
no
ne
P11
Leaf
let w
ill b
e pr
oduc
ed in
the
cour
se o
f 200
7 an
d m
ade
publ
icly
ava
ilabl
e in
man
y la
ngua
ges.
Page
52 of
77
Exp
loit
able
K
now
ledg
e (d
escr
ipti
on)
Exp
loit
able
pr
oduc
t(s)
or
m
easu
re(s
)
Sect
or(s
) of
ap
plic
atio
n
Tim
etab
le
for
com
mer
cial
us
e
Pat
ents
or
othe
r IP
R
prot
ecti
on
Ow
ner
&
Oth
er
Par
tner
(s)
invo
lved
Com
men
ts
Com
puta
tion
al
mod
el to
cal
cula
te
acry
lam
ide
leve
ls
unde
r va
riou
s pr
oces
sing
co
ndit
ions
Fry
Sim
ulat
or
mod
el
Dut
ch P
otat
o In
dust
ry
Neg
otia
tion
s st
art Q
4/20
06N
one
P11
& P
7
Tab
ulat
ed
info
rmat
ion
on
know
n co
okin
g/
proc
essi
ng
cont
amin
ants
Publ
icat
ion
Food
Ind
ustr
ies
Food
Age
ncie
s 20
07-
2008
P12
Tab
ulat
ed in
form
atio
n.
Kno
wle
dge
of
met
abol
ism
and
in
tern
al d
oses
of
acry
lam
ide
(and
its
met
abol
ite
glyc
idam
ide)
Impr
oved
ca
ncer
ris
k as
sess
men
t fo
r ac
ryla
mid
e
1. A
utho
riti
es
(exp
osur
e lim
its,
e.
g. A
DI)
2.
Can
cer
rese
arch
(e
pide
mio
logy
)
P14
A
mai
n co
mpo
nent
in a
n im
prov
ed c
ance
r ri
sk
asse
ssm
ent o
f acr
ylam
ide
is c
ompa
riso
n of
inte
rnal
do
ses
of g
lyci
dam
ide
- th
e ge
noto
xic
met
abol
ite
of
acry
lam
ide
- in
hum
ans
and
anim
als
that
hav
e be
en
used
in c
ance
r te
sts.
Oth
er p
aram
eter
s th
at a
re
nece
ssar
y (i
nter
med
iate
goa
ls)
for
esti
mat
ion
of th
e ra
nge
of r
isk:
inte
r- a
nd in
tra-
indi
vidu
al v
aria
tion
in
inte
rnal
dos
es, b
ioav
aila
bilit
y of
acr
ylam
ide
in
diff
eren
t foo
dstu
ffs,
pos
sibl
e en
doge
nous
form
atio
n of
ac
ryla
mid
e.
Prob
abili
stic
ex
posu
re c
alcu
lati
on
Mon
te C
arlo
R
isk
Ass
essm
ent
prog
ram
me
(MC
RA
) fo
r sc
enar
io
anal
ysis
of
the
effe
ct o
n ex
posu
re o
f ac
ryla
mid
e re
duct
ion.
1. P
olic
y 2.
Ind
ustr
y 20
07
2008
- -
P17
M
CR
A h
as b
een
deve
lope
d in
oth
er p
roje
cts
and
is in
th
e co
nsor
tium
agr
eem
ent
defin
ed a
s pr
e-ex
isti
ng
know
ledg
e. T
he u
se o
f MC
RA
in a
cryl
amid
e an
d ot
her
heat
-tre
ated
toxi
cant
s ha
s be
en e
xplo
ited
and
w
ill b
e fu
rthe
r ex
ploi
ted
for
rese
arch
que
stio
ns o
f Fo
od I
ndus
try
(in
coop
erat
ion
wit
h IL
SI)
rega
rdin
g th
e ef
fect
iven
ess
of s
ever
al m
itig
atio
n st
eps
as
prop
osed
in th
e T
OO
LBO
X o
f the
CIA
A. F
urth
er
appl
icat
ion
of th
e M
CR
A s
oftw
are
also
in r
elat
ion
to
quan
tita
tive
ris
k-be
nefit
eva
luat
ions
are
fore
seen
. T
here
is a
lso
an in
tere
st o
f nat
iona
l Foo
d St
anda
rd
Age
ncie
s to
use
the
MC
RA
sof
twar
e in
coo
pera
tion
w
ith
the
RIK
ILT
in th
e ne
arby
futu
re.
Page
53 of
77
Exp
loit
able
K
now
ledg
e (d
escr
ipti
on)
Exp
loit
able
pr
oduc
t(s)
or
m
easu
re(s
)
Sect
or(s
) of
ap
plic
atio
n
Tim
etab
le
for
com
mer
cial
us
e
Pat
ents
or
othe
r IP
R
prot
ecti
on
Ow
ner
&
Oth
er
Par
tner
(s)
invo
lved
Com
men
ts
Iden
tific
atio
n of
SU
LT1A
1 as
the
prin
cipa
l hum
an
enzy
me
invo
lved
in
the
toxi
ficat
ion
of
HM
F
Mou
se li
nes
hum
aniz
ed
for
SULT
1A1
1.
Phar
mac
euti
cal
Indu
stry
2.
Tox
icol
ogic
al
cont
ract
rse
arch
2008
or
2009
A
pplic
atio
n pa
tent
en
visa
ged
for
2008
P18
Tox
icok
inet
ics
rela
ted
to
acry
lam
ide
or
glyc
idam
ide
P22
Exp
ress
ion
prof
iling
of h
uman
MC
F7 c
ells
exp
osed
to
diff
eren
t con
cent
rati
ons
of a
cryl
amid
e or
gly
cida
mid
e R
eal-
tim
e re
vers
e-tr
ansc
ript
ase
PCR
val
idat
ion
of 2
5 tr
ansc
ript
s in
MC
F7 c
ells
and
hum
an C
aCo-
2 ce
lls
Mea
sure
men
t of
intr
acel
lula
r gl
utat
hion
e le
vels
aft
er
trea
tmen
t wit
h ac
ryla
mid
e or
gly
cida
mid
e D
eter
min
atio
n of
nuc
lear
fact
or-•
B a
ctiv
ity
usin
g a
repo
rter
gen
e as
say
Stan
dard
toxi
city
and
cel
l via
bilit
y as
says
.
Kno
wle
dge
on
vacu
um fr
ying
to
redu
ce a
cryl
amid
e le
vel i
n po
tato
es
cris
ps
In
dust
ry
P23
Vac
uum
fryi
ng w
ith
and
wit
hout
pre
-tre
atm
ents
has
sh
own
to p
rodu
ce lo
w le
vel a
cryl
amid
e cr
isps
. It i
s po
ssib
le to
impr
ove
thei
r qu
alit
y ch
arac
teri
stic
s. T
he
pre-
trea
tmen
ts r
epre
sent
a p
ract
ical
indu
stri
al
alte
rnat
ive
to r
educ
e ac
ryla
mid
e le
vel i
n po
tato
es
cris
ps
Page
54 of
77
Dis
sem
inat
ion
of K
now
ledg
e SC
IEN
TIF
IC J
OU
RN
AL
Yea
r 20
04
Par
tner
A
utho
r
Jour
nal
Tit
le
See
list,
pa
ge 6
. M
urko
vic,
M.
J. B
ioch
em. B
ioph
ys. M
eth.
61,
161
-16
7, 2
004
Acr
ylam
ide
in A
ustr
ian
Food
s P2
Surd
yk, N
., R
osen
, J.,
And
erss
on, R
. and
Åm
an, P
J.
Agr
ic. F
ood
Che
m.
Eff
ects
of a
spar
agin
es, f
ruct
ose,
and
bak
ing
cond
itio
ns o
n ac
ryla
mid
e co
nten
t in
yeas
t-le
aven
ed w
heat
bre
ad
P4
Fred
riks
son,
H.,
Tal
lvin
g, J
, R
osen
, J. a
nd Å
man
, P.
Cer
eal C
hem
. Fe
rmen
tati
on r
educ
es fr
ee a
spar
agin
es in
dou
gh a
nd a
cryl
amid
e co
nten
t in
brea
d P4
Hus
øy, T
., A
bram
sson
-Z
ette
rber
g, L
., Ø
lstø
rn, H
., Pa
ulse
n J
E.,
and
Ale
xand
er J
.
Mut
atio
n R
esea
rch,
580
, 103
-110
. A
deno
mat
ous
poly
posi
s co
li in
fluen
ces
the
effe
ct o
f the
hea
t ind
uced
m
utag
ens,
PhI
P an
d ac
ryla
mid
e, o
n th
e pr
oduc
tion
of m
icro
nucl
ei in
m
ice
eryt
hroc
ytes
.
P9
Dur
ling,
L. a
nd L
ilian
ne
Abr
amss
on-Z
ette
rber
g, L
.*
Mut
atio
n R
esea
rch,
580
, 103
-110
.
A c
ompa
riso
n of
gen
otox
icit
y be
twee
n th
ree
com
mon
het
eroc
yclic
am
ines
and
acr
ylam
ide.
P9
Dun
ovsk
á L.
, Haj
šlov
á J.
, aj
ka
T.,
Hol
adov
á K
. and
Háj
ková
K.
Cze
ch J
ourn
al o
f Foo
d Sc
ienc
e, V
ol.
22, 2
83-2
86
Cha
nges
of A
cryl
amid
e Le
vels
in F
ood
Prod
ucts
dur
ing
Tec
hnol
ogic
al
Proc
essi
ng
P10
E. B
erm
udo,
V. R
uiz-
Cal
ero,
L.
Puig
nou,
M.T
. Gal
cera
n E
lect
roph
ores
is
Mic
roem
ulsi
on
elec
trok
inet
ic
chro
mat
ogra
phy
for
the
anal
ysis
of
ac
ryla
mid
e in
food
. P1
2
Year
200
5
Hel
lenä
s K
.-E
., A
bram
sson
-Z
ette
rber
g L.
, and
Sko
g K
. Jo
urna
l of A
OA
C I
nter
nati
onal
, Ja
nuar
y/Fe
brua
ry V
ol. 8
8, N
o. 1
, 242
-24
5.
The
HE
AT
OX
pro
ject
,
P1
P9
Bag
dona
ite,
K.,
Mur
kovi
c, M
. C
zech
J. F
ood
Sci.
22, 2
2-24
, 200
5 Fa
ctor
s af
fect
ing
the
form
atio
n of
acr
ylam
ide
in c
offe
e P2
Mus
tafa
, A.,
Ade
rsso
n, R
., R
osen
, J.,
Kam
al-E
ldin
A. a
nd
Åm
an P
.
J. A
gric
. Foo
d C
hem
. 200
5, 5
3, 5
985-
5989
Fa
ctor
s in
fluen
cing
acr
ylam
ide
cont
ent a
nd c
olor
in r
ye c
risp
bre
ad
P4
Ols
son,
E.E
.M.,
Trä
gård
h, A
.C.
& A
hrné
, L.M
. J.
Foo
d Sc
i. 70
(8),
E48
4-49
1 (2
005)
E
ffec
t of N
ear-
infr
ared
Rad
iati
on a
nd J
et I
mpi
ngem
ent H
eat T
rans
fer
on C
rust
For
mat
ion
of B
read
P6
Page
55 of
77
Kno
l,J.J
; Van
Loo
n,W
.A.M
. van
; Li
nsse
n,J.
P.H
.; R
uck,
A.L
.; V
an
Boe
kel,M
.A.J
.S.;
Vor
agen
,A.G
J.
J. o
f Agr
ic. F
ood
Che
m. 2
005,
53,
61
33-6
139.
T
owar
d a
Kin
etic
Mod
el fo
r A
cryl
amid
e Fo
rmat
ion
in a
Glu
cose
-A
spar
agin
e R
eact
ion
Syst
em.
P7
L. C
astle
and
S. E
riks
son
Jour
nal o
f AO
AC
Int
erna
tiona
l, 20
05,
88, 2
74-2
84.
Ana
lyti
cal m
etho
ds u
sed
to m
easu
re a
cryl
amid
e co
ncen
trat
ions
in
food
s.
P8
L. M
. Ow
en, L
. Cas
tle, J
. Kel
ly,
L. W
ilson
and
A. S
. Llo
yd
Jour
nal o
f AO
AC
Int
erna
tiona
l, 20
05,
88, 2
85-2
91.
Acr
ylam
ide
anal
ysis
: Ass
essm
ent o
f res
ults
from
six
rou
nds
of F
ood
Ana
lysi
s Pe
rfor
man
ce A
sses
smen
t Sch
eme
(FA
PAS)
pro
ficie
ncy
test
ing.
P8
P. F
ohge
lber
g, J
. Ros
én,
K.-
E.
Hel
lenä
s, L
. Abr
amss
on-
Zet
terb
erg*
Food
and
Che
mic
al T
oxic
olog
y, 4
3: 9
51-
959.
T
he a
cryl
amid
e in
take
via
som
e co
mm
on b
aby
food
for
child
ren
in
Swed
en d
urin
g th
eir
first
yea
r of
life
- (
an im
prov
ed m
etho
d fo
r an
alys
is o
f acr
ylam
ide)
.
P9
Pete
rsso
n E
.V.,
Ros
én J
., T
urne
r C
., D
anie
lsso
n R
., H
elle
näs
KE
. A
naly
tica
Chi
mic
a A
cta
Cri
tica
l fac
tors
and
pit
falls
aff
ecti
ng th
e ex
trac
tion
of a
cryl
amid
e fr
om
food
s: A
n op
tim
isat
ion
stud
y
P9
Dun
ovsk
á L.
, aj
ka T
. and
H
ajšl
ová
J.
Che
mic
ké li
sty
Jour
nal,
99 2
79-2
80,
2005
, Spe
cial
Iss
ue 1
4 A
cryl
amid
e Le
vels
in F
oods
tuff
s fr
om C
zech
mar
ket
P10
E. B
erm
udo,
O. N
úñez
, L.
Puig
nou,
M.T
. Gal
cera
n Jo
urna
l of C
hrom
atog
raph
y A
A
naly
sis
of a
cryl
amid
e in
food
sam
ples
by
capi
llary
zon
e el
ectr
opho
resi
sP1
2
E. B
arce
ló-B
arra
chin
a, F
.J.
Sant
os, L
. Pui
gnou
, M.T
. G
alce
ran
Ana
lyti
ca C
him
ica
Act
a D
eter
min
atio
n of
het
eroc
yclic
am
ines
in a
mea
t ext
ract
as
dim
ethy
lform
amid
e di
alky
lace
tal d
eriv
ativ
es b
y ga
s ch
rom
atog
raph
y m
ass
spec
trom
etry
.
P12
Hag
mar
, L.,
Wir
fält
E.,
Paul
sson
, B.,
Tör
nqvi
st, M
. M
utat
ion
Res
earc
h 58
0, 1
57.1
65
Diff
eren
ces
in h
emog
lobi
n ad
duct
leve
ls o
f acr
ylam
ide
in th
e ge
nera
l po
pula
tion
wit
h re
spec
t to
diet
ary
inta
ke, s
mok
ing
habi
ts a
nd g
ende
r P1
4
Tho
mas
Bje
llaas
, Kar
el J
anak
, E
lsa
Lund
anes
, Geo
rg B
eche
r
Xen
obio
tica
D
eter
min
atio
n an
d qu
anti
ficat
ion
of u
rina
ry m
etab
olit
es a
fter
die
tary
ex
posu
re to
acr
ylam
ide
P16
Boo
n PE
, Mul
A d
e, V
oet H
van
de
r, D
onke
rsgo
ed G
van
, Bre
tte
M, K
lave
ren
JD v
an
Mut
atio
n R
esea
rch
580,
143
-155
; 20
05
Cal
cula
tion
s of
die
tary
exp
osur
e to
acr
ylam
ide
P17
H. R
. Gla
tt, H
. Sch
neid
er, Y
.-G
. Li
u
Mut
atio
n R
esea
rch
580
(200
5) 4
1-52
V
79-h
CY
P2E
1-hS
ULT
1A1,
a c
ell l
ine
for
the
sens
itiv
e de
tect
ion
of
geno
toxi
c ef
fect
s in
duce
d by
car
bohy
drat
e py
roly
sis
prod
ucts
and
oth
er
food
-bor
ne c
hem
ical
s
P18
H. R
. Gla
tt, W
. Mei
nl
Met
hods
in E
nzym
olog
y 40
0 (2
005)
: 23
0-24
9 Su
lfotr
ansf
eras
es a
nd a
cety
ltran
sfer
ases
in m
utag
enic
ity
test
ing:
te
chni
cal a
spec
ts
P18
Page
56 of
77
Yea
r 20
06
Bag
dona
ite,
K.,
Vik
lund
, G.,
Skog
, K.,
Mur
kovi
c, M
. J.
Bio
chem
. Bio
phys
. Met
hods
, 69,
21
5-22
1, 2
006
Ana
lysi
s of
3-a
min
opro
pion
amid
e: A
pot
enti
al p
recu
rsor
of a
cryl
amid
e P2
, P1
Mur
kovi
c, M
., Pi
chle
r, N
. M
olec
ular
Nut
riti
on &
Foo
d R
esea
rch,
50
, 842
-846
(20
06)
Ana
lysi
s of
5-h
ydro
xym
ethy
lfurf
ural
in c
offe
e, d
ried
frui
ts a
nd u
rine
. P2
Mur
kovi
c, M
., D
erle
r, K
. J.
Bio
chem
. Bio
phys
. Met
hods
, 69,
25-
32, 2
006
Ana
lysi
s of
am
ino
acid
s an
d ca
rboh
ydra
tes
in g
reen
cof
fee.
P2
Low
, M. Y
.; K
outs
idis
, G.;
Park
er, J
. K.;
Elm
ore,
J. S
.; D
odso
n, A
. T.;
Mot
tram
, D. S
.
Jour
nal o
f Agr
icul
tura
l and
Foo
d C
hem
istr
y E
ffec
t of c
itri
c ac
id a
nd g
lyci
ne a
ddit
ion
on a
cryl
amid
e an
d fla
vor
in a
po
tato
mod
el s
yste
m.
NB
Wor
k pa
rtly
supp
orte
d by
Hea
tox
P3
Mar
tin
Fink
, Rog
er A
nder
sson
, Jo
han
Ros
én, a
nd P
er Å
man
C
erea
l Che
mis
try
Eff
ect o
f Add
ed A
spar
agin
e an
d G
lyci
ne o
n A
cryl
amid
e C
onte
nt in
Y
east
-Lea
vene
d B
read
P4
S. R
oman
i*, A
. Gas
parr
i*, M
. B
acch
iocc
a*, E
. Coc
ci*,
P. R
occu
li*,
M. D
alla
Ros
a*
Indu
stri
e al
imen
tari
(c
hiri
otti
ed.
, pin
erol
o –
to –
ital
y)
Con
tenu
to in
acr
ilam
mid
e e
cara
tter
isti
che
qual
itat
ive
di p
atat
e fr
itte
in
diff
eren
ti a
ppar
ati t
ecno
logi
ci –
Acr
ylam
ide
cont
ent a
nd c
hara
cter
siti
cs
of p
otat
o fr
ied
in d
iffer
ent f
ryer
s
P5
S. R
oman
i, M
. Bac
chio
cca,
P.
Roc
culi,
M. D
alla
Ros
a A
ccep
ted
to b
e pu
blis
hed
on E
urop
ean
Food
Res
earc
h A
nd T
echn
olog
y E
ffec
t of f
ryin
g ti
me
on a
cryl
amid
e co
nten
t and
qua
lity
aspe
cts
of
Fren
ch fr
ies
P5
Wen
zl, T
., K
aras
ek, L
., R
osen
, J.
, Hel
lena
es, K
-E.,
Cre
ws,
C.,
Cas
tle, L
. and
Ank
lam
, E.
Jour
nal
of
Chr
omat
ogra
phy
A.
2006
11
32,
211-
21.
Col
labo
rati
ve tr
ial v
alid
atio
n st
udy
of tw
o m
etho
ds, o
ne b
ased
on
high
pe
rfor
man
ce li
quid
chr
omat
ogra
phy–
tand
em m
ass
spec
trom
etry
and
on
gas
chr
omat
ogra
phy–
mas
s sp
ectr
omet
ry fo
r th
e de
term
inat
ion
of
acry
lam
ide
in b
aker
y an
d po
tato
pro
duct
s.
P8
Has
nip,
S.,
Cre
ws,
C. a
nd
Cas
tle, L
. Fo
od A
ddit
ives
and
Con
tam
inan
ts,
2006
, 23
, 219
-227
. So
me
fact
ors
affe
ctin
g th
e fo
rmat
ion
of fu
ran
in h
eate
d fo
ods.
P8
Gol
man
n, T
.,Per
isse
t, A
., B
erth
olet
, M.-
C.,
Stad
ler,
R. H
., Pe
ters
son,
E. V
., H
elle
näs,
K.-
E.
Food
Add
it. C
onta
m.,
23(5
), 4
37-4
45
Impa
ct o
f ext
ract
ion
cond
itio
ns o
n th
e co
nten
t of a
cryl
amid
e in
mod
el
syst
ems
and
food
. P9
Wen
zl, T
., K
aras
ek, L
., R
osén
, J.
, Hel
lena
es, K
.-E
., C
rew
s, C
., C
astle
, L.,
and
Ank
lam
, E.
J. C
hrom
. A.,
doi:1
0.10
16/j
.chr
oma.
2006
.07.
007
Col
labo
rati
ve tr
ial v
alid
atio
n st
udy
of tw
o m
etho
ds, o
ne b
ased
on
high
pe
rfor
man
ce li
quid
chr
omat
ogra
phy–
tand
em m
ass
spec
trom
etry
and
on
gas
chr
omat
ogra
phy–
mas
s sp
ectr
omet
ry fo
r th
e de
term
inat
ion
of
acry
lam
ide
in b
aker
y an
d po
tato
pro
duct
s.
P9
Dun
ovsk
á L.
, aj
ka T
., H
ajšl
ová
J., H
olad
ová
K.
Ana
lyti
ca C
him
ica
Act
a, V
olum
e 57
8,
Issu
e 2,
25
Sept
embe
r 20
06, 2
34-2
40
Dir
ect d
eter
min
atio
n of
acr
ylam
ide
in fo
od b
y ga
s ch
rom
atog
raph
y-hi
gh-r
esol
utio
n ti
me-
of-f
light
mas
s sp
ectr
omet
ry
P10
Page
57 of
77
E. B
erm
udo,
O. N
úñez
, L.
Puig
nou,
M.T
. Gal
cera
n J.
Chr
omat
ogr.
A, 1
120
(200
6) 1
99-
204
A
naly
sis
of a
cryl
amid
e in
food
sam
ples
by
capi
llary
zon
e el
ectr
opho
resi
sP1
2
E. B
erm
udo,
O. N
úñez
, L.
Puig
nou,
M.T
. Gal
cera
n J.
Chr
omat
ogr.
A, 1
129
(200
6) 1
29-
134
Ana
lysi
s of
acr
ylam
ide
in fo
od p
rodu
cts
by in
-lin
e pr
econ
cent
rati
on
capi
llary
zon
e el
ectr
opho
resi
s P1
2
E. T
eixi
dó, F
.J. S
anto
s, L
. Pu
igno
u, M
.T. G
alce
ran
J. C
hrom
atog
r. A
, 113
5 (2
006)
85-
90
Ana
lysi
s of
5-h
ydro
xym
ethy
lfurf
ural
in fo
ods
by g
as c
hrom
atog
raph
y-m
ass
spec
trom
etry
P1
2
T B
jella
as, L
H S
tøle
n, M
H
auge
n, J
E P
auls
en, J
A
lexa
nder
, E L
unda
nes
and
G
Bec
her
Food
and
Che
mic
al T
oxic
olog
y
2006
Dec
19;
[E
pub
ahea
d of
pri
nt]
Uri
nary
acr
ylam
ide
met
abol
ites
as
biom
arke
rs fo
r sh
ort-
term
die
tary
ex
posu
re to
acr
ylam
ide.
P1
6
Year
200
7
G. V
iklu
nd, F
. Men
doza
, I.
Sjö
holm
and
K. S
kog
LW
T -
Foo
d Sc
ienc
e an
d T
echn
olog
y,
40, i
ssue
6, 1
066-
1071
A
n ex
peri
men
tal s
et-u
p fo
r st
udyi
ng a
cryl
amid
e fo
rmat
ion
in p
otat
o cr
isps
P1
G. V
iklu
nd,.
Ols
son,
K.,
Sjöh
olm
I. a
nd K
. Sko
g su
bmit
ted
Eff
ect o
f pot
ato
vari
ety,
yea
r an
d st
orag
e co
ndit
ions
for
the
form
atio
n of
acr
ylam
ide
in c
risp
s P1
G. V
iklu
nd,.
Ols
son,
K.,
Sjöh
olm
I. a
nd K
. Sko
g su
bmit
ted
Bla
nchi
ng r
educ
es p
recu
rsor
con
tent
and
acr
ylam
ide
form
atio
n in
fo
rmat
ion
in c
risp
s P1
Mus
tafa
, A.,
Åm
an, P
., A
nder
sson
, R. K
amal
-Eld
in A
. Fo
od C
hem
istr
y,
subm
itte
d A
naly
sis
of fr
ee a
min
o ac
ids
in c
erea
l pro
duct
s P4
Ahr
ne, L
., A
nder
sson
, C.-
G.,
Flob
erg,
P.,
Ros
én, J
. &
Ling
nert
, H.
LWT
– F
ood
Scie
nce
and
Tec
hnol
ogy,
In
pre
ss
Eff
ect o
f cru
st te
mpe
ratu
re a
nd w
ater
con
tent
on
acry
lam
ide
form
atio
n du
ring
bak
ing
of w
hite
bre
ad: s
team
and
falli
ng te
mpe
ratu
re b
akin
g P6
J.J.
Kno
l, G
. Vik
lund
, J.P
.H.
Lins
sen,
I. S
jöho
lm, K
. Sko
g,
M.A
.J.S
. van
Boe
kel
Jour
nal o
f Mol
ecul
ar N
utri
tion
and
Fo
od R
esea
rch
Kin
etic
Mod
elin
g: A
Too
l to
Und
erst
and
the
Form
atio
n of
Acr
ylam
ide
in F
ood
In p
repa
ratio
n
P7, P
1
C. C
rew
s an
d L.
Cas
tle
Tre
nds
in F
ood
Scie
nce
and
Tec
hnol
ogy,
in p
ress
R
evie
w o
f the
occ
urre
nce,
form
atio
n an
d an
alys
is o
f fur
an in
hea
t-pr
oces
sed
food
s P8
C. C
rew
s, S
Has
nip,
D. P
. T.
Rob
erts
and
L. C
astle
Fo
od A
ddit
ives
& C
onta
min
ants
,
in p
ress
. Fa
ctor
s af
fect
ing
the
anal
ysis
of f
uran
in h
eate
d fo
ods.
P8
C. C
rew
s an
d L.
Cas
tle.
LC-G
C E
urop
e, in
pre
ss
The
det
erm
inat
ion
of fu
ran
in fo
ods-
cha
lleng
es a
nd s
olut
ions
. P8
D. P
. T. R
ober
ts, C
. Cre
ws
and
H. G
rund
y M
anus
crip
t in
prep
arat
ion.
E
ffec
t of c
onsu
mer
coo
king
on
fura
n in
con
veni
ence
food
s
P8
Page
58 of
77
Vol
lebr
egt,
H.M
.; Z
onde
rvan
, C
. J.
Hea
t Tra
nsfe
r A
kin
etic
and
ther
mod
ynam
ic m
odel
to p
redi
ct le
vels
of a
cryl
amid
e in
he
ated
food
pro
duct
s (t
enta
tive
) P1
1
M.S
. Alta
ki, F
.J. S
anto
s, M
.T.
Gal
cera
n J.
Chr
omat
ogr.
A
Ana
lysi
s of
fura
n in
food
s by
hea
dspa
ce s
olid
-pha
se m
icro
extr
acti
on-g
as
chro
mat
ogra
phy-
ion
trap
mas
s sp
ectr
omet
ry
P12
E. B
erm
udo,
O. N
úñez
, E.
Moy
ano,
L. P
uign
ou, M
.T.
Gal
cera
n
J. C
hrom
atog
r. A
Fi
eld
Am
plifi
ed S
impl
e In
ject
ion-
Cap
illar
y E
lect
roph
ores
is-T
ande
m
Mas
s Sp
ectr
omet
ry fo
r th
e an
ális
is o
f acr
ylam
ide
in fo
odst
uffs
P12
A V
ikst
röm
, S E
riks
son,
B
Paul
sson
, P K
arls
son
and
M
Tör
nqvi
st
subm
itte
d C
ompa
riso
n of
bio
avai
labi
lity
of a
cryl
amid
e in
diff
eren
t foo
ds-a
stu
dy
in m
ice
P14
E W
irfä
lt, B
Pau
lsso
n, M
T
örnq
vist
, A A
xmon
and
L
Hag
mar
Eur
opea
n Jo
urna
l of C
linic
al N
utri
tion
(i
n pr
ess,
doi
10.
1038
/sj.e
jcn.
1602
704)
Ass
ocia
tion
s be
twee
n es
tim
ated
acr
ylam
ide
(AA
) in
take
s, a
nd H
b A
A-
addu
cts
in a
sam
ple
from
the
Mal
mö
Die
t and
Can
cer
coho
rt
P14
T B
jella
as*,
† , P T
honn
ing
Ole
sen‡ , H
Fra
ndse
n‡ , M
Hau
gen* , L
L-H
Stø
len* , J
E
Paul
sen* , J
Ale
xand
er* , E
Lu
ndan
es† a
nd G
Bec
her*,
†
Tox
icol
ogic
al S
cien
ces,
sub
mit
ted
Com
pari
son
of e
stim
ated
die
tary
inta
ke o
f acr
ylam
ide
wit
h ha
emog
lobi
n ad
duct
s of
acr
ylam
ide
and
glyc
idam
ide
P15
R.V
. Hed
egaa
rd, H
. Fra
ndse
n,
K. G
ranb
y, A
. Apo
stol
opou
la
and
L. S
kibs
ted
Jour
nal o
f foo
d an
d A
gric
ultu
ral
Che
mis
try,
sub
mit
ted
Influ
ence
of w
ater
act
ivit
y on
acr
ylam
ide
form
atio
n fr
om g
luco
se a
nd
aspa
ragi
nes
in a
queo
us g
lyce
rol
P15
Cle
men
t F, D
ip R
, Nae
geli
H
Can
cer
Epi
dem
iolo
gy B
iom
arke
rs a
nd
Prev
enti
on (
Subm
itte
d)
Exp
ress
ion
Sign
atur
e of
Gly
cida
mid
e, th
e R
eact
ive
Met
abol
ite
of th
e W
ides
prea
d Fo
od C
arci
noge
n A
cryl
amid
e P2
2
Page
59 of
77
Boo
k C
hapt
ers
Y
ear
2006
Aut
hor
Boo
k T
itle
P
artn
ner
Mot
tram
, D. S
; Low
, M. Y
.; E
lmor
e, J
. S
In: A
cryl
amid
e an
d ot
her
Haz
ardo
us
Com
poun
ds in
Hea
t-T
reat
ed F
oods
. K
Sko
g; J
Ale
xand
er (
Eds
), W
oodh
ead
Publ
ishi
ng
The
Mai
llard
rea
ctio
n an
d it
s ro
le in
the
form
atio
n of
acr
ylam
ide
and
othe
r po
tent
ially
haz
ardo
us c
ompo
unds
in fo
ods
P3
Q. C
haud
hry,
J. C
otte
rill,
R.
Wat
kins
and
N. P
rice
In
‘Acr
ylam
ide
and
othe
r he
alth
ha
zard
ous c
ompo
unds
in h
eat-
trea
ted
food
s’. p
p. 1
32-1
59.
K
. Sko
g an
d J.
Ale
xand
er (
Eds
),
Woo
dhea
d Pu
blis
hing
, 200
6. I
SBN
97
8-1-
8456
9-01
1-3.
A m
olec
ular
mod
ellin
g ap
proa
ch to
pre
dict
the
toxi
city
of c
ompo
unds
ge
nera
ted
duri
ng h
eat t
reat
men
t of f
ood.
P8
L. C
astle
In
‘Acr
ylam
ide
and
othe
r he
alth
ha
zard
ous c
ompo
unds
in h
eat-
trea
ted
food
s’. p
p. 1
17-1
31.
K
. Sko
g an
d J.
Ale
xand
er (
Eds
),
Woo
dhea
d Pu
blis
hing
, 200
6. I
SBN
-13
: 978
-1-8
4569
-011
-3.
Ana
lysi
s fo
r ac
ryla
mid
e in
food
s.
P8
Oth
er P
ubli
cati
ons
Y
ear
2004
Hül
ya E
rol G
ezgi
n D
ünya
GID
A-M
ayıs
200
4 Is
ıl i
lem
Gör
en G
ıdal
arda
Kan
ser
Ris
ki (
Can
cer
Ris
k in
Hea
t Tre
ated
Fo
ods)
P13
K G
ranb
y, H
Fra
ndse
n, K
Kac
k,
BB
B J
ense
n, J
A N
iels
en
Alim
enta
A
cryl
amid
i fø
deva
rer
P15
Yea
r 20
05
Per
Åm
an
Livs
med
el I
Fok
us, 2
005,
7 p
p 57
A
kryl
amid
i br
öd
P4
J.J.
Kno
l, J.
P.H
. Lin
ssen
and
M
.A.J
.S. v
an B
oeke
l B
i-/t
rien
nial
per
iodi
cal o
n cu
rren
t re
sear
ch o
f the
gra
duat
e sc
hool
VLA
G
Kin
etic
Mod
elin
g of
Acr
ylam
ide
Form
atio
n in
Mod
el S
yste
ms
P7
Page
60 of
77
L C
astle
FS
A F
ood
Surv
ey I
nfor
mat
ion
Shee
t 71
/05.
Jan
uary
200
5.
http
://w
ww
.food
.gov
.uk/
mul
tim
edia
s/fs
is71
2005
#pag
e=5
Ana
lysi
s of
Tot
al D
iet S
tudy
sam
ples
for
acry
lam
ide.
P8
Hül
ya Ö
lmez
-Son
gün
Dem
irel
D
ünya
GID
A-O
cak
2005
T
ahıl
Baz
lı G
ıdak
lard
a A
krila
mid
(A
cryl
amid
e in
cer
eal b
ased
pr
oduc
ts)
P13
Jaco
b D
. van
Kla
vere
n, P
olly
E.
Boo
n an
d A
nika
de
Mul
W
oodh
ead
publ
ishi
ng. (
subm
itte
d)
Mod
ellin
g of
die
tary
exp
osur
e to
acr
ylam
ide,
Acr
ylam
ide
and
othe
r he
alth
haz
ardo
us c
ompo
unds
in h
eat-
trea
ted
food
s.
P17
H. R
. Gla
tt
In: H
uman
Sul
phot
rans
fera
ses (
G. M
. Pa
cific
i, M
. W. H
. Cou
ghtr
ie, e
ds.)
T
aylo
r &
Fra
ncis
, Lon
don,
p. 2
81-3
06
Act
ivat
ion
and
inac
tiva
tion
of c
arci
noge
ns b
y hu
man
sul
fotr
ansf
eras
es
P18
H. R
. Gla
tt, W
. Teu
bner
, W.
Mei
nl
In: 1
5th
Inte
rnat
iona
l Sym
posiu
m o
n M
icro
som
es a
nd D
rug
Oxi
datio
ns, h
eld
in M
ainz
(G
erm
any)
(F.
Oes
ch, e
d.),
M
edim
ond/
Mon
duzz
i, B
olog
na, p
. 9-
15
Sulfo
tran
sfer
ases
exp
ress
ed in
the
hum
an g
astr
o-in
test
inal
trac
t and
th
eir
abili
ty to
act
ivat
e pr
omut
agen
s/ca
rcin
ogen
s P1
8
Yea
r 20
06
Skog
K. a
nd A
lexa
nder
J. (
Eds
) A
cryl
amid
e an
d ot
her
haza
rdou
s co
mpo
unds
in h
eat-
trea
ted
food
s,
Woo
dhea
d Pu
blis
hing
Ltd
BO
OK
wit
h co
ntri
buti
ons
of m
any
HE
AT
OX
par
tner
s P1
, P16
Skog
K, H
elle
näs
K-E
. and
Li
ngne
rt H
. Li
vsm
edel
i fo
kus
CIA
A-w
orks
hop
krin
g ak
ryla
mid
P1
, P6,
P9
Sk
og K
. Fo
od S
cien
ce a
nd T
echn
olog
y, in
pre
ss
The
HE
AT
OX
pro
ject
: Hea
t-ge
nera
ted
food
toxi
cant
s: I
dent
ifica
tion
, C
hara
cter
isat
ion
and
Ris
k M
inim
isat
ion,
P1
Mur
kovi
c M
Acr
ylam
ide
and
othe
r ha
zard
ous
com
poun
ds in
hea
t tre
ated
food
s, S
kog
K, A
lexa
nder
J (
Eds
.)
Woo
dhea
d Pu
blis
hing
Ltd
, Cam
brid
ge
Mec
hani
sm fo
r th
e fo
rmat
ion
of P
hIP
P2
Mur
kovi
c M
Scha
dsto
ffe
in L
eben
smit
teln
und
Fu
tter
mit
teln
, Cic
hna-
Mar
kl M
, So
ntag
G, S
tidl
R (
Eds
.) G
esel
lsch
aft
Öst
erre
ichi
sche
r C
hem
iker
, pp.
2-1
0,
Wie
n, 2
006
Bild
ung
von
hete
rozy
klis
chen
aro
mat
isch
en A
min
en b
eim
Erh
itze
n vo
n Fl
eisc
h
P2
Page
61 of
77
Bag
dona
ite
K, M
urko
vic
M
Sc
hads
toff
e in
Leb
ensm
itte
ln u
nd
Futt
erm
itte
ln, C
ichn
a-M
arkl
M,
Sont
ag G
, Sti
dl R
(E
ds.)
Ges
ells
chaf
t Ö
ster
reic
hisc
her
Che
mik
er, p
p. 2
09-
214,
Wie
n, 2
006
Form
atio
n of
acr
ylam
ide
in c
offe
e
P2
Low
, M. Y
.; E
lmor
e, J
. S.;
Mot
tram
, D. S
. In
: Fla
vour
Sci
ence
- R
ecen
t Adv
ance
s an
d T
rend
s. W
. Bre
die;
M. P
eter
sen
(Eds
), E
lsev
ier.
Rel
atio
nshi
p be
twee
n ac
ryla
mid
e fo
rmat
ion
and
the
gene
rati
on o
f fla
vour
in h
eate
d fo
ods
NB
Wor
k pa
rtly
supp
orte
d by
Hea
tox
P3
Pehr
sson
, S
NyT
ekni
k, 4
8(20
06),
5
Så b
akar
du
akry
lam
idfr
itt
P6
M T
örnq
vist
, B P
auls
son
and
S O
ster
man
-Gol
kar
Acr
ylam
ide
and
othe
r ha
zard
ous
com
poun
ds in
hea
t-tr
eate
d fo
ods,
K
erst
in S
kog
and
Jan
Ale
xand
er
Bio
-mon
itor
ing
of a
cryl
amid
e P1
4
SH H
anse
n, E
J Sø
derl
und,
D
And
erso
n, A
Bau
mga
rtne
r, R
G
opal
an a
nd G
Bru
nbor
g
14th
Eur
opea
n T
esti
s W
orks
hop,
20
06. P
rogr
amm
e an
d m
inip
oste
rs
Abs
trac
t: D
NA
dam
agin
g ef
fect
s of
the
food
con
tam
inan
t acr
ylam
ide
in m
ale
germ
cel
ls
P16
Jaco
b D
. van
Kla
vere
n, P
olly
E.
Boo
n an
d A
nika
de
Mul
Boo
k: A
cryl
amid
e an
d ot
her
heal
th
haza
rdou
s co
mpo
unds
in h
eat-
trea
ted
food
s (W
oodh
ead
publ
ishi
ng)
Mod
ellin
g of
die
tary
exp
osur
e to
acr
ylam
ide,
Acr
ylam
ide
and
othe
r he
alth
haz
ardo
us c
ompo
unds
in h
eat-
trea
ted
food
s
P17
H. R
. Gla
tt, Y
. Som
mer
In
: Acr
ylam
ide
and
Oth
er H
ealth
H
azar
dous
Com
poun
ds in
Hea
t-tr
eate
d Fo
ods (
K. S
kog,
J. A
lexa
nder
, eds
.),
Woo
dhea
d Pu
blis
hing
, Cam
brid
ge, p
. 32
8-35
7
Hea
lth r
isks
by
5-hy
drox
ymet
hylfu
rfur
al (
HM
F) a
nd r
elat
ed
com
poun
ds
P18
Flur
ina
Cle
men
t, Su
sann
e H
alle
r-B
rem
U
NIM
AG
AZ
IN, U
nive
rsit
y of
Zür
ich
Car
cino
geni
c br
eakf
ast c
erea
ls?
P22
Yea
r 20
07
Skog
K
Food
Sci
ence
and
Tec
hnol
ogy,
21,
25-
27
The
HE
AT
OX
pro
ject
: Hea
t-ge
nera
ted
food
toxi
cant
s: I
dent
ifica
tion
, C
hara
cter
isat
ion
and
Ris
k M
inim
isat
ion
P1
Mur
kovi
c, M
., C
onsu
mer
Pro
tect
ion
thro
ugh
Food
Pr
oces
s Im
prov
emen
t a&
Inn
ovat
ion
in th
e R
eal W
orld
, Ed.
E. L
azos
; Vol
3,
2007
, pp.
502
-510
Tox
ic s
ubst
ance
s in
hea
ted
food
s
P2
Page
62 of
77
J.S.
Elm
ore
& Q
.Cha
udhr
y D
atab
ase
in M
icro
soft
Acc
ess
The
Hea
tox
data
base
of p
oten
tial
toxi
cant
s re
sulti
ng fr
om li
pid
oxid
atio
n P3
J.S.
Elm
ore
& Q
.Cha
udhr
y D
atab
ase
in M
icro
soft
Acc
ess
The
Hea
tox
data
base
of p
oten
tial
toxi
cant
s re
sulti
ng fr
om th
e M
ailla
rd
reac
tion
P3
J.J.
Kno
l, J.
P.H
. Lin
ssen
and
M
.A.J
.S. v
an B
oeke
l B
i-/t
rien
nial
per
iodi
cal o
n cu
rren
t re
sear
ch o
f the
gra
duat
e sc
hool
VLA
G
Kin
etic
Mod
elin
g of
Acr
ylam
ide
Form
atio
n in
Mod
el S
yste
ms
P7
J.J.
Kno
l Ph
D T
hesi
s, W
agen
inge
n U
nive
rsit
y,
The
Net
herl
ands
K
inet
ic M
odel
ing
of A
cryl
amid
e Fo
rmat
ion
(In
prep
arat
ion)
P7
C. Z
onde
rvan
V
oedi
ngsm
idde
len
Tec
hnol
ogie
(2
007,
in p
repa
rati
on)
Acr
ylam
ide
in N
eder
land
se v
oeds
elpr
oduc
ten
P11
Zon
derv
an, C
.; D
e M
ul, A
. V
oedi
ngsm
idde
lent
echn
olog
ie
Acr
ylam
ide
in v
erhi
tte v
oeds
elpr
oduc
ten:
inna
me
en v
oork
omen
van
in
nam
e (f
inal
title
to b
e co
nfir
med
by
publ
ishe
r). [
Acr
ylam
ide
in h
eate
d fo
od p
rodu
cts:
inta
ke a
nd r
educ
tion
of in
take
]
P11
T B
jella
as
The
sis,
Dis
erta
tion
for
the
degr
ee o
f Ph
iloso
phia
e D
octo
r, U
nive
rsit
y of
O
slo,
200
7
Bio
mar
kers
of d
ieta
ry e
xpos
ure
to a
cryl
amid
e P1
6
Wag
enin
gen
Uni
vers
ity,
The
N
ethe
rlan
ds
Publ
icat
ion:
Bi-
/tri
enni
al p
erio
dica
l on
curr
ent r
esea
rch
of th
e gr
adua
te s
choo
l V
LAG
Hig
her
educ
atio
n In
dust
ry
Res
earc
h
P7
Wag
enin
gen
Uni
vers
ity,
The
N
ethe
rlan
ds
Publ
icat
ion
in p
repa
rati
on: J
ourn
al o
f M
olec
ular
Nut
riti
on a
nd F
ood
Res
earc
h
Hig
her
educ
atio
n R
esea
rch
P7/P
1
Wag
enin
gen
Uni
vers
ity,
The
N
ethe
rlan
ds
Subm
itte
d A
bstr
act:
9th I
nter
nati
onal
Sy
mpo
sium
on
the
Mai
llard
Rea
ctio
n H
ighe
r ed
ucat
ion
Indu
stry
R
esea
rch
P7
Page
63 of
77
PO
STE
RS
/ OR
AL
P
RE
SEN
TA
TIO
NS
Yea
r 20
03
Aut
hor
Eve
nt
Tit
le
Par
tner
K
erst
in S
kog
E
xper
ts o
n C
onta
min
ants
In
Food
”,
Wor
ksho
p -
Way
s to
Red
uce
Leve
ls o
f A
cryl
amid
e in
Foo
d, B
russ
els,
20-
21
Oct
ober
200
3
“The
HE
AT
OX
pro
ject
” P1
Ker
stin
Sko
g, I
ngeg
erd
Sjöh
olm
W
orks
hop
on A
cryl
amid
e Fo
rmat
ion
in F
ood,
(E
FSA
), 1
7 N
ovem
ber
2003
, B
russ
els.
“The
HE
AT
OX
pro
ject
”
P1
Lilia
Mas
son
Con
fere
ncia
Soc
ieda
d C
hile
na d
e T
ecno
logí
a de
Alim
ento
s Sa
ntia
go, C
hile
“Acr
ilam
ida
En
Alim
ento
s, ¿
Un
Can
ceri
geno
Pot
enci
al”
y “
The
H
EA
TO
X p
roje
ct”
P23
Year
200
4
Ker
stin
Sko
g M
eeti
ng b
efor
e th
e st
art o
f CO
ST 9
27
acti
on, 2
8 Fe
brua
ry 2
004,
Vie
nna.
“T
he H
EA
TO
X p
roje
ct”
P1
Ker
stin
Sko
g N
etw
ork
mee
ting
“A
kryl
amid
i m
aten
”, L
I (S
wed
ish
Food
Ind
ustr
ies)
“T
he H
EA
TO
X p
roje
ct”
P1
Ker
stin
Sko
g ”L
ivsm
edel
sfor
skar
daga
rna”
, G
othe
nbur
g, 1
7 N
ovem
ber
2004
“T
he H
EA
TO
X p
roje
ct”,
P1
Gun
illa
Vik
lund
Li
vsm
edel
sfor
skar
daga
rna,
Göt
ebor
g,
Swed
en
Acr
ylam
ide
in p
otat
o cr
isps
P1
Bag
dona
ite
Öst
erre
ichi
sche
Leb
ensm
itte
lche
mik
er
Tag
e, 1
1.5.
– 1
2.5.
2004
Fo
rmat
ion
of a
cryl
amid
e in
roa
sted
cof
fee
P2
Ahr
né, A
nder
sson
, Flo
berg
, R
osén
, Lin
gner
t Sw
edis
h Fo
od r
esea
rch
Day
s,
Got
henb
urg,
Sw
eden
E
ffec
t of t
empe
ratu
re a
nd w
ater
con
tent
on
acry
lam
ide
form
atio
n du
ring
bak
ing
of w
hite
bre
ad (
Post
er)
P6
Ling
nert
, Han
s JI
FSA
N A
cryl
amid
e U
pdat
e, C
hica
go,
USA
A
cryl
amid
e in
food
– M
itig
atio
n op
tion
s P6
Ling
nert
, Han
s Le
athe
rhea
d K
ey I
ssue
Day
–
Acr
ylam
ide
Up-
date
, Lea
ther
head
, UK
A
cryl
amid
e in
food
– M
itig
atio
n op
tion
s P6
Ling
nert
, Han
s N
ordi
c St
arch
Net
wor
k M
eeti
ng,
Got
henb
urg,
Sw
eden
A
cryl
amid
e an
d st
arch
– S
tate
of t
he a
rt
P6
Page
64 of
77
Ling
nert
, Han
s C
IAA
acr
ylam
ide
Tec
hnic
al E
xper
t W
orks
hop,
Bru
ssel
s H
EA
TO
X U
pdat
e P6
Ling
nert
, Han
s Sw
edis
h In
dust
ry A
cryl
amid
e N
etw
ork,
M
ay 2
004,
Got
henb
urg
HE
AT
OX
Upd
ate
P6
Ling
nert
, Han
s Sw
edis
h In
dust
ry A
cryl
amid
e N
etw
ork,
O
ctob
er 2
004,
Got
henb
urg
HE
AT
OX
Upd
ate
P6
Ling
nert
, Han
s N
OR
DA
CR
YL
– N
ordi
c N
etw
ork
Proj
ect M
eeti
ng, O
slo,
Nor
way
H
EA
TO
X U
pdat
e P6
Hel
lenä
s, K
.-E
. JR
C-i
rmm
wor
k-sh
op, G
eel,1
7 D
ecem
ber.
T
he H
EA
TO
X p
roje
ct
P9
Dun
ovsk
á L.
, Haj
šlov
á J.
, aj
ka
T.,
Hol
adov
á K
. and
Háj
ková
K.
Che
mic
al R
eact
ions
in F
oods
V.
Prag
ue, C
zech
Rep
ublic
, 29.
thSe
pt -
1.
stO
ct.,
2004
Cha
nges
of A
cryl
amid
e Le
vels
in F
ood
Prod
ucts
dur
ing
Tec
hnol
ogic
al
Proc
essi
ng
P10
J. v
an G
ijsse
l
Pota
to c
onve
ntio
n, J
une
2004
A
mst
erda
m
EU
HE
AT
OX
and
Pot
ato
Proc
essi
ng
P11
E. T
eixi
dó, F
.J. S
anto
s, M
.T.
Gal
cera
n SE
CyT
A (
Mad
rid
2004
) A
naly
sis
of h
ydro
xym
ethy
lfurf
ural
in h
oney
by
gas
chro
mat
ogra
phy
mas
s sp
ectr
omet
ry
P12
Elis
abet
Ber
mud
o, E
ncar
na
Moy
ano,
Lui
s Pu
igno
u, M
aria
T
eres
a G
alce
ran
Reu
nión
Nac
iona
l de
Esp
ectr
omet
ría
de M
asas
LC
-APC
I-M
S/M
S (L
TQ
) fo
r th
e de
term
inat
ion
of a
cryl
amid
e in
fo
odst
uffs
. P1
2
Ele
na B
arce
ló-B
arra
chin
a,
Enc
arna
Moy
ano,
MT
Gal
cera
n
Reu
nión
Nac
iona
l de
Esp
ectr
omet
ría
de M
asas
A
ccur
ate
mas
s m
easu
rem
ent u
sing
a Q
-TO
F fo
r ch
arac
teri
sati
on a
nd
dete
rmin
atio
n of
het
eroc
yclic
am
ines
in fo
od.
P12
H F
rand
sen,
P O
lese
n M
eeti
ng w
ith
Dan
ish
Food
Ind
ustr
y A
cryl
amid
e, r
isk
asse
ssm
ent
P15
Siri
Hel
land
Han
sen
and
Gun
nar
Bru
nbor
g R
epro
duct
ive
Tox
icol
ogy
in th
e A
dult,
R
epro
Safe
Sum
mer
Sch
ool,
Swed
en.
Shor
t pre
sent
atio
n of
the
PhD
pro
ject
- ”
Acr
ylam
ide
in fo
od -
effe
cts o
n re
prod
uctio
n”
P16
H. R
. Gla
tt, W
. Mei
nl, G
. D
obbe
rnac
k, R
. Kol
lock
, U.
Pabe
l, Y
. Som
mer
, et a
l
15th
Int
erna
tiona
l Sym
posiu
m o
n M
icro
som
es a
nd D
rug
Oxi
datio
ns,
Mai
nz, J
uly
2004
Sulp
hotr
ansf
eras
es. (
invi
ted
oral
pre
sent
atio
n)
P18
Y. S
omm
er, H
. Sch
neid
er, H
. R.
Gla
tt
Firs
t Eur
opea
n N
utri
geno
mic
s C
onfe
renc
e, W
agen
inge
n, S
epte
mbe
r 20
04
Haz
ard
iden
tific
atio
n of
the
mut
agen
5-s
ulfo
xym
ethy
l-2-
furf
ural
(S
MF)
form
ed fr
om th
e co
mm
on M
ailla
rd p
rodu
ct 5
-hyd
roxy
met
hyl-
2-fu
rfur
al (
HM
F) b
y in
divi
dual
hum
an s
ulfo
tran
sfer
ases
. (po
ster
)
P18
H. R
. Gla
tt
Sem
inar
s of
Ins
titu
te o
f Pha
rmac
olog
y an
d T
oxic
olog
y, F
acul
ty o
f Vet
erin
ary
Med
icin
e, U
nive
rsit
y of
Gie
ssen
, G
erm
any,
Sep
tem
ber
2004
Sulfo
tran
sfer
asen
als
toxi
fizie
rend
e E
nzym
e [S
ulfo
tran
sfer
ases
as
toxi
fyin
g en
zym
es]
(sem
inar
talk
) P1
8
Page
65 of
77
H. R
. Gla
tt
Cou
rse
on D
rug
Met
abol
ism
, O
rgan
ized
by
the
Ger
man
Soc
iety
of
Tox
icol
ogy,
in D
ortm
und,
Ger
man
y,
Sept
embe
r 20
04
Sulfo
tran
sfer
asen
und
Hyd
rola
sen
(lec
ture
) P1
8
H. R
. Gla
tt
19th
Eur
opea
n W
orks
hop
on D
rug
Met
abol
ism, A
ntal
ya, T
urke
y, O
ctob
er
2004
Act
ivat
ion
of g
enot
oxic
ants
by
cDN
A-e
xpre
ssed
cyt
ochr
omes
P45
0,
alco
hol d
ehyd
roge
nase
s, a
cety
ltran
sfer
ases
and
sul
fotr
ansf
eras
es:
diff
eren
ces
betw
een
rode
nt a
nd h
uman
enz
yme
form
s (l
ectu
re)
P18
H. R
. Gla
tt
Post
grad
uate
stu
dy c
ours
e on
T
oxic
olog
y, U
nive
rsit
y of
Lei
pzig
, G
erm
any,
Dec
embe
r 20
04
Sulfo
tran
sfer
asen
(le
ctur
e)
P18
H. R
. Gla
tt
Sem
inar
s of
Ins
titu
te fo
r Fo
od
Che
mis
try
and
Env
iron
men
tal
Tox
icol
ogy,
Tec
hnic
al U
nive
rsit
y of
K
aise
rsla
uter
n, G
erm
any,
Dec
embe
r 20
04
Hum
anis
iert
e M
odel
lsys
tem
e zu
m E
rfas
sen
gent
oxis
cher
Wir
kung
en
von
Pyro
lyse
prod
ukte
n un
d N
atur
stof
fen
in L
eben
smit
teln
[H
uman
ized
mod
el s
yste
ms
for
the
dete
ctio
n of
gen
otox
ic e
ffec
ts o
f he
at-i
nduc
ed a
nd n
atur
al fo
od c
onst
itue
nts]
(se
min
ar ta
lk)
P18
Lilia
Mas
son
Con
gres
o N
acio
nal d
e T
ecno
logí
a de
A
limen
tos,
Mar
del
Pla
ta, A
rgen
tina
C
urso
de
Frit
ura
en P
rofu
ndid
ad, M
odul
o 4:
For
mac
ión
de A
crila
mid
a y
“The
HE
AT
OX
pro
ject
” P2
3
Lilia
Mas
son
XII
I Se
min
ario
Lat
inoa
mer
ican
o Y
Del
C
arib
e C
ienc
ia Y
Tec
nolo
gía
De
Alim
ento
s, M
onte
vide
o, U
rugu
ay
“Acr
ilam
ida
En
Alim
ento
s, ¿
Un
Can
ceri
geno
Pot
enci
al”
y “
The
H
EA
TO
X p
roje
ct”
P23
Yea
r 20
05
K S
kog
ILSI
Eur
ope,
wor
ksho
p A
cryl
amid
e ta
sk fo
rce
mee
ting
bra
inst
orm
ing
sess
ion
on r
isk
posi
tion
ing
P1
K S
kog
Nor
dic
Min
istr
y C
ounc
il, M
ölle
(S
wed
en),
8 S
epte
mbe
r, 2
005
“The
HE
AT
OX
pro
ject
”
P1
G V
iklu
nd, K
Ols
son,
I
Sjöh
olm
, K S
kog
16th T
rien
nial
Con
fere
nce
of th
e E
APR
E
ffec
t of s
tora
ge o
n ac
ryla
mid
e in
pot
ato
cris
ps fr
om fi
ve S
wed
ish
grow
n po
tato
var
ieti
es
P1
Skog
K
Frui
t, ve
geta
ble
and
pota
to p
roce
ssin
g,
Bru
gge,
Fo
rmat
ion
of a
cryl
amid
e in
cri
sps
P1
Mur
kovi
c M
iniw
orks
hop
at U
nive
rsit
y of
H
elsi
nki
Form
atio
n of
HM
F P2
Page
66 of
77
M. M
urko
vic
1st I
nter
nati
onal
Sym
posi
um o
f the
H
uman
Nut
riti
on &
Met
abol
ism
R
esea
rch
and
Tra
inin
gs C
ente
r, G
raz
Aus
tria
Eff
ects
of h
eati
ng o
n fo
od
P2
Pich
ler,
Mur
kovi
c In
vin
o an
alyt
ica
scie
ncia
200
5, 7
.7. –
9.
7.20
05, M
onte
pelli
er, F
ranc
e D
eter
min
atio
n of
HM
F in
win
es a
nd b
alsa
mic
vin
egar
s by
RP-
HPL
C
P2
Der
ler
K.,
Mur
kovi
c M
.
XV
IIth I
nter
nati
onal
Bot
anic
al
Con
gres
s, 1
7.7.
– 2
3.7.
2005
, Wie
n,
Aus
tria
Det
erm
inat
ion
of m
ono
and
disa
ccha
ride
s in
gre
en c
offe
e an
d th
eir
nutr
itio
nal s
igni
fican
ce
P2
Bag
dona
ite,
Mur
kovi
c 8th
Sym
posi
um o
n In
stru
men
tal
Ana
lysi
s, 2
59. –
28.
9.20
05
3-A
PA a
pos
sibl
e in
term
edia
te in
the
path
way
lead
ing
to a
cryl
amid
e P2
Der
ler,
Mur
kovi
c 8th
Sym
posi
um o
n In
stru
men
tal
Ana
lysi
s, 2
59. –
28.
9.20
05
Ana
lysi
s of
am
ino
acid
s an
d ca
rboh
ydra
tes
in g
reen
cof
fee
P2
Mei
Yin
Low
, Don
ald
S.
Mot
tram
and
J. S
teph
en E
lmor
e 11
th W
eurm
an F
lavo
ur R
esea
rch
Sym
posi
um, R
oski
lde,
Den
mar
k
Rel
atio
nshi
p be
twee
n A
cryl
amid
e Fo
rmat
ion
and
the
Gen
erat
ion
of
Flav
our
in H
eate
d Fo
ods
P3
Åm
an P
. N
I m
eeti
ng in
Ice
land
A
cryl
amid
e in
Bre
ad
P4
Mus
tafa
A.
Food
Sci
ence
Day
in U
ppsa
la
Stra
tegi
es to
Red
uce
Acr
ylam
ide
in B
read
P4
M. D
alla
Ros
a W
orks
hop
on b
read
and
bak
ery
prod
ucts
– V
eron
a (I
taly
) –
Ital
ian
Ass
ocia
tion
in F
ood
Tec
hnol
ogy
(AIT
A)
UE
res
earc
h ac
tivi
ty o
n ac
ryla
mid
e an
d to
xica
nts
in h
eat t
reat
ed fo
ods
P5
S. R
oman
i 7th
Ita
lian
Con
gres
s in
Foo
d Sc
ienc
e an
d T
echn
olog
y (C
ISE
TA
) –
Cer
nobb
io (
Ital
y)
Com
pari
son
amon
g di
ffer
ent f
ryer
s on
som
e qu
alit
y ch
arac
teri
stic
s an
d ac
ryla
mid
e co
nten
t of f
ried
pot
atoe
s P5
Ling
nert
Sw
edis
h In
dust
ry A
cryl
amid
e N
etw
ork,
M
arch
200
5, U
ppsa
la
HE
AT
OX
Upd
ate
P6
Ling
nert
Sa
fe F
ood
for
the
Futu
re; Ö
resu
nd
Food
Net
wor
k, L
und,
Sw
eden
A
cryl
amid
e P6
Ling
nert
N
OR
DA
CR
YL
– N
ordi
c N
etw
ork
Proj
ect M
eeti
ng, R
eykj
avik
, Ice
land
H
EA
TO
X U
pdat
e P6
Ling
nert
Sw
edis
h In
dust
ry A
cryl
amid
e N
etw
ork,
O
ctob
er 2
005,
Got
henb
urg
HE
AT
OX
Upd
ate
P6
Page
67 of
77
Ling
nert
K
SLA
Sem
inar
, Sto
ckho
lm, S
wed
en
Eve
nts
in fo
ods
duri
ng h
eati
ng –
Mai
llard
rea
ctio
n pr
oduc
ts, i
nclu
ding
ac
ryla
mid
e P6
Wag
enin
gen
Uni
vers
ity
Bi-
/tri
enni
al p
erio
dica
l on
curr
ent
rese
arch
of t
he g
radu
ate
scho
ol V
LAG
O
ne-p
age
sum
mar
y of
pro
ject
of p
artn
er 7
P7
J.J.
Kno
l, J.
P.H
. Lin
ssen
and
M
.A.J
.S. v
an B
oeke
l So
crat
es I
P ‘F
ood
& H
ealth
’, In
tern
atio
nal A
dvan
ced
Cou
rse
Tow
ard
a K
inet
ic M
odel
for
Acr
ylam
ide
Form
atio
n in
Mod
el S
yste
ms
(Pos
ter
pres
enta
tion
) P7
J.J.
Kno
l, J.
P.H
. Lin
ssen
and
M
.A.J
.S. v
an B
oeke
l M
eeti
ng w
ith
mem
bers
of D
utch
Po
tato
Pro
cess
ors’
Ass
ocia
tion
(D
PPA
/VA
VI)
Tow
ard
a K
inet
ic M
odel
for
Acr
ylam
ide
Form
atio
n in
Mod
el S
yste
ms
(Pos
ter
pres
enta
tion
) P7
Pete
rsso
n E
.V.,
Ros
én J
., T
urne
r C
., D
anie
lsso
n R
., H
elle
näs
K.-
E.
1st I
nter
nati
onal
Foo
d an
d N
utri
tion
C
ongr
ess,
Ist
anbu
l, T
urke
y, 1
5-18
Ju
ne, 2
005
Cri
tica
l fac
tors
and
pit
falls
aff
ecti
ng th
e ex
trac
tion
of a
cryl
amid
e fr
om
food
s (a
n op
tim
isat
ion
stud
y)
P9
Wag
enin
gen
Uni
vers
ity
Indu
stry
Po
ster
: Mee
ting
wit
h m
embe
rs o
f Dut
ch P
otat
o Pr
oces
sors
’ Ass
ocia
tion
(D
PPA
/VA
VI)
P7
Pete
rsso
n E
.V.,
Ros
én J
., T
urne
r C
., D
anie
lsso
n R
., H
elle
näs
K.E
.
2nd
Inte
rnat
iona
l Sym
posi
um o
n R
ecen
t Adv
ance
s in
Foo
d A
naly
sis,
Pr
ague
, Cze
ch R
epub
lic, 2
-4
Nov
embe
r, 2
005
Iden
tific
atio
n of
cri
tica
l fac
tors
for
extr
acti
on o
f acr
ylam
ide
from
food
s (a
n op
tim
isat
ion
stud
y us
ing
anal
ysis
by
LC-M
S/M
S)
P9
Hel
lenä
s, K
.-E
. SA
NC
O E
xper
t com
mit
tee
on
envi
ronm
enta
l and
indu
stri
al
cont
amin
ants
, Bru
ssel
s, 1
4 Ja
nuar
y.
The
HE
AT
OX
pro
ject
upd
ate.
P9
Hel
lenä
s, K
.-E
. SA
NC
O E
xper
t com
mit
tee
on
envi
ronm
enta
l and
indu
stri
al
cont
amin
ants
, Bru
ssel
s, 9
Jun
e.
Upd
ate
on r
ecen
t HE
AT
OX
act
ivit
ies
on a
cryl
amid
e an
d Fu
ran.
P9
Hel
lenä
s, K
.-E
. B
EU
C m
eeti
ng, B
russ
els,
13
Nov
embe
r.
The
HE
AT
OX
pro
ject
. P9
Hel
lenä
s, K
.-E
. Fo
od s
afet
y in
a E
urop
ean
pers
pect
ive,
Se
min
ar a
nd w
ork-
shop
, Osl
o, 7
-8
Dec
embe
r
Acr
ylam
ide
– St
ill a
mat
ter
of c
once
rn?
P9
Gib
ala
P., N
ovot
ný O
., Pe
ters
son
E.V
., D
unov
ská
L.,
Vác
lavi
k L.
Haj
slov
á J.
, Hel
lenä
s K
.-E
., R
osén
J.
2nd
Inte
rnat
iona
l Sym
posi
um o
n R
ecen
t Adv
ance
s in
Foo
d A
naly
sis,
Pr
ague
, Cze
ch R
epub
lic, 2
-4
Nov
embe
r, 2
005
Can
alk
alin
e ex
trac
tion
rel
ease
mas
ked
acry
lam
ide?
P9
P1
0
Page
68 of
77
Gib
ala
P., P
eter
sson
E.,
Dun
ovsk
á L.
, Vác
laví
k L.
, N
ovot
ný O
., H
ajšl
ová
J.,
Hel
lenä
s K
. E.,
Ros
én J
.
2nd I
nter
nati
onal
sym
posi
um o
n re
cent
ad
vanc
es in
food
ana
lysi
s, P
ragu
e,
Cze
ch R
epub
lic, 2
-4 N
ovem
ber,
200
5
Can
alk
alin
e ex
trac
tion
rel
ease
mas
ked
acry
lam
ide?
P1
0
Dun
ovsk
á L.
, aj
ka T
. and
H
ajšl
ová
J.
3rd M
eeti
ng o
n C
hem
istr
y &
Life
Brn
o,
Cze
ch R
epub
lic, 2
0-22
Sep
tem
ber,
20
05
Acr
ylam
ide
Leve
ls in
Foo
dstu
ffs
from
Cze
ch m
arke
t P1
0
Dun
ovsk
á L.
, Haj
šlov
á J.
, H
olad
ová
K.,
Gib
ala
P., S
chre
k J.
2nd I
nter
nati
onal
sym
posi
um o
n re
cent
ad
vanc
es in
food
ana
lysi
s, P
ragu
e,
Cze
ch R
epub
lic, 2
-4 N
ovem
ber,
200
5
Ana
lyti
cal s
trat
egie
s fo
r ex
amin
atio
n of
roa
sted
cof
fee:
vol
atile
mar
kers
of
acr
ylam
ide
form
atio
n P1
0
J. v
an G
ijsse
l
Acr
ylam
ide
stak
ehol
ders
mee
ting
, 14
Janu
ary
2005
, Bru
ssel
H
EA
TO
X W
P 2.
For
mat
ion
and
Proc
essi
ng, P
rogr
ess
in th
e fir
st y
ear.
P1
1
E. B
erm
udo,
O. N
úñez
, L.
Puig
nou,
M.T
. Gal
cera
n X
I Jo
rnad
as d
e A
nális
is I
nstr
umen
tal
(Bar
celo
na 2
005)
D
evel
opm
ent o
f new
cap
illar
y el
ectr
opho
resi
s m
etho
dolo
gies
for
the
anal
ysis
of a
cryl
amid
e in
food
stuf
fs
P12
E. T
eixi
dó, F
.J. S
anto
s, M
.T.
Gal
cera
n X
I Jo
rnad
as d
e A
nális
is I
nstr
umen
tal
(Bar
celo
na 2
005)
G
as c
hrom
atog
raph
y co
uple
d to
mas
s sp
ectr
omet
ry fo
r th
e an
alys
is o
f hy
drox
ymet
hylfu
rfur
al in
food
s P1
2
M. S
. Alta
ki, F
.J. S
anto
s, M
.T.
Gal
cera
n X
I Jo
rnad
as d
e A
nális
is I
nstr
umen
tal
(Bar
celo
na 2
005)
A
naly
sis
of fu
ran
in fo
od b
y he
adsp
ace
solid
-pha
se m
icro
extr
acti
on
coup
led
wit
h ga
s ch
rom
atog
raph
y-m
ass
spec
trom
etry
. P1
2
Elis
abet
Ber
mud
o, O
scar
Núñ
ez,
Luis
Pui
gnou
, Mar
ia T
eres
a G
alce
ran
HPL
C 2
005
Cap
illar
y Z
one
Ele
ctro
phor
esis
for
the
dete
rmin
atio
n of
acr
ylam
ide
in
food
stuf
fs
P12
Ele
na B
arce
ló-B
arra
chin
a,
Enc
arna
ción
Moy
ano,
Llu
ís
Puig
nou
and
Mar
ia T
eres
a G
alce
ran
HPL
C 2
005
Dev
elop
men
t of m
etha
cryl
ate
mon
olit
hs fo
r th
e an
alys
is o
f het
eroc
yclic
am
ines
by
capi
llary
ele
ctro
chro
mat
ogra
phy
P12
Mar
ia T
eres
a G
alce
ran
1st
Int
erna
tion
al W
orks
hop
of M
S/M
S sp
ectr
omet
ry
Tan
dem
Mas
s Sp
ectr
omet
ry a
nd A
ccur
ate
Mas
s M
easu
rem
ent f
or th
e an
alys
is o
f gen
otox
ic h
eat-
indu
ced
com
poun
ds in
food
. P1
2
Hül
ya Ö
lmez
T
ÜB
TA
K M
RC
1st
Int
erna
tion
al
Food
and
Nut
riti
on C
ongr
ess-
15-1
8 Ju
ne 2
005,
Ist
anbu
l / O
ral
pres
enta
tion
Gıd
alar
da A
krila
mid
(A
cryl
amid
e in
food
s)
P13
Hül
ya Ö
lmez
T
V P
rogr
amm
e -
AT
V C
hann
el /
sp
eake
r A
cryl
amid
e ri
sk in
food
s P1
3
Page
69 of
77
Hül
ya Ö
lmez
T
V P
rogr
amm
e –
CN
BC
-e C
hann
el /
sp
eake
r A
cryl
amid
e ri
sk in
food
s P1
3
P O
lese
n O
ral p
rese
ntat
ion
inte
rnal
ly in
DFV
F A
kryl
amid
– a
naly
se a
f glo
bina
dduk
ter
I hu
man
e pr
øver
(pr
ojec
t H
EA
TO
X)
P15
Tho
mas
Bje
llaas
N
orw
egia
n So
ciet
y fo
r M
ass
Spec
trom
etry
, Haf
jell
Janu
ary
2005
Po
ster
– “
Det
erm
inat
ion
of a
cryl
amid
e m
etab
olit
es in
hum
an u
rine
us
ing
solid
-pha
se e
xtra
ctio
n an
d LC
-MS/
MS
dete
ctio
n”
P16
Tho
mas
Bje
llaas
, Heg
e B
. Ø
lstø
rn, J
an E
rik
Paul
sen
Nor
dic
mee
ting
for
NO
RD
AC
RY
L –
june
, Ice
land
Pr
esen
tati
on o
f bio
avai
labi
lity
of a
cryl
amid
e fr
om fo
ods
stud
y in
mic
e us
ing
met
abol
ic c
ages
P1
6
Tho
mas
Bje
llaas
E
urop
ean
mee
ting
for
HE
AT
OX
–
(Net
herl
ands
), M
ay 2
005
Post
er –
“D
eter
min
atio
n of
acr
ylam
ide
met
abol
ites
in h
uman
uri
ne
usin
g so
lid-p
hase
ext
ract
ion
and
LC-M
S/M
S de
tect
ion”
P1
6
Tho
mas
Bje
llaas
N
orw
egia
n In
stit
ute
of P
ublic
Hea
lth
Pres
enta
tion
– A
pilo
t stu
dy o
n ur
inar
y ex
cret
ion
acry
lam
ide
met
abol
ites
in h
uman
s P1
6
Heg
e B
. A. Ø
lstø
rn, J
an E
rik
Paul
sen,
Jan
Ale
xand
er
Nor
weg
ian
Inst
itut
e of
Pub
lic H
ealth
Pr
esen
tati
on o
f pre
limin
ary
resu
lts –
Acr
ylam
ide
and
inte
stin
al c
ance
r P1
6
Siri
Hel
land
Han
sen,
Eri
k Sø
derl
und,
Ado
lf B
aum
gart
ner,
D
And
erso
n an
d G
Bru
nbor
g
3rd C
open
hage
n W
orks
hop
on
Env
iron
men
t, R
epro
duct
ive
Hea
lth a
nd
Fert
ility
, 15-
18 J
anua
ry 2
005
Post
er a
nd a
bstr
act –
“T
he p
ater
nal r
epro
duct
ive
toxi
city
of t
he fo
od
cont
amin
ant a
cryl
amid
e P1
6
Siri
Hel
land
Han
sen
and
Gun
nar
Bru
nbor
g N
orw
egia
n In
stit
ute
of P
ublic
Hea
lth,
Div
isio
n of
Env
iron
men
tal M
edic
ine
Pres
enta
tion
(in
Nor
weg
ian)
– “
Kan
akr
ylam
id p
åvir
ke d
en m
annl
ige
repr
oduk
sjon
en?”
P1
6
Jan
Ale
xand
er
9th I
nter
nati
onal
Con
fere
nce
on
Env
iron
men
tal M
utag
ens,
San
Fr
anci
sco,
Sep
t. 20
05
Ris
k A
sses
smen
t of A
cryl
amid
e in
Foo
d P1
6
Jaco
b D
. van
Kla
vere
n H
eato
x 18
th m
onth
mee
ting
, Sy
mpo
sium
wit
h D
utch
pot
ato
proc
esso
rs A
ssoc
iati
on
Exp
osur
e as
sess
men
t of H
EA
TO
X c
ompo
unds
; P1
7
Jaco
b D
. van
Kla
vere
n, A
nika
de
Mul
D
utch
Foo
d A
utho
rity
(V
WA
) w
ork-
mee
ting
E
xpos
ure
asse
ssm
ent o
f hea
t-tr
eate
d to
xica
nts
P17
Ani
ka d
e M
ul
Food
Sta
ndar
d A
genc
y (U
K)
, wor
k-m
eeti
ng
Prob
abili
stic
acr
ylam
ide
inta
ke c
alcu
lati
ons
wit
h M
CR
A a
nd
Com
pari
son
of a
cryl
amid
e in
take
by
Bri
tish
and
Dut
ch to
ddle
rs
P17
H. R
. Gla
tt
Sym
posi
um o
f Sem
inar
s of
B
unde
sins
titu
t für
Ris
ikob
ewer
tung
([
Ger
man
] Fe
dera
l Ins
titu
te o
f Ris
k A
sses
smen
t), F
ebru
ary
2005
Gen
toxi
zitä
ts-U
nter
such
unge
n in
vitr
o m
it v
ersc
hied
enen
K
ohle
nhyd
rat-
Pyro
lyse
prod
ukte
n [G
enot
oxic
ity
stud
ies
in v
itro
on
vari
ous
carb
ohyd
rate
pyr
olys
is p
rodu
cts]
(ta
lk)
P18
Page
70 of
77
H. R
. Gla
tt
Ann
ual M
eeti
ng o
f the
“A
rbei
tskr
eis
Ges
undh
eit
und
Lebe
nsm
itte
lsic
herh
eit”
[w
ork
grou
p H
ealth
and
Foo
d Sa
fety
] of
the
“Bun
desv
erba
ndes
der
Deu
tsch
en
Süßw
aren
indu
stri
e e.
V.”
[Fe
dera
l A
ssoc
iati
on o
f the
Ger
man
Ind
ustr
y on
Sw
eet P
rodu
cts]
, Ber
ghol
z-R
ehbr
ücke
, G
erm
any,
Jun
e 20
05
Hea
tox:
G
ento
xizi
täts
-Unt
ersu
chun
gen
mit
ve
rsch
iede
nen
Koh
lenh
ydra
t-Py
roly
sepr
oduk
ten
[Hea
tox:
G
enot
oxic
ity
stud
ies
on
vari
ous
carb
ohyd
rate
pyr
olys
is p
rodu
cts]
(ta
lk)
P18
H. R
. Gla
tt
Sem
inar
s of
the
Inst
itut
e fo
r C
ance
r R
esea
rch,
Uni
vers
ity
of V
ienn
a,
Aus
tria
, Jun
e 20
05
Gen
toxi
sche
und
kan
zero
gene
Wir
kung
en v
on L
eben
smit
tel-
Inha
ltsst
offe
n in
hum
anis
iert
en M
odel
lsys
tem
en u
nd d
eren
Pr
även
tion
/Pot
enzi
erun
g du
rch
wei
tere
Ern
ähru
ngsf
akto
ren
[Gen
otox
ic a
nd c
arci
noge
nic
effe
cts
of fo
od-b
orne
sub
stan
ces
in
hum
aniz
ed m
odel
sys
tem
s, a
nd th
eir
prev
enti
on/p
oten
tiat
ion
by o
ther
fo
od fa
ctor
s] (
sem
inar
talk
)
P18
H. R
. Gla
tt
Post
grad
uate
stu
dy c
ours
e on
T
oxic
olog
y, U
nive
rsit
y of
Vie
nna,
A
ustr
ia, J
une
2005
Gen
etis
ch v
erän
dert
e Z
elle
n un
d T
iere
zur
Unt
ersu
chun
g de
s Fr
emds
toff
-Met
abol
ism
us [
Gen
etic
ally
eng
inee
red
cells
and
ani
mal
s fo
r in
vest
igat
ions
on
xeno
biot
ic m
etab
olis
m]
(lec
ture
)
P18
H. R
. Gla
tt
Con
tinu
ous
Edu
cati
on in
Tox
icol
ogy,
Sy
mpo
sium
„R
ücks
tand
stox
ikol
ogie
, ne
ue A
spek
te u
nd E
ntw
ickl
unge
n 20
05“
Uni
vers
ity
of Z
uric
h,
Swit
zerl
and,
Nov
embe
r 20
05
Gen
otox
isch
e Py
roly
sepr
oduk
te u
nd N
atur
stof
fe in
Leb
ensm
itte
ln
[Gen
otox
c py
roly
sis
prod
ucts
and
nat
ural
com
poun
ds in
food
s]
(lec
ture
)
P18
Bar
bara
Gal
lani
B
EU
C F
ood
offic
ers’
mee
ting
29
-30
Nov
embe
r 20
05
Acr
ylam
ide:
reg
ulat
ory
appr
oach
and
com
mun
icat
ion
to c
onsu
mer
s P2
0
Hel
ga O
dden
Rek
snes
Fo
od p
ract
ices
at h
ome;
wha
t are
the
risk
s?
Wor
ksho
ps I
rela
nd a
nd N
orth
ern
Irel
and
17 a
nd 1
8 M
ay 2
005
The
rol
e of
ris
k co
mm
unic
atio
n po
licy
in E
urop
e in
ass
urin
g fo
od
safe
ty to
con
sum
ers
(man
uscr
ipt a
nd p
pt-p
rese
ntat
ion)
P21
Hel
ga O
dden
Rek
snes
D
G S
anco
, CC
AB
mee
ting
H
EA
TO
X –
Del
iver
able
s –
Rsi
k C
omm
unic
atio
n an
d di
ssem
inat
ion
(ppt
-pre
sent
atio
n)
P21
Han
spet
er N
aege
li C
ours
e in
Tox
icol
ogy
Swis
s Fe
dera
l Ins
titu
te o
f Tec
hnol
ogy
The
app
licat
ion
of “
-om
ic”
tech
nolo
gies
in fo
od to
xico
logy
P2
2
Flur
ina
Cle
men
t In
tern
atio
nal S
ympo
sium
of
Phar
mac
olog
y an
d T
oxic
olog
y. V
ienn
a G
ene
expr
essi
on p
rofil
es in
duce
d by
gly
cida
mid
e in
hum
an c
ells
P2
2
Page
71 of
77
Flur
ina
Cle
men
t A
nnua
l Mee
ting
of t
he S
wis
s T
oxic
olog
y N
etw
ork
(XE
RR
) G
ene
expr
essi
on p
rofil
es in
duce
d by
gly
cida
mid
e in
hum
an c
ells
P2
2
Flur
ina
Cle
men
t A
nnua
l Mee
ting
of t
he X
enob
ioti
c an
d E
nvir
onm
enta
l Ris
k R
esea
rch
Cen
ter
Zür
ich,
Sw
itze
rlan
d
Tra
nscr
ipto
mic
s of
Acr
ylam
ide
and
Gly
cida
mid
e P2
2
Lilia
Mas
son
VI
Sem
inar
io L
atin
oam
eric
ano
de
Cie
ncia
de
Alim
ento
s, C
ampi
nas,
B
rasi
l
“Acr
ilam
ida
En
Alim
ento
s, ¿
Un
Can
ceri
geno
Pot
enci
al”
y “
The
H
EA
TO
X p
roje
ct”
P23
Lilia
Mas
son
Con
gres
o Fe
dera
ción
Far
mac
éuti
ca
Suda
mer
ican
a, S
anti
ago,
Chi
le
“Acr
ilam
ida
En
Alim
ento
s, ¿
Un
Can
ceri
geno
Pot
enci
al”
y “
The
H
EA
TO
X p
roje
ct”
P23
Year
200
6
Skog
K
Env
iron
men
tal a
nd I
ndus
tria
l C
onta
min
ants
wor
ksho
p- a
cryl
amid
e,
(CIA
A)
Bru
ssel
s
The
HE
AT
OX
pro
ject
- H
eat –
gen
erat
ed F
ood
Tox
ican
ts.
Iden
tific
atio
n, C
hara
cter
izat
ion
and
Ris
k M
inim
isat
ion
Skog
K
HE
AT
OX
wor
ksho
p, G
raz
Hom
e-co
okin
g an
d ac
ryla
mid
e
P1
Skog
K
Wor
ksho
p at
Nes
tlé, L
ausa
nne
The
HE
AT
OX
pro
ject
P1
Skog
K
Form
as v
isit
to L
U
Bild
ning
av
toxi
ska
ämne
n vi
d liv
smed
elsp
roce
ssni
ng
Bild
ning
smek
anis
mer
och
häl
soef
fekt
er
P1
G V
iklu
nd, K
Ols
son,
I
Sjöh
olm
, K S
kog
Livs
med
elsf
orsk
arda
garn
a, N
ovem
ber
21-
22, 2
006,
Upp
sala
, Sw
eden
A
cryl
amid
e in
cri
sps
from
Sw
edis
h gr
own
Satu
rna
pota
toes
P1
G V
iklu
nd, K
Ols
son,
I
Sjöh
olm
, K S
kog
Mee
ting
NO
RD
AC
RY
L E
ffec
t of s
tora
ge o
n ac
ryla
mid
e in
pot
ato
cris
ps fr
om fi
ve S
wed
ish
grow
n po
tato
var
ieti
es
P1
Bag
dona
ite
K, M
urko
vic
M
Ö
ster
reic
hisc
he L
eben
smit
telc
hem
iker
T
age,
12.
9. –
14.
9.20
06, W
ien
Acr
ylam
ide
form
atio
n in
cof
fee
P2
Bor
nik
MA
, Jöb
stl D
, Der
ler
K,
Mur
kovi
c M
Öst
erre
ichi
sche
Leb
ensm
itte
lche
mik
er
Tag
e, 1
2.9.
– 1
4.9.
2006
, Wie
n K
inet
ik d
er H
MF-
und
HM
FS-B
ildun
g im
Lau
fe d
er K
affe
erös
tung
, in
Mod
ells
yste
men
P2
Per
Åm
an
Arr
ange
d a
mee
ting
in c
olla
bora
tion
w
ith
Nor
dic
Inno
vati
on C
entr
e (N
ICe
Mee
ting
NO
RD
AC
RY
L)
Acr
ylam
ide
P4
Page
72 of
77
Arw
a M
usta
fa
NIC
e M
eeti
ng N
OR
DA
CR
YL
Ana
lysi
s of
Fre
e A
min
o A
cids
P4
Arw
a M
usta
fa
Lect
ure
at S
udan
Uni
vers
ity,
K
hart
oum
, Sud
an
Acr
ylam
ide
in F
ood
Prod
ucts
P4
Arw
a M
usta
fa
Lect
ure
at A
hfad
Uni
vers
ity,
O
mdu
rman
, Sud
an
Acr
ylam
ide
in F
ood
Prod
ucts
P4
M. D
alla
Ros
a C
IAA
Bru
ssel
les
Mee
ting
H
eat –
gen
erat
ed fo
od to
xica
nts.
Ide
ntifi
cati
on, c
hara
cter
izat
ion
and
risk
min
imis
atio
n. H
ome
cook
ing
P5
S. R
oman
i, M
. Bac
chio
cca,
P.
Roc
culi,
A. A
ngio
loni
, M. D
alla
R
osa
CE
Food
Con
gres
s: 3
th C
entr
al
Eur
opea
n C
ongr
ess
on F
ood.
(So
fia -
B
ulga
ria)
Eff
ect o
f fry
ing
tim
e on
som
e qu
alit
y as
pect
s an
d ac
ryla
mid
e co
nten
t of
Fren
ch f
ries
P5
S. R
oman
i, M
. Bac
chio
cca,
P.
Roc
culi,
M. D
alla
Ros
a IU
FOST
, 13th
Wor
ld C
ongr
ess
of
Food
Sci
ence
and
Tec
hnol
ogy
(Nan
tes
– Fr
ance
)
Eff
ect o
f som
e fr
ying
con
diti
ons
on a
cryl
amid
e co
nten
t in
fren
ch fr
ies
P5
Ling
nert
, Han
s Sw
edis
h In
dust
ry A
cryl
amid
e N
etw
ork,
M
arch
200
6, U
ppsa
la
HE
AT
OX
Upd
ate
P6
Ling
nert
, Han
s H
EA
TO
X W
orks
hop,
Jun
e 20
06,
Gra
z, A
ustr
ia
Min
imis
atio
n O
ptio
ns -
Ind
ustr
y P6
Ling
nert
, Han
s
CIA
A A
cryl
amid
e G
roup
, 5 J
uly
2006
, B
russ
els,
Bel
gum
H
EA
TO
X d
eliv
erab
les
in r
elat
ion
to th
e C
IAA
Too
lbox
P6
Ling
nert
, Han
s Sw
edis
h In
dust
ry A
cryl
amid
e N
etw
ork,
O
ctob
er 2
006,
Got
henb
urg
HE
AT
OX
Upd
ate
P6
Hel
lenä
s, K
.-E
. SA
NC
O/C
IAA
Wor
ksho
p, B
russ
els,
16
Mar
ch.
Acr
ylam
ide
extr
acti
on a
t hig
h pH
. P9
Ling
nert
, Han
s Sw
edis
h Fo
od R
esea
rch
Day
s, U
ppsa
la,
21-2
2 N
ovem
ber
2006
A
kryl
amid
– p
å go
tt o
ch o
nt
Vác
laví
k L.
, Bar
táko
vá V
., H
olad
ová
K.,
Haj
šlov
á J.
JA
KO
ST O
BIL
OV
IN -
200
6 (Q
ualit
y of
cer
eals
-200
6),
Kro
míž
, C
zech
Rep
ublic
, 9.1
1.20
06
Acr
ylam
ide
– ne
w fo
od c
arci
noge
n P1
0
Haj
šlov
á J.
, Nov
otný
O.,
Vác
laví
k L,
Dun
ovsk
á L.
C
OST
IM
AR
S Jo
int W
orks
hop,
N
apol
i, It
aly,
24.
- 2
7. 5
. 200
6 St
udy
of "
mas
ked"
acr
ylam
ide
orig
in
P10
Vác
laví
k L,
aj
ka T
., D
unov
ská
L., H
ajšl
ová
J.
CO
ST I
MA
RS
Join
t Wor
ksho
p,
Nap
oli,
Ital
y, 2
4. -
27.
5. 2
006
LC-M
S/M
S an
d G
C/H
RT
OF-
MS
met
hods
for
anal
ysis
of a
cryl
amid
e in
food
stuf
fs
P10
Vác
laví
k L.
, Bar
táko
vá V
., H
olad
ová
K.,
Haj
šlov
á J.
JA
KO
ST O
BIL
OV
IN -
200
6 (Q
ualit
y of
cer
eals
-200
6),
A
cryl
amid
e –
new
food
car
cino
gen
P10
Page
73 of
77
Zon
derv
an, C
. W
orks
hop
wit
h D
PPA
N
ov 2
8, 2
006
Inst
ruct
ions
on
usin
g th
e fr
ench
fry
mod
ellin
g to
ol: F
ry S
imul
ator
v.2
P1
1
E. T
eixi
dó, E
. Moy
ano,
F.J
. Sa
ntos
, M.T
. Gal
cera
n II
I R
euni
ón d
e la
Soc
ieda
d E
spañ
ola
de
Esp
ectr
omet
ría
de M
asas
(O
vied
o 20
06)
Ana
lysi
s of
hyd
roxy
met
hylfu
rfur
al in
food
stuf
fs b
y liq
uid
chro
mat
ogra
phy
coup
led
to ta
ndem
mas
s sp
ectr
omet
ry
P12
L. P
uign
ou
Jorn
ada
de G
enot
òxic
s en
Alim
ents
(R
eus
2006
) T
óxic
os a
limen
tari
os e
ndog
énic
os e
n al
imen
tos
proc
esad
os
térm
icam
ente
P1
2
E. B
erm
udo
Jorn
ada
de G
enot
òxic
s en
Alim
ents
(R
eus
2006
) A
nális
is d
e ac
rila
mid
a en
alim
ento
s P1
2
E. T
eixi
dó
Jorn
ada
de G
enot
òxic
s en
Alim
ents
(R
eus
2006
) A
nális
is d
e hi
drox
imet
ilfur
fura
l med
iant
e té
cnic
as c
rom
atog
ráfic
as y
es
pect
rom
etrí
a de
mas
as
P12
M. S
. Alta
ki
Jorn
ada
de G
enot
òxic
s en
Alim
ents
(R
eus
2006
) Fu
ran:
For
mat
ion
and
anal
ysis
P1
2
E. B
erm
udo,
O. N
úñez
, E.
Moy
ano,
L. P
uign
ou, M
.T.
Gal
cera
n
SEC
yTA
(V
igo
2006
) A
pplic
atio
n of
ele
ctro
phor
etic
tech
niqu
es fo
r th
e de
term
inat
ion
of
acry
lam
ide
in fo
ods
P1
2
M. S
. Alta
ki, F
.J. S
anto
s, M
.T.
Gal
cera
n SE
CyT
A (
Vig
o 20
06)
Ana
lysi
s of
fura
n in
food
by
on-l
ine
head
spac
e so
lid-p
hase
m
icro
extr
acti
on a
nd g
as c
hrom
atog
raph
y io
n tr
ap m
ass
spec
trom
etry
. P1
2
A V
ikst
röm
, S E
riks
son,
B
Paul
sson
, P K
arls
son
and
M
Tör
nqvi
st
Pres
enta
tion
at Å
bo A
cade
my
Stud
y of
rel
ativ
e bi
oava
ilabi
lity
of a
cryl
amid
e in
diff
eren
t foo
dstu
ffs
P14
A V
ikst
röm
Se
min
ar a
t Sto
ckho
lm U
nive
rsit
y B
ackg
roun
d ex
posu
re to
acr
ylam
ide
and
its
geno
toxi
c m
etab
olit
e gl
ycid
amid
e P1
4
A V
ikst
röm
, B P
auls
son,
& M
T
örnq
vist
C
ours
e at
Kar
olin
ska
Inst
itut
et,
Soln
a,Sw
eden
(E
CN
IS),
pos
ter
pres
enta
tion
Poss
ible
influ
ence
of p
olym
orph
ism
on
inte
rnal
dos
e of
acr
ylam
ide
and
glyc
idam
ide
P14
M T
örnq
vist
H
EA
TO
X w
orks
hop
13-1
4 Ju
ne
The
acr
ylam
ide
stor
y P1
4 A
nika
de
Mul
IL
SI, E
xper
t gro
up m
eeti
ng ‘R
isk
Ben
efit
Ana
lysi
s of
Mit
igat
ion
Mea
sure
s’
Acr
ylam
ide;
Exp
osur
e an
d R
educ
tion
sce
nari
os
P17
Jaco
b D
. van
Kla
vere
n N
orw
egai
n Fo
od A
utho
rity
(V
KM
) Pr
obab
ilist
ic m
odel
ing
in p
ersp
ecti
ve o
f fut
ure
risk
ass
essm
ent
P17
Jaco
b D
. van
Kla
vere
n N
aam
van
het
sym
posi
um. T
he
HE
AT
OX
Wor
ksho
p
Exp
osur
e an
d R
educ
tion
Sce
nari
os
P17
Page
74 of
77
Jaco
b D
. van
Kla
vere
n T
elev
isio
n pr
ogra
mm
e fo
r co
nsum
ers
(Kas
sa)
Acr
ylam
ide
leve
ls in
food
and
its
risk
s P1
7
H. R
. Gla
tt
Con
tinu
ous
Edu
cati
on fo
r M
edic
al
Doc
tors
, Cou
rse
“Ern
ähru
ngs-
med
izin
”, P
otsd
am-R
ehbr
ücke
, G
erm
any,
Mar
ch 2
006
Kre
bser
zeug
ende
Sto
ffe
in L
eben
smit
teln
[Fo
od-b
orne
car
cino
gens
] (l
ectu
re)
P18
H. R
. Gla
tt
Cou
rse
on D
rug
Met
abol
ism
, O
rgan
ized
by
the
Ger
man
Soc
iety
of
Tox
icol
ogy,
in D
ortm
und,
Ger
man
y,
Sept
embe
r 20
06
Sulfo
tran
sfer
asen
und
Hyd
rola
sen
(lec
ture
) P1
8
H. R
. Gla
tt, Y
. Som
mer
, G.
Dob
bern
ack,
W. M
einl
43
rd C
ongr
ess o
f the
Eur
opea
n So
ciet
ies
of T
oxic
olog
y an
d 6t
h C
ongr
ess o
f T
oxic
olog
y in
Dev
elop
ing
Cou
ntri
es,
Cav
tat/
Dub
rovn
ik, C
roat
ia, S
epte
mbe
r
Het
erol
ogou
s an
d tr
ansg
enic
mod
els
for
stud
ying
gen
otox
ic e
ffec
ts o
f co
ntam
inan
ts p
rodu
ced
by h
eat-
trea
tmen
t of f
ood
(inv
ited
talk
),
Abs
trac
t pub
lishe
d in
: Tox
icol
. Let
t. 16
4 S
(200
6) 6
2-63
.
P18
Jaco
b D
. van
Kla
vere
n T
rain
ing
for
Nor
weg
ian
Food
A
utho
rity
(V
KM
) Pr
obab
ilist
ic d
ieta
ry in
take
cal
cula
tion
s fo
r ac
ryla
mid
e P1
7
H. R
. Gla
tt, F
. Tau
gner
, S.
Flor
ian,
Y. S
omm
er
CO
ST 9
26/9
27 C
onfe
renc
e:
Inte
rnat
iona
l Con
fere
nce
on M
olec
ular
an
d Ph
ysio
logi
cal E
ffect
s of B
ioac
tive
Food
Com
pone
nts,
Vie
nna,
Aus
tria
, O
ctob
er 2
006
Hea
lth r
isks
by
5-hy
drox
ymet
hylfu
rfur
al (
HM
F) a
nd r
elat
ed
com
poun
ds (
invi
ted
talk
), S
ome
data
acc
essi
ble
at C
OST
926
web
pa
ge: h
ttp:
//w
ww
.uoc
hb.c
as.c
z/Z
prav
y/C
OST
_926
/
P18
H. R
. Gla
tt, G
. Dob
bern
ack,
W.
Mei
nl
CO
ST 9
26/9
27 C
onfe
renc
e:
Inte
rnat
iona
l Con
fere
nce
on M
olec
ular
an
d Ph
ysio
logi
cal E
ffect
s of B
ioac
tive
Food
Com
pone
nts,
Vie
nna,
Aus
tria
, O
ctob
er 2
006
Exp
ress
ion
of h
uman
pha
se-I
I en
zym
es in
mod
el s
yste
ms
for
dete
ctin
g ge
noto
xica
nts
and
prot
ecti
ve a
gent
s (i
nvit
ed ta
lk),
Som
e da
ta a
cces
sibl
e at
CO
ST92
6 w
eb p
age:
htt
p://
ww
w.u
ochb
.cas
.cz/
Zpr
avy/
CO
ST_9
26/
P18
Bar
bara
Gal
lani
C
IAA
/EC
Wor
ksho
p 16
-17
Mar
ch 2
006
Acr
ylam
ide:
info
rmat
ion
and
com
mun
icat
ion
to c
onsu
mer
s P2
0
Hel
ga O
dden
Rek
snes
A
cryl
amid
e W
orks
hop
EU
/CIA
A
Min
utes
from
wor
king
gro
up o
n ho
me
cook
ing
P21
Hel
ga O
dden
Rek
snes
C
CFA
C W
orks
hop
on R
isk
com
mun
icat
ion
as a
man
agem
ent t
ool
for
cont
amin
ants
Con
sum
ptio
n A
dvic
e on
Con
tam
inan
ts
- a
brie
f com
mun
icat
ion
case
stu
dy o
n A
cryl
amid
e
P21
Wor
king
Gro
up, H
EA
TO
X
Wor
ksho
p
The
HE
AT
OX
Wor
ksho
p R
epor
t P2
1
Page
75 of
77
Hel
ga O
dden
Rek
snes
T
he H
EA
TO
X W
orks
hop
The
HE
AT
OX
Aqu
ariu
m
P21
Hel
ga O
dden
Rek
snes
T
he H
EA
TO
X 3
0 M
mee
ting
Gra
z Su
mm
ary
Wor
ksho
p G
raz
P21
Flur
ina
Cle
men
t M
eeti
ng o
f the
DN
A R
epai
r-R
eplic
atio
n-R
ecom
bina
tion
Net
wor
k Z
üric
h, S
wit
zerl
and
Tra
nscr
ipto
mic
s of
Gly
cida
mid
e, th
e G
enot
oxic
Met
abol
ite
of
Acr
ylam
ide
P22
Year
200
7
G V
iklu
nd, K
Ols
son,
I
Sjöh
olm
, K S
kog
4th E
APR
/FN
K/U
EIT
P Po
tato
Pr
oces
sing
Con
fere
nce
and
EA
PR
Eng
inee
ring
& U
tilis
atio
n Se
ctio
n M
eeti
ng, J
anua
ry 1
7, 2
007,
Gra
ntha
m,
UK
Acr
ylam
ide
in c
risp
s fr
om S
wed
ish
grow
n Sa
turn
a po
tato
es
P1
G V
iklu
nd, K
Ols
son,
I
Sjöh
olm
, K S
kog
Firs
t Eur
opea
n W
orks
hop
on F
ood
Eng
inee
ring
and
Tec
hnol
ogy,
EFF
oST
E
FChE
Foo
d W
orki
ng P
arty
Con
gres
s,
May
200
7, B
erlin
, Ger
man
y
Acr
ylam
ide
in p
otat
o cr
isps
from
Sw
edis
h gr
own
pota
toes
P1
G V
iklu
nd, K
Ols
son,
I
Sjöh
olm
, K S
kog
CO
ST 9
27 A
ctio
n W
orks
hop,
Sof
ia,
Bul
gari
a A
cryl
amid
e in
roa
sted
pot
ato
wed
ges
P1
Skog
K
Sym
posi
um o
n C
hem
istr
y an
d T
oxic
olog
y of
Acr
ylam
ide,
Bos
ton,
US,
A
ugus
t, 20
07.
Hea
t der
ived
toxi
cant
s in
food
– s
ome
findi
ngs
of a
col
labo
rati
ve
Eur
opea
n re
sear
ch p
roje
ct
P1
Mur
kovi
c, M
. 5th
Int
erna
tion
al C
ongr
ess
on F
ood
Tec
hnol
ogy,
9.3
.-11
.3.2
007
The
ssal
onik
i/G
reec
e
Form
atio
n of
pot
enti
ally
toxi
c su
bsta
nces
dur
ing
heat
ing
of fo
ods
P2
And
erss
on, C
-G &
Lin
gner
t, H
Fä
rsk
Fors
knin
g, S
IK, G
öteb
org,
7
Febr
uary
200
7 A
tt u
ndvi
ka a
kryl
amid
när
man
bak
ar (
To
avoi
d ac
ryla
mid
e w
hen
baki
ng)
P6
J.J.
Kno
l, J.
P.H
. Lin
ssen
and
M
.A.J
.S. v
an B
oeke
l Ph
D s
tudy
trip
USA
, pre
sent
atio
ns a
t di
ffer
ent u
nive
rsit
ies
in th
e U
SA
Kin
etic
Mod
elin
g: A
Too
l to
Und
erst
and
the
Form
atio
n of
Acr
ylam
ide
(Ora
l pre
sent
atio
ns)
P7
J.J.
Kno
l, G
. Vik
lund
, J.P
.H.
Lins
sen,
I. S
jöho
lm, K
. Sko
g,
M.A
.J.S
. van
Boe
kel
CO
ST 9
27 A
ctio
n W
orks
hop,
Sof
ia,
Bul
gari
a K
inet
ic M
odel
ing:
A T
ool t
o U
nder
stan
d th
e Fo
rmat
ion
of A
cryl
amid
e in
Foo
d (O
ral p
rese
ntat
ion)
P7
Vác
laví
k L.
, Bar
táko
vá V
., H
olad
ová
K.,
Haj
šlov
á J.
H
eato
x Pr
ojec
t Mee
ting
Pra
gue,
Cze
ch
Rep
ublic
, 21.
-23.
1.20
07
Fast
and
sim
ple
met
hod
for
acry
lam
ide
anal
ysis
and
con
tent
of
acry
lam
ide
in C
zech
bre
ads
P10
Page
76 of
77
Siri
Hel
land
Han
sen
and
Gun
nar
Bru
nbor
g In
tern
al s
emin
ar a
t the
Div
isio
n of
E
nvir
onm
enta
l Med
icin
e, N
orw
egia
n In
stit
ute
of P
ublic
Hea
lth
Inta
ke o
f cri
sp b
read
and
cof
fee
whi
le m
akin
g ch
ildre
n –
mut
ually
ex
clus
ive?
(in
Nor
weg
ian)
P1
6
Siri
Hel
land
Han
sen,
Min
h H
oang
, Eri
k Sø
derl
und
and
Gun
nar
Bru
nbor
g
HE
AT
OX
36
mon
th m
eeti
ng, P
ragu
e,
The
Cze
ch R
epub
lic, 2
1st-2
3rd o
f Jan
. 20
07
Gen
otox
ic e
ffec
ts o
f gly
cida
mid
e on
mou
se te
stic
ular
cel
ls
P16
J.J.
Kno
l, J.
P.H
. Lin
ssen
and
M
.A.J
.S. v
an B
oeke
l 9t
h In
tern
atio
nal S
ympo
sium
on
the
Mai
llard
Rea
ctio
n Fa
te o
f acr
ylam
ide,
mel
anoi
dins
, sug
ars,
asp
arag
ine,
org
anic
aci
ds a
nd
pH in
an
aque
ous
fruc
tose
-asp
arag
ine
reac
tion
sys
tem
at p
H 5
.5: a
ki
neti
c an
alys
is (
Subm
itte
d fo
r O
ral p
rese
ntat
ion)
P17
Oth
er w
ays o
f di
ssem
inat
ion
Ani
ka d
e M
ul, P
olly
E. B
oon,
Ja
cob
D. v
an K
lave
ren
Tra
inin
g fo
r E
FSA
and
Nat
iona
l Foo
d A
utho
riti
es
Tra
inin
g pr
ogra
m o
n pr
obab
ilist
ic e
xpos
ure
asse
ssm
ent f
or
inte
rnat
iona
l and
nat
iona
l foo
d sa
fety
aut
hori
ties
P17
Page 77 of 77