The nightmare of doping i-series - Shimadzuods of the previous LC-2010 sys-tem or non-Shimadzu...

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arst

ellu

ng, N

ASA

/JPL

-Cal

tech

The nightmare of doping offenders

Analysis of doping agents using thenew LCMS-8050 Triple Quadrupole Mass Spectrometer

Including part 3 of the bigbreakfast test

i -series – new integrated (U)HPLCsystems

Nexera-i / Prominence-i – high speed,performance, maintainability and effi-ciency

New: ICPE-9800Espresso in Lake Constance

Five application examples of simulta-neous elemental analysis sensitivity

Changing forest – what happens during Autumn? 2

About knackwursts and sausages –The big breakfast test using the EZ-Test-X Texture Analyzer (part 3) 7

Partial oxidation of isobutane – New analysis method using GC 8

Plastics – Water content as indicatorfor ageing processes 10

The nightmare of doping offenders –Analysis of doping agents using thenew LCMS-8050 Triple QuadrupoleMass Spectrometer 12

Espresso in Lake Constance – Fiveapplication examples of simultaneouselemental analysis sensitivity 15

Bilirubin determination in cerebro -spinal fluid following a subarachnoidhemorrhage – UV spectroscopy sup-ports fast response 17

New: TOC Application Handbook, Version 2 19

„What can i do for you?” The new i -series – Nexera-i / Prominence-i –new integrated (U)HPLC systems 4

APPLICATION

PRODUCTS

Precise temperature control of samplecells – New CPS-100 accessory forUV-VIS spectrophotometers 6

LATEST NEWS

SHIMADZU NEWS 3/2014

CONTENT

Changing forest – whathappens during Autumn?D uring Autumn leaves losetheir green coloring, a pro -cess also known as senes-cence. This is a generic term forthe process of ageing in livingorganisms. It is assumed that se -nescence is associated with thechlorophyll contained in plants(see infobox). These interrelation-ships and processes have been wellinvestigated scientifically and canbe described as follows:

During photosynthesis, chloro-phylls absorb energy from sun-light to convert carbon dioxideand water into carbohydrates andoxygen. During this process, chlo -

and can lead to the destruction ofvital plant organelles. This is whychlorophyll must be degradedvery rapidly after separation fromthe protein. This process is accom -panied by the color change [2].Each chlorophyll molecule alsocontains four nitrogen atoms,which can be recycled for newprocesses in the plant.

Approach

For a more detailed investigationof senescence, the chlorophyllcontent of the leaves of an oak(Quercus) was determined. Theoak leaves were also analyzed with

rophylls are bound to apoproteinswhereby the absorbed energy canenter metabolism.

With increased ageing, this pro -cess becomes less stable and theproteins are gradually removed.Should chlorophylls, at this point,not decompose as well, the energyproduced would need to flow intoother processes. In this case, oxy-gen molecules would be availableto absorb the energy. Oxygen mol -ecules in the ground state (O2)and in the triplet state (3O2) willyield activated (O2*) and singletoxygen (1O2) [1]. Such an oxygenmolecule is much more reactive

Figure 1: Structure of chlorophyll and its various naturally occurring species [3]

NEWS – Shimadzu’s customer maga -zine is also available as WebApp(for iOS and Android) via: www.shimadzu-webapp.euor as app for iPhone in the AppStore.

CH2

CH3

CH3

H3C

H3C

H3C H3C 3

H3COOCR3OOC

Phytyl:

R1

R2

Mg

N N

NN

O

CH2

CH3

H3C

H3C

H3COOC

R1

R2

OHOOC

Mg

N N

NN

R1= CH3R2= C2H5R3= Phytyl

R1= CHO

R2= C2H5R3= Phytyl

c1c2c3

R1= CH3 R2= C2H5R1= CH3 R2= CH = CH2R1= COOCH3 R2= CH = CH2

Chlorophyll a Chlorophyll b Chlorophyll c

Figure 2: Determination of chlorophyll a and b in oak leaf extracts using HPLC-F-AAS.

a: an oak leaf still green in autumn, b: a withered oak leaf.

b

a

Time [min]5 6 7 8 9 10 11

10

15

20

25

30

10

15

20

25

30

Inte

nsit

y [m

V]

3SHIMADZU NEWS 3/2014

APPLICATION

cules contain magnesium as thecentral atom (see figure 1). Asbefore, the chlorophyll specieswere separated via HPLC (Pro minence Modular HPLC).For a more specific detection,

flame AAS (AA-7000) was subse-quently used. The absorption(output as current in μA) can betapped off via the analog port ofthe AA-7000 and can be time-de -pendently re corded.

As chlorophyll is ashed in theflame and the magnesium is thenpresent in its atomic form, a con-ventional water-based magnesiumstandard can be used. This stan-dard is diluted accordingly using

respect to their other naturallyoccurring chlorophyll species (seefigure 1). Aim of the investigationwas direct comparison of greenand yellow, already withered (se -nesced) leaves. In order to assurethe direct comparability of sam-ples with different extent of age-ing, a tree was sampled in autumn.At that time, the tree bore greenas well as yellow, already witheredleaves.

Methanol was used to extract thechlorophylls from the leaves. Pre -liminary experiments showed thatan influence on the different chlo -rophyll species could be largelyexcluded.

For detection purposes, the UV-VIS activity of chlorophyll is gen-erally taken advantage of. Thechlorophyll species are separatedusing HPLC and subsequentlydetected using a diode-array de -tector (DAD). This method wouldhowever require the use of refer-ence standards which are onlyavailable for chlorophyll a and b,to ensure an accurate analysis. Areliable statement on other chlo -rophyll species (see figure 1)would not be possible.

A different approach was there-fore used, in which the specificstructure of chlorophyll was takenadvantage of, as each of the mole-

»Chlorophyll and photosynthesis«

The term ‘chlorophyll’ is derived from ancient Greek and means ‘greenleaf.’ Chlorophyll is responsible for the green coloring of leaves. But it isnot only present in plants since other organisms also contain chlorophyll.

It is necessary for photosynthesis, the photochemical process that pro-duces glucose, oxygen and energy from water and carbon dioxide. This is, however not always the case. Oxygen production only occurs in oxy-genic phototrophs.

Photosynthesis can also generate compounds other than oxygen and glucose and this type of photosynthesis is carried out by anoxygenic phototrophs. Both the products generated and the chlorophylls involved in these processes differ.

methanol, so that the sample ma -trix and the standard matrix areidentical. The water content of theMg-standard does not cause anyproblems due to the high dilutionfactor and can be neglected.

Result

Except for one intermediate in thechlorophyll degradation process(chlorophyllide), none of the de -gradation products contain thebound magnesium. This is whythe measurement of senescencewas limited to the decrease in in -tensity of chlorophyll a and b.

The concentration of chlorophylla decreased by a factor of 8.5 dueto senescence. In chlorophyll b,this decrease was even more dra-matic. The samples only contained6.1 % chlorophyll b in compari-son to the sample with the non-withered leaves. Before senes-cence, the ratio between chloro-phyll a and b was 4.4 :1. The natu-ral ratio is 3 :1. This indicates thatthe green leaves were also at theinitial stage of senescence, whichcould be confirmed by the time ofsampling (autumn). After ageing,the ratio increased to 8.4 :1. Chlo -rophyll b was therefore degradedfaster than chlorophyll a. Chloro -phyll degradation is accompaniedby the change of color when otherplant substances such as carot -enoids (yellow/orange) and tannin(brown) are emerging.

This problem can be addressedusing the AA-7000 coupled to themodular Prominence HPLC. Inparticular, the possibility to cali-brate using cost-effective water-

based magnesium standards in -stead of expensive chlorophyllstandards makes the coupling ofHPLC and AAS extremely lucra-tive. The possibility to studyother chlorophyll species andtheir degradation is also interest-ing. Chlorophyll c, for instance, ismainly present in algae. Althoughalgae do not age depending onseason, they are however limitedin their life expectancy by ecologi-cal changes.

AuthorsFlorian Pahlke, Prof. Dr. Jürgen Schram

Laboratory for Element and Species Analysis

Faculty of Chemistry

University of Applied Sciences

Krefeld, Germany

Literature[1] T. Masuda und Y. Fujita, „Regulation and

evolution of chlorophyll metabolism“,

Photochemical & Photobiological Sciences,

Bd. 7, Nr. 10, p. 1131, 2008.

[2] S. Hörtensteiner, „The loss of green color

during chlorophyll degradation – a prereq-

uisite to prevent cell death?“, Planta,

Bd. 219, Nr. 2, pp. 191-194, 2004.

[3] Chlorophylle. In: Römpp Online. Georg

Thieme Verlag, (May, 19. 2014).

Further information

on this article:

• Application Note

Analysis of Mg by HPLC

FAAS (PDF)

www.shimadzu.eu/shimadzu-news-2014

Figure 2: Placing the RID-20A on top of

the i -series

of the current status of the instru-ment or monitoring of the currentdata acquisition pro cess. Thecolor touch panel also enablesvisualization of the re corded chro-matograms, in addition to com-plete instrument control with

method and batch creation (fig-ure 3).

Intelligence

Intelligence is embodied in the i-series by straightforward meth -od transferability, whereby meth-ods of the previous LC-2010 sys-tem or non-Shimadzu systems canbe adapted easily. Furthermore,the compact HPLCs of the i-seriescan carry out fully automated anal -ysis workflows.

An overview is presented in table 1.

The ‘Auto Validation’ function(figure 4) supports the user in ob taining a report on whether the system is working in a stable

PRODUCTS

H igh speed and outstandingperformance, maintainabil-ity and efficiency – the new i-series with the compactPromi nence-i (HPLC) and Nexera-i(UHPLC) versions meets theneeds of large as well as small lab-oratories. The letter i stands forinnovation, intelligence and intu-itive operation, all of which arecombined in the i-series concept.

The new Nexera-i and the Promi nence-i systems are perfectfor routine analyses in the chemi-cal, pharmaceutical, food and envi -ronmental industries. The systemscan be operated via workstationfor a single user or a large client-server system for multiple-user

carryover increases accuracy ofthe analytical results.

In addition to the absorption de -tector (UV or PDA) included inthe standard configuration, a fluo-rescence detector or refractive

touch panel or a PC, is highlyinnovative (figure 1). Furthermoremethods or sequences can be bi-directionally synchronized fromthe instrument to the software.

The excellent baseline stability,even at fluctuating ambient tem-perature, is also an innovative fea-ture due to a unique dual-temper-ature controlled detector unit (de -tector cell and detector optics).An injection cycle of 14 secondsre duces analysis times and the low

operation. The small surface areareduces space requirements in thelaboratory and simplifies possiblerelocation.

Further development of the LC-2010 predecessor model is charac-terized by the improvement of allsystem components (degassingunit, quaternary pumps, autosam-pler, oven) and also enables, in ad -dition to optional cooling of theautosampler, the configuration ofan UV or PDA detector. Further -more, the Prominence-i allowsworking in a pressure range of upto 440 bar, while the Nexera-iallows working at pressures up to660 bar.

Innovation

The ICM (Interactive Commu -nica tion Mode), which allowsoperators to edit and launch meth -ods and sequences directly from a

4 SHIMADZU NEWS 3/2014

“What can i do for you?”

index detector, as well as a lightscattering detector, can be added. It is also possible to implementdata acquisition via other detec-tors using an analog input. Whencombining the i-series with theequally innovative RID-20A re -fractive in dex detector (figure 2),it is possible to configure a simpleGPC system with a minimumfootprint.

Besides optional interactive com-munication modes enable re motemonitoring. This allows checking

Figure 1: Touch panel of the i -series

Figure 4: Auto Validation

Figure 5: New user interface for LabSolutions software

Figure 3: Chromatogram displayed on the

touch panel

The new i-series – Nexera-i / Prominence-i – new integrated (U)HPLC systems


PRODUCTS

range. Here, a fixed and thus atany point in time comparable pro-cedure is used that checks solventdelivery stability or the wave-length accuracy of the absorptiondetector. In addition, it is possibleto carry out an automated routineinspection and self-diagnosis ofthe instrument via the ‘SystemCheck’ function. Data recordedon consumables used, for instancethe number of injections, volumedelivered by the pump or usage ofthe detector lamp, is read. The re -sults of these two status reportscan be viewed on the instrumentmonitor or printed via the soft-ware.

Intuition

The new unified user interface ofthe i-series and the new version of the LabSolutions software are Table 1: Automation of the HPLC analysis workflow using the i -series

Figure 6: Quick Batch function of the LabSolutions software Figure 7: Product lines and categorization of Shimadzu HPLC and UHPLCs

System startup

Purging of the flow lines

Column equilibration

Starting the analysis

Sample measurement

Ending the analysis

Column cooling down Pump delivery and detectorlamp switches offColumn oven temperaturecontrol switches offSystem shutdown

Auto start-up

Auto purge

Baseline check

Analysis starts

Direct access mechanism

E-Mail notification

Auto shut-down

Automated system startup by setting a time

Automated purging of the flow lines with eluents prior to analysisMonitors baseline stability and determines when the analysis can be started, based on predefined noise/drift criteria Analysis is started automatically as soon as the baselinecheck is completedSamples can be added at any time during the ongoing analysisAfter completing the analysis, an email can be sent automatically to laboratory personnelAutomated instrument shutdown with 95 % reduction in energy consumption in the standby mode. Prior rinsing of the instrument or the column can be implemented

Workflow for implementing an HPLCanalysis

Automation Comment

high ly intuitive. This can be illus-trated via a simple comparison be -tween figures 1 and 5, in whichthe flowline of the i-series is visu-alized, thus enabling intuitive op -eration of the software via simpleselection of the various compo-nents.

Another novel software feature isthe ‘Quick Batch’ function whereusers can easily and quickly createa sequence of analyses by selec-ting the vial positions on the sam-ple rack which is graphically dis-played (figure 6).

The combination of i-series withLabSolutions and the system uni-fied graphical user interface andsoftware allows intuitive opera-tion independently of user experi-ence level.

Shimadzu’s comprehensive LCproduct portfolio meets all analyt-ical requirements, from conven-tional up to ultra-fast analyses.Next to the modular HPLC/UHPLC systems, figure 7 showsthe position of the i-series amongShimadzu’s LC products.

Further information

Read more about the new i-serieson www.shimadzu.eu/i-series

LATEST NEWS


S himadzu offers the mostdiverse range of accessoriesfor UV-VIS-NIR-spectro -photometers. A wide variety ofcell holders, with or without tem-perature control, allows custom -ized system configurations for allrequirements of the environmen-tal, pharmaceutical, food, life sci-ence applications and many more.

Whether a single standard cell atroom temperature is used or amulti-cell changer for high samplethroughput is required – the idealsolution for each analytical prob-lem is always available.

The latest release in the Europeanmarket is the CPS-100 thermo-electrically temperature-controlled6-cell changer.

The CPS-100 is a 6-cell changercapable of controlling the temper-ature of the sample cells in a rangefrom 16 °C to 60 °C with a preci-sion of +/- 0.1 °C. Combined with a spectrophotometer such as UV-1800 working in kineticsmode, the system can be config-ured to measure the enzyme activ-ity of up to six samples underconstant temperature conditions.Figure 1 shows the UV-1800 incombination with CPS-100.

The CPS-100 can also be used inthe following spectrophotometers:UVmini-1240 series, UV-2600/UV-2700 and UV-3600.

The UV series is intended formultiple applications. A widerange of accessories enables cus-

Precise temperature control of sample cellsNew CPS-100 accessory for UV-VIS spectrophotometers

Figure 1: UV-1800 with CPS-100

tomized solutions such as emis-sion optics, rotating film holder,sample cups for integrating spheres,diverse selection of sipper units or photomultipliers for the NIRrange.

A variety of software solutions forband gap analysis, Bilirubin deter-mination in CSF and sun protec-tion factor for milks, creams andtextiles complement the hardwarerange.


APPLICATION

F oods as a basic need of hu -man existence are subject toconstant inspections. TheShimadzu News regularly reportson new analytical capabilities.

In addition to taste and the inspec -tion of ingredients, questions arealways raised on the physical prop -erties of our food: how quicklydoes our bread get stale? Howcrisp are our sausages? What arethe differences between eggshellsoriginating from different eggfarming methods? ... These are thequestions that are addressed inthis and in the two previous issuesof the Shimadzu News – using thefoods that make a continentalbreakfast.

Those who wish to supplementtheir breakfast consisting of breadand eggs, might like to try somesausages. Depending on localtaste, these may vary in type andcan be prepared either fried orboiled. Regardless, however, oftheir production and preparation:all sausages are expected to be ten-der but still firm, as in addition tothe intense flavor, consumers alsowant an equivalent bite sensation.

Taste and bite sensation

Additionally to the questionwhich type of sausage best offersthis bite sensation, the questionon possible differences betweenindustrially produced or home-made type sausages needs to beanswered as well. To avoid anydifferences caused by the method

raw, the Nuremberg sausage ismuch crispier when fried com-pared to the cold version.

Part 1: How do you like your eggs? – The big

breakfast test using the EZ-Test-X

Texture Analyzer

Part 2: The daily bread – The big breakfast

test using the EZ-Test-X Texture

Analyzer

of preparation, the sausages werealso tested in the raw state.

All tests were carried out usingthe EZ-Test-X Texture Analyzer.As cut resistance and firmness areproportional, the sausages werecut using a blade test meth od, inorder to exclude differences inchewing behavior.

As can be seen in the table below,the cutting force for sausages thatare mainly eaten warm (for in -stance Krakauer Polish sausagesor Bavarian veal sausages) is sig-nificantly higher for the raw statecompared to after being cooked.This may be explained by the factthat the raw sausage is consider-ably softer in its initial raw con-sistency and, therefore, a muchhigher force is required to be ableto make the first incision. Duringcooking, the sausage mass expandsand the casing tightens.

In contrast, although Vienna sau -sages show no differences betweenthe raw and cooked state, as theyare suitable to be eaten cold aswell as warm, there is however asignificant difference with respectto cutting force between Viennasausages obtained from a discountsupermarket or from a butchershop.

The results for Bavarian veal sau -sages and Nuremberg grilled sau -sages, however, appear to vary.While the Bavarian veal sausagecan be cut into much more easilyin the cooked state than when

About knackwursts and sausagesThe big breakfast test using the EZ-Test-X Texture Analyzer (part 3)

Tabelle 1: Cutting resistance comparison

Further information

on this article:

•Full big breakfast test

(PDF)


ABABAB

8794

111911742

497098842044

Krakauer / Polish sausage

Mettenden / Beerstick sausage

Vienna sausage (Wiener)

Bavarian veal sausage

Nuremberg grilled sausage

B

B

43

13

31

27

Sausage typeDiscount supermarket (A) /

butcher (B)Cutting force [in N] raw

Cutting force [in N] boiled / fried

Figure 1: Cutting resistance test with EZ-Test LX

APPLICATION

P eroxides (R1-O-O-R2) andhydroperoxides (R1-O-OH)are increasingly being used inindustry and as oxidizing agents inorganic chemistry. Hydroperoxides(ROOH) are the initial, generallyless stable intermediate products inthe oxidation of hydrocarbons(Allara et al. [1968]). For instance,they occur when edible oils turnrancid.

Among the most stable representa-tives of this class is t-butyl hy -droperoxide (TBHP). Current re -search focuses on improving theefficiency of the industrial produc-tion process based on isobutane.Isobutane usually yields severalproducts, as the decomposition ofTBHP also plays a role. A prereq-uisite for increasing the efficiencyis, therefore, to carry out a com-prehensive study of the processes

pounds could be considered for thepreparation of the analysis sam-ples. This is why, in particular,potential solvents such as hexane,decane, ethanol, toluene, acetoni-trile and dioxane, as well as com-pounds that could be used as inter-nal standards or additives, such ast-butyl methyl ether, methyl ethylketone and n-hexane were tested.

The main reaction products areTBHP, t-butanol, acetone andmethanol; other byproducts canoccur due to strong thermal de -composition. An additional prob-lem is the sensitivity of hydroper-oxides to metals (including steel)that can catalyze the decomposi-tion.

Among the gaseous products, par-ticularly isobutane, isobutene, oxy-gen, nitrogen, carbon dioxide, car-

To increase the rate of the reaction,it would be desirable to carry outthe reaction at a higher tempera-ture. However, increasing the tem-perature also increases the decom-position of TBHP and in additionto gaseous products, alcohols, ke -tones and aldehydes are formed.This is why a method was neededthat could separate the complexmixture occurring during this pro -cess. For this purpose, however,only GC is suitable and Shimadzu’sGCMS-QP2010 Ultra was selected.

Sample preparation

For method development, poten-tially liquid and gaseous productsreported in the literature wereidentified. First, a mixture of theidentified liquid products was pre-pared in toluene. It should also beclarified which additional com-

and a detailed analysis of the com-plex reaction mixture, which has sofar been largely neglected.

Gas chromatography – the method of choice?

For many years, hydroperoxideshave not been analyzed using GC,since it was known that they easilythermally decompose. Later, it be -came evident that tertiary com-pounds, in particular (R3C-OOH),were relatively stable. Partial oxi-dation of isobutane is usually car-ried out at approximately 130 °C.


Partial oxidation of isobutaneNew analysis method using GC

Figure 1: Chromatogram of all separated compounds in the liquid phase (peak assignment

in table 2), (TIC = Total Ion Count)

Minutes

0.000.050.100.150.200.250.300.350.400.450.500.550.600.650.700.750.800.850.900.951.001.051.101.15

TIC (x 10,000,000) Max. Intensity: 11,897,460

1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0 12.0 13.0 14.0 15.0 16.0 17.0 18.0 19.0

Figure 2: Extension of the range 1 to 3 min in figure 1 with simultaneous representation

of the chromatograms of the GC grade toluene

Minutes

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

1.1

1.2

TIC (x 10,000,000) Max. Intensity: 81,126,853

1.1 1.2 1.3 1.4 1.5 1.6 1.7 1.8 1.9 2.0 2.1 2.2 2.3 2.4 2.5

Table 2: Retention times of the separated compounds

123456789

1011

Nr.

Hexanet-butyl methyl etherAcetaldehydeDi-t-butyl peroxideIsobutylene oxideAcetonet-ButanolMethanolMethyl ethyl ketoneEthanol Decane

Compound

1.3881.4881.5191.6381.6641.8252.0832.1212.2402.3353.054

Retention time /min

1213141516171819202122

Nr.

AcetonitrileTolueneDioxaneIsobutanolEthyl benzenep-Xyleneo-Xylenet-Butyl hydroperoxiden-HexanolAcetic acidFormic acid

Compound

3.2013.9004.1464.2315.4655.6755.7799.0899.648

11.40412.806

Retention time /min

Table 1: Temperature program

0 min5 min

16 min19 min

80 °C80 °C

190 °C250 °C

Time Temperature


APPLICATION

bon monoxide and methane – aswell as propene at higher tempera-tures – can be expected. In prepa-ration for the gas analysis, a cali-bration gas containing all thesegases was obtained from LindeEngineering AG. In order to carryout the analysis of both the gasphase as well as the liquid prod-ucts, a suitable GC configuration is needed.

Application

This problem could be solved viacolumn switching using MDGC-Switching (multidimensional GC).For the separation of polar com-pounds, an RT Stabilwax columnwas used (length 30 m, ID 0.25 mm,film thickness 0.25 μm). Separationand detection of the permanentgases is carried out using a molecu-lar sieve column (RT M-Sieve 5A30 m, ID 0.53 mm, film thickness0.50 μm), which is connected tothe Stabilwax column via theswitching valve, and a thermal con-ductivity detector (TCD). Afterapproximately two minutes, thevalve switches over from the TCDto the MS system. The productsretained by the Stabilwax columnare then detected using a massspectrometer, after passing througha stainless steel (SilcoTek GmbH,Silcosteel) restrictor (0.45 m, ID0.15 mm). The restrictors of theswitching valve were also manufac-tured from Silcosteel.

Separation of the mixture

a) Liquid phaseAt an isothermal phase of 40 °Cfor 5 minutes with subsequentheating to 250 °C at 10 °C/min, thecomponents of the liquid sample

Conclusions

All potential liquid reaction prod-ucts could be separated successful-ly, whereby the analysis of thestrongly polar liquid products aswell as the gases could be madepossible via column switching. Thepresent configuration, however,does not allow the separation of allgases. Propene was not found. Thisproblem is being addressed further.

AuthorDr. rer. nat. Thomas Willms

Helmholtz-Zentrum

Dresden-Rossendorf, Germany

Institute of Fluid Dynamics

LiteratureAbraham, M.H.; Davies, A.G.; Llewellyn,

D.R.; Thain, E.M.

The Chromatographic Analysis of organic

Peroxides. Analyt. Chim. Acta. 17 (1957),

499 - 502.

Allara, D.L.; Edelson, D.; Irwin, K.C.

Computational Modeling of the Mechanisms

of the free radical-chain Reaction of Alkanes

with Oxygen. The Oxidation of Isobutane,

n-Butane, and Isopentane.

Int. J. Chem. Kin. 4 (1972), 345 - 362.

Figure 2 shows that the GC gradetoluene (99.9 %, signal hidden) inaddition to the xylene isomers con-tains other compounds which areresponsible for the smaller signalsin the lower retention time range.The TBHP exhibits a significantlyhigher retention time than all thealcohols with a smaller or compa-rably large alkyl group. In the calibration of formic acid (figure3), the dependence is linear up to0.1 vol %.

b) Gas phaseThe permanent gases nitrogen,methane, oxygen and carbon mon -oxide detected via the TCD yieldthe gas chromatogram presented infigure 4. The remaining gases areretained on the Stabilwax columnand – in addition to a nitrogenresidue – are detected via the massspectrometer (figure 5). Calibra -tion of the gases is carried outusing one-point calibration basedon the mean of three reproducibili-ty measurements, which is com-mon in this case.

are only partially separated. The t-butanol and methanol signalsoverlap each other, although theyare separated when using the head-space technique with a retentiontime difference of approximately0.2 minutes. In the mass spectrum,the peak is therefore identified as2-pentanol. Separation is not possi-ble, as the mass of the main peakof methanol (m/z = 31) is the samefor both compounds. By increasingthe temperature of the isothermalphase from 40 °C to 80 °C (seetable 1), all compounds were wellseparated.

Figure 1 shows the chromatogramof such a separation. Althoughmethanol and t-butanol are notbaseline separated, the signals canbe clearly distinguished via theirmass peaks. This temperature isnot yet critical for TBHP and hadalready been used in the past(Abraham [1959]). The toluenepeak was hidden by switching offthe filament in the range of 3.8 -4.0 minutes.

Figure 3: Calibration of formic acid

Volume %

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

1.1

1.2

1.3

1.4

1.5

Area (x 1,000,000)

0.000 0.010 0.020 0.030 0.040 0.050 0.060 0.070 0.080 0.090 0.100

Figure 4: Chromatogram of the permanent gases nitrogen (1), oxygen (2),

carbon monoxide (3) and methane (4)

Minutes

-2.5-2.0-1.5-1.0-0.50.00.51.01.52.02.53.03.54.04.55.05.5

uV (x 1,000)

2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0 7.5 8.0 8.5 9.0 9.5 10.0 10.5

Figure 5: Chromatogram of carbon dioxide (1), isobutane (2) and isobutene (3).

(Upon switching it is not completely possible to avoid entry of nitrogen).

Minutes

0.5

1.0

1.5

2.0

2.5

3.0

3.5

4.0

4.5

5.0

1.50 1.55 1.60 1.65 1.70 1.75 1.80 1.85 1.90 1.95 2.00 2.05 2.10 2.15 2.20 2.25 2.30 2.35 2.40 2.45

Max. Intensity: 47,495,565TIC (x 10,000,000)

APPLICATION

6 billion tons of plastics havebeen produced worldwide inthe last 50 years. Due totheir many versatile properties,plastics can be found in virtuallyall products. ‘Low-density’ (LD-PE) and ‘high-density’ (HD-PE)polyethylenes in particular arealmost exclusively used in thepackaging industry as they arelong-lasting and persistent.

When plastics are used as packag-ing materials, it is important totest their water content duringquality control, as this solvent canpotentially transport productcomponents outside of the pack-aging, as well as components fromthe environment inward. In addi-tion, the water content can con-tain information about the condi-tion of the polymer, since water isformed in light-induced ageingprocesses of polyethylene [1]. Dueto capillary forces, water moves

granulates in various ageing stageshave been investigated – from newmaterials to recycled materials andplastic wastes from the oceans.

In the first step, the samples aresplit so that the outer surface andthe core of the respective granu-lates become accessible. Smallfibers are then cut from the outerand inner surfaces using a scalpel.These can be measured directlywith the FTIR microscope.

Reflectance mode with the mirroras supporting surface is particular-ly suitable for this purpose. Formeasurements in the transmissionmode, the use of a diamond com-pression cell is recommended. Theseparated fibers are pressed into athin film and can be measuredwithout any additional supportingsurface.

Evaluation of the spectra

The determination of adsorbedwater in polyethylene using FTIR

between the polymer chains, fromthe polymer surface into the coreand forms water clusters. In theevent of damage, a spatially re -solved analysis of water within thepackaging material could offerindications of potential causes.

In general, ageing processes ofplastics can be observed easilyusing infrared spectroscopy, dueto the high detection sensitivity ofthe resulting decomposition prod-ucts such as water, aldehydes orcarboxyl groups [1]. A great ad -vantage of this type of analysis isits non-destructiveness and itsminimal effort required. Complexsample preparation steps are notusually needed. The high spatialresolution required can beachieved using an FTIR micro-scope. In this way, even the small-est sample components can beexamined accurately for possibledamage.

In the present analysis, ‘low-den-sity’ and ‘high-density’ polymer


Plastics – Water contentas indicator for ageingprocessesPrecise measurements using FTIR microscopy

Figure 1: FTIR spectrum (absorption) of water. Typical broadband, finely split absorption

patterns in the wavelength range of 4,100 - 3,500 cm-1 and 2,000 - 1,200 cm-1 [2][5].

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Shimadzu NEWS, Customer Magazine of Shimadzu Europa GmbH, Duisburg

PublisherShimadzu Europa GmbHAlbert-Hahn-Str. 6 -10 · D-47269 DuisburgPhone: +49 - 203 - 76 87-0 Fax: +49 - 203 - 76 66 [email protected]

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APPLICATION

microscopy requires special che -mometric methods. Among otherreasons, this is due to water in theform of moisture being containedin the background signal (atmos-phere). In addition, water is ad -sorbed at ambient temperatureonto the surfaces of the measure-ment system.

These effects are counteracted bysubtracting the background spec-trum as well as using empiricalatmospheric corrections. A furtherchallenge lies in the complex vi -brational bands of water. At ambi-ent temperatures, there are manydifferent vibrational and rotationalstates due to van der Waals inter-actions. This results in very broadand finely split absorption pat-terns in the wavelength range of4,100 - 3,500 cm-1 and 2,000 -1,200 cm-1 (see figure 1) [2].

As a result of these two effects, asemi-quantitative comparativeanalysis of the spectra is recom-mended. In this method, the sec-ond derivative of the spectroscop-ic data, including a Savitzky-Golay smoothing, is first deter-mined. This corrects the baselineand enables a better comparisonbetween the individual samples[3]. Using conventional statisticalmethods such as the ‘two samplet-test’ and the ‘effect size’, theranges of the IR spectrum exhibit-ing significant differences between

the sample’s inner and outer sur-face can subsequently be isolated(figure 2). The t-test determinesthe significance of a differencebetween two mean values, whilethe effect size describes the extentof that difference [4].

Results and perspectives

Using FTIR microscopy, it can beshown that in terms of water con-tent, there are significant differ-ences between ‘low density’ and‘high density’ polyethylenes, aswell as between new materials andplastic wastes. Much smaller waterclusters are present in HD-PE,which can be attributed to theincreased crystallinity. However,the theory on ageing processes ofpolyethylene can also be con-firmed, as comparatively largequantities of water can be deter-mined in the significantly agedplastic waste samples. A compari-son of the spectra of all measuredsamples as well as a comparativeanalysis is shown in figure 3. The samples were measured usingShimadzu’s AIM-8800 FTIR mi -croscope.

The method presented can beextended through the use of amicrotome for improved spatialresolution and reproducibility. Inaddition, as part of a multivariatedata analysis, a calibration of thewater content is conceivable.

AuthorsGerrit Renner, Jürgen Schram

Department of Chemistry

Frankenring 20, 47798 Krefeld

University of Applied Sciences Niederrhein,

Germany

Albert van Oyen

CARAT GmbH Research

Harderhook 20, 46395 Bocholt

Germany

Literature[1] K. Jansen, Influence of acid on the

photochemical ageing of UV-stabilized

polyethylene films, Dissertation University

of Berlin, Germany (2003), 75-93

[2] T. Bruno, P. Svoronos, Handbook of

Basic Tables for Chemical Analysis,

CRC, (2003), 351

[3] B. C. Smith, Fundamentals of Fourier

Transform Infrared Spectroscopy, CRC,

(2011), 69-74

[4] J. Cohen, Statistical Power Analysis for the

Behavioral Sciences, LEA, (1988), 274

[5] G. Socrates, Infrared and Raman

Characteristic Group Frequencies, WILEY,

(2001), 26

Figure 2: (Bottom) A comparison of mean value FTIR spectra (absorption) of LD-PE granu -

lates (new material) from five measurements each of the outer and inner surfaces.

The inner surface exhibits an increased background absorption and a strongly pronounced

water absorption pattern. (Top) The extended effect size shows various ranges in the FTIR

spectrum, which differ significantly. The water absorption ranges are clearly visible.

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Figure 3: FTIR subtraction spectra (absorption inner surface – outer surface) in wavelength

ranges 1,750 - 1,500 cm-1 (water absorption) [2][5]. The plastic waste investigated shows

the most pronounced water absorption, which can be indicative of the quantitatively high -

est water content. LD-PE exhibits a slightly increased water content in the interior of the

granulate, while HD-PE exhibits even less water in the core. The recycled sample is a mix-

ture from both PE variants and lies between LD-PE and HD-PE.

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APPLICATION


An early form of testosterone dop -ing began with the discovery ofthe performance-enhancing effectsof bull testicles. Even animalswere not spared when it came todoping: already in the middle of

the 17th century, the performanceof horses was affected. This hasbeen shown in an official decreeof a small British town. In thosedays, however, horses were alsobeing poisoned, for example witharsenic, in order to place a bet ona competitor’s horse. As it had notbeen possible to detect illegal sub-

F aster, higher, further – dop-ing use has been associatedwith sports for many centu -

ries. Already during the Olym picGames held in ancient Greece,athletes were using special plantdiets to enhance their physical fit-ness. Beyond the sports, the an -cient Incas chewed on coca leavesas a stimulant, and the Chineseswore by the ephedrine-contain-ing Ma-Huang herb.

The nightmare of doping offendersAnalysis of doping agents using the new LCMS-8050 Triple Quadrupole Mass Spectrometer

stances, it was not until 1812 thatthe first doping case was discov-ered – only be cause the culpritwas caught in the act.

From nutritional supplement to doping

During World War II, pharmacists

discovered how to synthesizederivatives of testosterone: ana-bolic steroids. Initially, they wereused to feed emaciated prisonersof war. Soon athletes discoveredthese preparations, which werehenceforth considered to be the‘breakfast of champions’ – but notonly for human champions.

Figure 1: LCMS-8050 Figure 2: Comparison of data quality for flunitrazepam at different scanning speeds


APPLICATION

The American racehorse Hollo -way won one race after anotherafter receiving testosterone.

Also today, doping incidents arebeing reported time and again, inspite of all control measures. Dur -ing the Olympic Games in Bei -jing, six horses were tested posi-tive. A current example is the mostrecent doping scandal in endur -ance racing, a horse racing variantthat enjoys great popularity in theArab countries. The winner of theprestigious UAE Pres ident’s Cup,which was held in Abu Dhabi inSpring 2013, was banned for twoyears after a doping-related ste -roid was detected in his horse.Already in pure mathematicalterms, the race across the deserthappened at a pace that must havebeen harmful to the health of theanimal.

The seizure of over 120 bannedand illegally imported substancesincluding anesthetics, anti-inflam-matory drugs and antibiotics, at a British thoroughbred stud farmat the beginning of this year illus-trates that the well-being andhealth of horses are no longer pri-oritized, but that the animals arebeing downgraded to ‘sportsequip ment’ that must deliver max-imum performances during theday of the race.

What is doping?

For years, animal welfare organi-zations have been demanding con-sistent actions against the quitelenient anti-doping sanctions inequestrian sports. In Germany, theFN Anti Doping and Drug Con -

trol regulations (FN-Anti-Dop -ing- und Medikamentenkontroll-Regeln, ADMR), issued by theGerman Equestrian Federation(Deutsche Reiterliche Vereini -gung, FN) and adapted to interna-tional regulations, have been inforce since 2010 and are continual-ly updated.

Since April 1, 2011, German top-class riders must be prepared for‘house calls’ by the National AntiDoping Agency (NADA), in addi-tion to regular doping controlsduring tournaments. These con-trols on horses outside of thetour nament are unique worldwideand groundbreaking in the fightagainst doping.

In general terms, doping refers tothe use of substances from prohib -ited substance classes as well as tothe use of banned methods for per -formance enhancement. Horseracing is also about negative dop-ing, i.e. ‘doping to defeat’ by de -creasing a horse’s performance andgiving competitors an advantage,or of unauthorized medication. Inunauthorized medication, an exist-ing reduction in performance isassumed to be the result of an ill-ness. The horse is, for in stance,treated with an analgesic and canthus perform ‘normally’ again.Just like doping, performance isinfluenced, which is prohibited incompetition.

In unauthorized medication, how-ever, good intentions are assumed,which is reflected in lower sen-tencing. As broad-ranging as theapplication possibilities are, soalso are the varieties of substance

Figure 3: Chromatogram of a horse urine extract, 13 events / 28 MRMs

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classes used. Stimulants such asamphetamines that make horsesrun up to the point of completeexhaustion are commonly used.Methyl xanthines which includecaffeine also act against signs offatigue. An opposite effect is at -tained by sedatives that are usedto get extremely nervous horses‘ready for takeoff.’ The list of pro -hibited substances also in cludessteroid hormones, including tes -tosterone and peptide hormonessuch as EPO that are used in hu -man doping.

Ludger Beerbaum, one of themost successful show jumpers ofthe past 20 years, commented onhis approach to equestrian sportsover the past years in a 2009‘Frankfurter Allgemeine Sonntags -zeitung’ weekly Sunday paper asfollows: “Over the years, I havebecome accustomed to exhaustingall possibilities ... In the past, I hadthe attitude: anything that is notdetected, is allowed” (cited accord -ing to Wikipedia as well as SpiegelOnline). Today, even the smallesttraces of doping agents can be de -tected in blood and urine. Zerotolerance policies meanwhile ap -ply in equestrian sports, i.e. noprohibited substances may be de -tected in a horse’s blood or urineat the time of the competition.

Advanced analytics extenddetection possibilities

The analytical possibilities of alaboratory are crucial for the de -tection of a substance. Based onoptimized detection methods andconsistently further developedinstruments, substances are now

found whose detection had notbeen possible a few years ago. Inaddition, these substances can bedetected over a much longer timeperiod. Often, the time interval inwhich these substances can be de -tected surpasses the effect dura-tion of a substance.

In these cases, a sample can onlybe considered positive when theamount detected is still effective.This is why doping is sometimesunintentionally a topic of discus-sion when waiting periods or re -tention periods are not properlyobserved, especially when topical-ly applied ointments and gels alsolead to positive test results.

Very recently, the idea of a highlysensitive LCMS system, theyoungest member of Shimadzu’sUFMS (Ultra Fast Mass Spectro -metry) family, has become a night-mare for doping offenders. TheLCMS-8050 high-performancetriple quadrupole mass spectrome-ter (figure 1) features the world’sfastest data acquisition rates andthe highest sensitivity, enablingsimultaneous quantitative andqualitative analyses. This in -strument was developed specifi-cally for increasing quantificationrequirements at trace level in clin-ical research, food analysis andenvironmental analysis. �

APPLICATION


Excellent sensitivity combinedwith high-speed analytics isachieved via two improved tech-niques: the newly designed heatedESI source and the ad vancedUFsweeper-III collision cell withefficient fragmentation. To opti-mize desolvation, the new ly de -signed ESI source uses a heat inggas in combination with a neb -ulizer.

New techniques deliver superb qualitative data

The following application examplefrom horse doping illustrates theexcellent sensitivity as well as theclear advantage of ultra fast MStechnology. Real samples from ahorse doping laboratory were test -ed for various steroid hormones,in the form of free steroids or ste -roid esters. Small molecules ofbanned neuroleptics, benzodiaze -

High-quality data without loss of sensitivity

In the following example, a horseurine extract spiked with 1 pg/μLof a standard was screened fordoping substances using an MSscreening method in which 254MRMs are stored. Due to the highscan speed of the LCMS-8050, itis possible, even with this highnumber of MRMs, to obtain dataof excellent quality and withoutsignificant loss in sensitivity. This

only nine data points can be ac -quired across the marked peak. Asa rule of thumb, at least 10 datapoints should be acquired in orderto ob tain good reproducibility ofqualitative and quantitative re -sults.

A comparison of the standard devi -ations (RSD) for flunitraze pam,calculated for 3,000 u/sec and30,000 u/sec, demonstrates un -equiv ocally the advantage of fastdata acquisition (figure 2). Atthese ultra fast scan speeds, quali-ty as well as sensitivity of thesimultaneously acquired qualita-tive data often decreases, assumingthat the instrument even allowssimultaneous acquisition of MRMand scan data. Both flunitrazepammass spectra in figure 2 prove thatthe LCMS-8050 delivers first-classqualitative data, also at the highestpossible scan speed of 30,000 u/sec.

pines and opioids were also inves-tigated.

The samples were analyzed usingShimadzu’s LCMS-8050 triplequadrupole mass spectrometercoupled to a Nexera X2 UHPLC.

An outstanding feature of theLCMS-8050 is its ultra fast scanspeed for MS data acquisition. Fig -ure 2 illustrates the clear advan-tage of a fast analysis system.Using a scan speed of 3,000 u/sec,

Figure 5: Details of triamcinolone, comparison of chromatograms of 28 versus

254 MRMs

Figure 6: Reproducibilities (6 injections) of an extracted standard acquired using MRM only compared to a combined method MRM

and synchronized survey scan with 30,000 u/sec

-0.100.000.100.200.300.400.500.600.700.800.901.001.101.201.301.401.501.601.701.801.902.002.102.202.302.402.502.602.702.802.90

423.20>347.20(-)@7/ 423.20>347.20(-)@12/ 437.30>361.20(-)@10/ 437.30>361.20(-)@17/ 479.20>45.05(-)@13/ 479.20>45.05(-)@23/

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Fluoro prednisolone Dexa/Betamethasone Triamcinolone acetonide

can be clearly seen when compar-ing the chromatograms of figure 3(28 MRMs) and figure 4 (254MRMs). In addition, detailed as -sessment of a less intense peak(here: triamcinolone) (figure 5)proves the consistent high dataquality despite a considerablyhigher number of MRMs.

In further measurements, repro-ducibility (as a measure of gooddata quality) at simultaneous syn-chronized survey product ion scan

of a doping standard solution wasanalyzed. Shimadzu’s LabSolutionsoftware allows simultaneousMRM and product ion scan meas-urements. The scan does not runpermanently in the backgroundbut is triggered by an increase ofthe masses, stored in the MRMmethod, above a defined thresh-old.

The synchronized survey scan isnot just limited to product ions; itis also possible to define specific

masses in the method that willtrigger a scan. Table 1 (see furtherinformation) shows a comparisonof results for reproducibility ex -periments (n = 6) – working inMRM mode only and then ana-lyzed again in MRM mode with adwell time of 5 msec for all com-ponents plus simultaneous syn-chronized survey product ion scan(scanning speed 30,000 u/sec). Theconcentration of the injected stan-dard is equivalent to an extractedsample with a concentration of 2pg/mL. This comparison also con-firms that simultaneous measure-ments of quantitative (MRM) andqualitative (synchronized surveyscan) data are possible withoutsignificant losses in sensitivity andre producibility (see figure 6).

Regular doping controls usingmodern high-end analytical in -struments such as the LCMS-8050make life difficult for doping of -fenders in equestrian sports. State -ments like ”… anything that is notdetected, is allowed” are definitelya thing of the past and even recre-ational riders must be cautiousthat their animals will not sudden-ly test positive after careless use ofmedication.

Further information

on this article:

•Poster (PDF)

•Table 1, with compari-

son of results (Link in

text, PDF)


W ater is the best-knowntest medium used in ele-mental measuring tech-niques in instrumental analysis.However, modern instrument sys-tems such as Shimadzu’s AA-7000(atomic absorption spectroscopy,AAS) or ICPE-9800 (optical emis-sion spectroscopy with inductive-ly coupled plasma, ICP-OES) of -fer considerably more applicationpossibilities.

What are these possibilities andwhat special solutions are avail-able? One answer is provided bythe following application over -view of the ICPE-9800, which canmeasure more than 70 elements atthe same time for any applicationwithin the shortest possible time(simultaneous elemental analysis).Due to the high dynamic measure-ment range of the ICPE-9800, it isirrelevant whether the elements tobe determined are present in thelower ppb range (μg/L) or in thehigher ppm range (mg/L).

The ICPE-9800 with its flexibleinstrument configuration is a truemulti-tool. For instance, operationcan be switched between hydridesystem and ultrasonic nebulizer aswell as between the various sam-ple types, such as water and organ -ic samples.

Quality assurance of sulfuricacid for special applications

Sulfuric acid (H2SO4) plays a ma -jor role in synthetically producedchemicals. Its application range is

Espresso in LakeConstanceFive application examples of simultaneous elemental analysis sensitivity

very diverse and the re quirementsfor high purity are not alwaysmet, as for instance in the produc-tion of phosphate and am moniumsulfate fertilizers. [1]

However, there are also many spe-cial application areas requiringhigh-purity sulfuric acids such assample preparation for instrumen-tal analysis. Here, the acid is, forinstance, used for sample diges-tion of polymers, fats or some geo -logical samples such as aluminumoxide. [2]

Using the ICPE-9800, a sulfuricacid sample was measured in a1 :10 and 1 : 20 dilution. Using thestandard addition method as cali-

bration model, matrix effects canbe masked. When the results ofboth dilutions are comparable, itcan be assumed that the method isaccurate. In order to permit a finalvalidated method, a certified refer-ence material can be measured inaddition. The results (summary)are listed in table 1 and are deter-mined for each element at variousanalytical wavelengths.

Although the diluted sulfuric acidsolution still is quite aggressiveand its viscosity differs from thatof water, measurement can be car-ried out using the standard config-uration (Minitorch). In additionto the consistent results of the dif-ferent dilutions, the recoveries for

samples with added standard arealso very good (100 ± 3 %).

Rapeseed oil for biodiesel –determination of phosphorus,sodium and potassium

Vegetable oils in fuels contributeto the sustainable operation ofcombustion engines. However,they should not influence engineperformance and catalyst efficien-cy in order to guarantee the envi-ronmental friendliness of biofuels.[3]

The determination of phosphorusin rapeseed oil is therefore essen-tial, as this element impairs thefunctioning of catalysts. High lev-els of calcium and magnesium arealso detrimental, since both areassociated with increased ash de -posits in soot particle filters. [3]

For measurement using ICP-OES,the sample only needs to be dilut-ed. The use of an additional com-bustion gas (oxygen) is not neces-sary in this case. Nevertheless, thehighly sensitive axial plasma �

ICPE-9800 – the multi-tool


APPLICATION

APPLICATION


ob servation mode can be applied.Measurement results and detec-tion limits are compared in table2. When selecting a more sensitiveworking range, additional andmore sensitive wavelengths can be

used, whereby the detection limitis further reduced. The stability ofmeasured values for the analysisresults is 100 ± 2 % (measuringinterval 1 hour).

The results show that the limitvalue can be determined very sat-isfactorily using the ICPE-9800and that the broad measuringrange makes it possible to alsounequivocally and reliably meas-ure increased concentrations.

Based on the analysis results, thesample can be assessed and isfound to be clearly unsuitable foruse in biofuels. The considerablyincreased concentration of thethree elements may, for instance,be due to the increased proportionof immature seeds used in produc-tion. Other influencing factors arethe shelling of the seeds and pro-portion of broken grains as well asthe pressing parameters such asthe pressure head temperature.

When using the ICPE-9800 foranalysis of other types of organicsamples, for instance samples fromthe petrochemical industry, the

use of an Ar/O2 mixed gas supplykit is recommended.

Along with the standard argoncarrier gas, an additional gas is in -troduced via the quadruple torchfor sample transport and mainte-nance of the plasma. It in creasesthe decomposition of the matrix,thereby reducing backgroundnoise.

By using this instrument configu-ration, the plasma can be observedaxially and radially despite thecarbon-rich sample material, anddetection limits in the lower ppbrange can be achieved. For exam-ple, for tin in toluene, a detectionlimit of 2.0 ppb could be attained.Other sample types for this in -strument configuration are, forinstance, kerosene, xylene, methylisobutyl ketone (MIBK), isopro -pyl alcohol (IPA), ethyl alcohol oralso the volatile tetrahydrofuran(THF).

Table 1: Analysis results of sulfuric acid. For determination of the recovery, 0.5 mg/L (ppm) of the element was added. n.d. = not detectable.

H2SO4 Sample A

Spike Recovery

H2SO4 Sample A

Spike Recovery

20

10

n.d.

98 %

n.d.

97 %

2.62 ppm

102 %

2.62 ppm

97 %

0.052 ppm

99 %

0.073 ppm

99 %

0.056 ppm

100 %

0.038 ppm

98 %

1.68 ppm

100 %

1.79 ppm

98 %

0.032 ppm

99 %

0.033 ppm

97 %

Dilution factor Cadmium, Cd Chromium, Cr Copper, Cu Magnesium, Mg Nickel, Ni Zinc, Zn

Elemental analysis in limitedsample volumes – 1 mL andless!

The intelligent instrument designof the ICPE-9800 not only re -duces the consumption of argonvia its Minitorch, but also ensures

efficiency in other aspects: a CCDchip continuously detects the en -tire spectrum and therefore ac -quires information on the elementcontent of a sample significantlyfaster than the earlier sequentialICP-OES technique.

Unlike conventional sampleamounts of 5 - 10 mL, the ICPE-9800 only needs 1 mL or less sam-ple volume, depending on the ap -plication. In this way, the smallestsample amounts can be investigat-ed without the need for dilution.In addition to the timesaving ef -fect, not having to dilute the sam-ples eliminates the danger of con-tamination and the loss of sensi-tivity. Within the shortest possibletime, it is possible to obtain mean-ingful results from a triplicate de -termination including axial and ra -dial plasma observation.

The measurement of small sampleamounts is not only useful for lim -ited volumes. When larger vol-umes of test media are to be meas-ured for changes in element con-tent over a longer period of time,it is also advisable to only usesmall samples in order to mini-mize the effect on the test system.

Rare earths in electronic waste – an alternative source of raw materials?

Which elements are actually con-tained in waste? This is a keyquestion in a world with limitedquantities of raw materials andtheir continuously increasing con-sumption. Bottlenecks in the sup-ply of rare earths are already im -portant topics covered in the me -dia.

Rare earths belong to a group ofelements that include neodymium,dysprosium or cerium, and addspecial features to advanced elec-tronic components. They make itpossible to design small and com-pact mobile phones that are, at thesame time, quite powerful. But

Table 2: Results obtained with the ICPE-9800. Analysis results of rapeseed oil as well as detec-

tion limits and maximum admissible content according to DIN 51627-6 (*dilution factor regarded).

PhosphorusCalciumMagnesium

3.0 mg/kg1.0 mg/kg1.0 mg/kg

0.085 [177.499 nm]0.025 [183.801 nm]0.005 [285.213 nm]

13.617.4

1.80

ElementLimit value toDIN 51627-6

Detection limit*[mg/kg]

Analysis result*[mg/kg]

these elements are also used atton-scale, for instance in the greenenergy sector as in generators ofmodern wind turbines.

As raw material sources are be -coming limited, recycling possibil-ities are moving into focus, espe-cially when extraction of rareearth oxides and subsequent puri -fication to pure metal form re -

quires extensive use of chemicals.In 2010, the EU classified rareearths as raw materials with a highsupply risk [5]. Todays REE recy-cling rate is around 1 %.

Is recycling from electronic scrapworthwhile? To answer this ques-tion, various electronic compo-nents were crushed (homoge-nized) and digested in a laborato-ry microwave oven. The digestionsolution can be measured usingthe ICP-OES and yields the re -sults listed in table 3.

This study shows that the analysisof solids using the ICPE-9800 ispossible and that modern wastecontains many elements which arein high demand on the interna-tional raw materials market. Notevery sample may contain a vari-ety or a large amount of rareearths, but recycling of used mo -bile phones (>1 g/kg Nd) couldbe quite conceivable after selectivepresorting. For more informationon analysis of REE in e-wasteplease see literature note.

Literature[1] Schwefelsäure (sulfuric acid) In: Römpp

Online. Georg Thieme Verlag, retrieved

9 May 2014.

[2] Application database CEM Corporation

[3] Remmele, Die neue Norm für Rapsölkraft -

stoff DIN 51605, Technologie- und Förder -

zentrum TFZ, Straubingen,

http://www.duesse.de/znr/pdfs/2010/2010-

11-25-biokraftstoffe-03.pdf

[4] Qualitätssicherung bei der dezentralen

Pflanzenölerzeugung, 12. Bericht aus dem

TFZ, Straubingen

http://www.tfz.bayern.de/mam/cms08/biok

raftstoffe/dateien/12_bericht.pdf

[5] Report of the Ad-hoc Working Group on

defining critical raw materials: Critical raw

materials for the EU, 2010

http://ec.europa.eu/enterprise/policies/raw

materials/files/docs/report-b_en.pdf

the environmental sector are notincluded in the above list. But it isalso possible for these elements toattain detection limits in the pptrange. In this case, the ability ofthe elements to pass from the liq-uid into the gas phase after con-version with sodium borohydrate(NaBH4) is taken advantage of.Instead of the aerosol, the gasstream is introduced into the plas-ma, completely separated from thematrix. As, Sb, Se, Sn and Hg canbe detected in the ppt range usingthis variant.

ICPE-9800 is therefore a goodand economical alternative toICP-MS for detection in the ultratrace range in order to addressspecific analytical problems.

Ultra-trace analysis in the ppt range

When elemental analysis in the pptrange (ng/L) rather than in theppb range (μg/L) is required,questions can often be answeredusing ICP-MS. But this concen-tration range can also be coveredusing advanced ICP-OES such asthe ICPE-9800. In this way, detec-tion limits for more than 10 ele-ments (Ba, Be, Ca, Eu, Lu, Mg,Mn, Sc, Sr, Y, Yb) lie within singleor double-digit ppt range usingthe standard configuration includ-ing the Minitorch. For more sen-sitive detection of other elements,various other approaches areavailable.

Special nebulizers generating sam-ple aerosols more efficiently andreproducibly with a smaller drop -let size distribution reduce detec-tion limits very easily without theneed to significantly change theinstrument configuration. An ad -ditional ultrasonic nebulizer can

Table 3: Analysis results in mg/kg for rare earths in electronic scrap.

n.d. = not detectable.

Mobile phone

LCD (Display)

Printed circuit board

Sample

1.040

33

n.d.

Nd

107

51

n.d.

88

n.d.

n.d.

48

n.d.

n.d.

4,6

7,8

1,3

4,6

n.d.

n.d.

n.d.

86

32

La Pr Dy Y Er Ce

be connected to the ICPE-9800,enabling more sensitive measure-ments by up to a factor of 20. Be -sides the elements mentionedabove, which can be calibrated inthe ppt range using the standardconfiguration, 27 more elementswith detection limits in the dou-ble-digit ppt range and lower areadded (Ag, Cd, Ce, Co, Cr, Cu,Dy, Er, Fe, Gd, Hf, Ho, La, Li,Mo, Nb, Nd, Pd, Pr, Sm, Ta, Tb,Ti, V, Zn, Zr). Strontium has thelowest detection limit of 1 ppt(parts per trillion – this is theequivalent of a cup of espresso inLake Constance [Bodensee] sur-face area = 536 km2).

Elements such as arsenic and mer-cury that are highly important for


APPLICATION

Bilirubin determination in cerebrospinal fluid following asubarachnoid hemorrhage Stroke, aneurysm – UV spectroscopy supports fast response

Subarachnoid hemorrhage is a pathological event in the central nervoussystem. It is characterized by bleeding into the subarachnoid space, whichis filled with cerebrospinal fluid (CSF, liquor cerebrospinalis). Subarachnoidhemorrhage accounts for up to 10 percent of all strokes.

This particular form of stroke is, in most cases, caused by rupture of anarterial vessel due to an abnormality. Subarachnoid hemorrhage is accom-panied by sudden onset of a very severe headache and neck stiffness. This can lead to short-term decreased levels of consciousness, but also tosevere permanent brain disorders (quoted from Wikipedia).

Bilirubin determination, such as the determination of bilirubin in CSF, is ananalytical method used in clinical analysis. In CSF-bilirubin determination,cerebrospinal fluid is analyzed for blood fragments. CSF bilirubin = cere-brospinal fluid bilirubin.

In case of illness, red blood cellswill initially enter the cerebrospi -nal fluid. These cells contain oxy-hemoglobin, which is converted tobilirubin by enzymatic processes.In addition to bilirubin, methe-moglobin may also be formed.

As the process is time-dependent,the bilirubin concentration andthe presence of oxidation productssuch as oxyhemoglobin provideinformation on the state of thepatient. In addition, it is possibleto estimate the duration of theoxidation process. �

T he UV spectroscopic appli-cation presented here is basedon the ‘National Guidelinesfor analysis of Cerebrospinal Fluidfor Bilirubin in suspected Sub -arach noid Hemorrhage’ ac cordingto Beetham [1].

The causes of cerebral hemorrhagecan be diverse, such as a stroke oran aneurysm. In order to take ap -propriate measures for treatmentof a patient, it is important for thephysician to determine which typeof blood (old, fresh or none) ispresent in the cerebrospinal fluid.

Further information

on this article:

• Application Note

ICP-OES Spectroscopy

(PDF)

• Poster Rare Earth Elements in Electronic

Waste (PDF)


APPLICATION

This shifting baseline presents achallenge for the automated analy-sis of very small maxima or shoul-ders in a signal pattern. Variousanalytical methods are applied toseparate these pigments (HPLC).Fast testing methods can also beperformed, such as use of a gratingspectrometer for the rapid screen-ing of hemoglobin.

Defined measuring method

Beetham has defined the measur-ing procedure according to whichmeasurement should be carriedout in the visible range from 350to 600 nm. The analytical wave-lengths for the hemoglobin colorpigments are listed in table 1. Themethod requires corrected signalheights, realized via the baseline/tangents under the signals/bands.Net bilirubin absorption is deter-mined using a fixed wavelength of476 nm.

Multi-component measurementrequired

The color pigments mentionedabove are suitable for spectroscop-ic analysis in the visible range.Quantitative but also qualitativeassessment as well as color repre-sentation are the domain of UV-VIS spectroscopy. Since cerebro -spinal fluid is a mixture not onlyrepresented by hemoglobin, ittherefore requires a multi-compo-nent measurement, as all spectra ofthe individual components overlap(superposition). In this case, astraight baseline as would be ex -pected in the analysis of liquids isnot measured, but rather a risingbaseline extending into the UVrange. This effect occurs due tosubstances responding to UV irra-diation, such as proteins or otherhigh-energy compounds (pi elec-trons) in materials whose spectraare expected to be similar tohemoglobin spectra.


Table 1: Analytical wavelengths for hemoglobin

OxyhemoglobinBilirubin

Methemoglobin

Pigment

between 410 and 418 nmbroad bands of 450 to 460 nm or as a shoulder in the oxyhemoglobin signalmore rarely occurring pigment; when present it occurs as a broader signal between 403 to 410 nm

Absorption range

Figure 1: Representation of a UV-VIS absorption spectrum of a bilirubin sample exhibiting

an oxyhemoglobin signal at 415 nm and a bilirubin shoulder at 450 to 460 nm recorded

using the UVProbe software

nm.

0.014

0.020

0.030

0.040

0.050

0.060

350.0 400.0 450.0 500.0 550.0 600.0

T %

Figure 2: Representation of an absorption spectrum of a bilirubin sample and the

representation of the baseline used for the correction of the bands

nm.

Oxyhemoglobin with NBA below thereference value

[Abs

]

0

0.01

0.02

0.03

0.04

0.05

0.06

350 400 450

––– Abs

500 550 600

Table 2: Calculation of the net bilirubin content (NBA) after graphical evaluation

365 0.0289 476 0.021NBA at 475 nm

0.00385714Limit value< = 0.007

EvaluationOK

Base point A = 350 - 400 Base point B = 430 - 530

Beetham specifies the evaluationof the spectra. One specification isthat the baseline under the signalshould never intersect the signalitself. This strict instruction infersthat automated evaluation cannotlead to the correct result. Manualevaluation is called for, in whichthe base points under the signalcan be specifically adjusted/cor-rected.

To solve this problem, an Excelspreadsheet was developed inwhich it is possible to measure, torepresent the spectrum, and tomanually shift the baseline. With -in the evaluation range, all re -quired values of the sample couldbe represented.

A representation of the spectrumfrom the Excel sheet is shown infigure 2.

The evaluation is presented intable 2.

Shimadzu’s UV-1800 spectropho-tometer was used for the analysis.The spectrum was measured in therange of 350 to 600 nm. The sam-ple was measured using a 1 cmcuvette. The limit value in thisexample was below 0.007 absorp-tion units.

Summary

Using the Excel method, the cal-culation of the net bilirubin con-

centration can be easily deter-mined. Using a macro simplifiesthe adjustment to local conditions,such as different output formatsand adaptation to individualforms. On the other hand, it iseasy to extend the macro withother applications. The directinstrument control of the UV-1800 allows for direct data evalua-tion. It is also possible to importthe results from the UV-Probesoftware, which is supplied asstandard with the UV-1800.

Literature[1] „National Guidelines for Analysis of

Cerebrospinal fluid for Bilirubin in Sus -

pected Subarachnoid Haemorrhage“,

Draft 1; R. Beetham, M.N. Fahie-Wilson,

I. Holbrook, I.D. Watson, P.R. Wenham,

P.A.E. White, P. Thomas, A.M. Ward,

A. Cruickshank, G. Keir, W. Egner, K. Allen


APPLICATION

New: TOC Application Handbook,Version 2 Online and Laboratory TOCs

A s a leader in TOC technol-ogy, Shimadzu has beensetting standards with itsanalyzers for decades. The wealthof Shimadzu’s experience continu-ously flows into the developmentof its TOC systems. In conse-quence, Shimadzu’s online analyz-ers, as well as laboratory TOCsystems offer the highest flexibili-ty, high availability, tremendousrobustness and stability, straight-forward and intuitive operationaccompanied by advanced controland evaluation software. Usersbenefit from personal support;many additional functions facili-tate their work and offer latitudefor other important tasks.

In 2012, Shimadzu has summa-rized its available experience andinformation in the first TOC Ap -plication Handbook. In the cur-rently published second revision,the existing chapters have beenextended and now include 50 ap -plication and information notes inmore than 100 pages. The TOCApplication Handbook containsthe following chapters:

2. Pharmaceutical industryThe TOC determination is de -scribed in the European Pharma -copoeia (EP). The sum parameterserves as a measure for contami-nation by organic components. Not only the method itself is de -scribed, but also a test to verifythe suitability of a TOC analyzerfor the analysis. Recent changes inthe American Pharmacopoeia andtheir effects on validation are pre-sented in a new application note.

concentration ranges, conditionsare always diverse, such as salinityor number of particles.

1. Environmental analysisTOC analysis is carried out in awide variety of environmentalmatrices – from groundwater toseawater, from drinking water towastewater, from soils to sewagesludge. This chapter discusses thediversity in environmental applica-tions and their many varying chal-lenges. In addition to the different

TOC Application Handbook

Furthermore, the various samplersfor TOC analysis in cleaning vali-dation are highlighted.

3. Chemical industryIncoming goods control plays animportant role in the chemical in -dustry. Impurities present in re -agents often also constitute theimpurities in products. In additionto the targeted analysis of knowncompounds, sum parameters canhelp to assess the raw chemicalswith regard to their impurities.TOC is important here: this pa -rameter describes the contamina-tion by organic compounds andspecifies the total amount of or -ganic carbon. TOC can, therefore,also be used for the assessment ofinorganic chemicals.

Descriptions for TOC determina-tions in phosphoric acid and in di -luted hydrofluoric acid have beenadded. �

APPLICATION

20

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WMFNovember 10 -12, 2014Vienna, Austriawww.shimadzu.eu/world-mycotoxin-forum

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4. TOC special applicationsDue of its informative signifi-cance, the TOC sum parameter iswidely applicable. It mirrors thetotal concentration of organicallybound carbon or organic com-pounds.

Besides the environmental, phar-maceutical or chemical industry,the TOC parameter is used innumerous other complex applica-tions. Often, it is the scientificcuriosity and ingenuity of userswho want to solve an analyticalproblem or simplify analytics thatleads to finding the key to theanswer in the TOC pa rameter.

5. TOC in daily practiceThis chapter deals with the indi -vid ual modules, kits, options andsup ported functions of Shimadzu’sTOC analyzers. Concepts and

methods are also described. In -for mation on related sum param-eters such as BOD (biochemicaloxygen demand) and COD (chem -i cal oxygen demand) are includ-ed.

Many users want to calculate theCOD from the TOC and, there-fore, need correlation factors.These correlations depend on var -ious parameters. Examples showhow the COD factor can bemathematically determined. Theyfurther illustrate why COD fac-tors may be so different, due tothe sample components. Hence,the bandwidth of the conversionfactors ranges from < 1 to > 5, de -pending on how much oxygen isalready bound to the organiccompounds. In addition, inor-ganic compounds such as nitrites,bromides, iodides, metal ions and

sulfur compounds are also oxi-dized and co-determined in aCOD determination, and there-fore may also have an effect theconversion factor.

6. TOC process analysisParticularly during process con-trol, it is important to obtain fast,continuous and informative dataon the organic pollution levels ofwaters. In TOC process analysis,the sample is continuously fed tothe measuring system for subse-quent measuring. This way, thecontrol room can react promptlyto any possible process changes.

One of the most important attri -butes of a TOC process analyzeris its versatility. Since a TOC pro -cess system is not available ‘offthe shelf’, each measurement mustbe customized to the particular

measuring problem, the ma trixand the sampling location. Theanalyzer can be tailored to thespecific measuring task. Manyanalyzers are used in wastewatercontrol where, depending on thetype of industry, the wastewatermay have a very different matrix.A further application area is con-tinuous TOC determination of‘clean’ water, such as conden-sates. For this purpose, there isan option for which the specifica-tions were described in a separateapplication note.

Further information

on this article:

• TOC Application

Handbook (PDF)

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