Extraction of proteins from Palmaria palmata
Rósa Jónsdóttir
Background
2
• Increasing demand for high quality protein both for food and feed
• The access to high quality protein is becoming more challenging
• Important to find alternative food sources including protein alternatives
that contain all the essential amino acids fulfilling human requirements
Utilisation of Palmaria palmata
3
• Palmaria palmata - dulse
• Good nutritional value
• Suitable for human food and animal feed application
• Utilized as food in Iceland for centuries
Background
✓High protein content (up to 35%)
– Comparable to high protein vegetables e.g. soybeans
✓Main polysaccharide is xylan (34-35%)
✓ P. palmata has a rigid cell wall consisting mainly of β-(1→4)/β-(1→3)-D-xylans along with some fractions of cellulose and β-(1→4)-xylans
✓Due to the strength of the cell wall, traditional protein extraction methods might give limited results
Objectives
✓Develop fractionation processes for preparation of protein-rich products and extracts to be used as food, nutraceutical or feed ingredients
✓ Explore the use of enzymes (xylanse and protease) as processing tool
✓ Study protein yield, quality and the bioactivity of the different fractions
• P. palmata from Norway (NIBIO)
• Preprocessing: wet-milling
• Enzyme hydrolyzation
• Filtration
• Chemical analysis (water, fat, salt, ash, protein)
• Protein
– SDS-PAGE; Nitrogen;
–Amino acids (SINTEF)
• Bioactivity (TPC, ORAC, DPPH, MC, ACE)
Material and methods
Enzyme ProducerOptimum conditions
Activity
pH Temperature
Umamizyme Amano enzymes Inc. 7.0 50Endo- and exo-peptidase
complex
ProteAX Amano enzymes Inc. 6.0 60Endo- and exo-peptidase
complex
Xylanase Matis ltd 6.5 60 Endo-1,4-β-xylanase
Xylanase is derived from a bacterium belonging to the genus Planifilum, of the familyThermoactinomycetaceae according to 16S rRNA sequence analysis and blasting to the NCBI nrdatabase (Matís, unpublished results).
EnzymesOptimum conditions and activity
Flow chart of the extraction process
Freeze drying Freeze drying
100 µm sieve
Extraction yields of dry matter of both fractions liquid extract and solid phase
0
10
20
30
40
50
60
70
Control Umamizyme Xylanase Xylanase andUmamizyme
Umamizyme
Extr
acti
on
yie
ld (
%d
w)
Yield
Liquid extract
Solid phase
Amino acid composition and protein content of P. palmata
10
Values for untreated seaweed (n=1) are presented as % of total amino acid content.Values for control, xylanase, xylanase + Umamizyme and Umamizyme (n=2) are presented as % of total amino acid content. Values for nitrogen content are presented as % dry weight ± SD (n=4).
Essential amino acids, % of total amino acid content
Amino acid composition and protein content of P. palmata
11
• All EAA were present in the samples
• EAA ratio higher in all solid phase samples compared to liquid extract
• EAA ration higher in liquid extract when treated with protease
― Protease break down proteins into peptides and amino acids passing
the 100 µm sieve after the hydrolysis
=> resulting in higher amino acid content in liquid extract.
Essential amino acids (EAA)
Protein content of P. palmata, untreated and enzymatically treated, using different N conversion factor
54.9
33.4
0
5
10
15
20
25
30
35
40
45
50
55
60
Untreated Liquidextract
Solidphase
Liquidextract
Solidphase
Liquidextract
Solidphase
Liquidextract
Solidphase
Control Xylanase Xylanase +Umamizyme
Umamizyme
Pro
tein
co
nte
nt
(% d
w)
N = 6.25
Calculated conversion factor
Values are presented as
% dry weight ± SD (n=4).
Conversion factor 4.7 3.6 4.1 2.5 3.8 2.9 3.5 3.2 3.4
Bioactivity of different fractions of P. palmata antioxidant activity
13
0.0
1.0
2.0
3.0
4.0
5.0
6.0
7.0
8.0
9.0
Control ProteAX ProteAX andUmamizyme
Umamizyme Xylanase Xylanase andUmamizyme
Umamizyme
DP
PH
, IC
50
valu
es
Liquid extract
Solid phase
0
10
20
30
40
50
60
70
80
90
100
Control ProteAX ProteAX +Umamizyme
75
Umamizyme75
Xylanase Xylanse +Umamizyme
200
Umamizyme200
Met
al c
hel
atin
g ab
ility
(%
)
Liquid extact
Solid phase
DPPH antioxidant activity, IC50 valuesHigh antioxidant activity in liquid extracts
Metal chelating ability (%) High antioxidant activity in some solid phase and liquid extract
ACE (angiotensin converting enzyme) inhibition
activity of different fractions of P. palmata
14
IC50 (mg/mL)
Liquid extract Solid phase
Control >1.00 >1.00
ProteAX 0.45 ± 0.11 >1.00
ProteAX + Umamizyme 75 0.38 ± 0.02 >1.00
Umamizyme 75 0.69 ± 0.05 >1.00
Xylanase >1.00 >1.00
Xylanase + Umamizyme 200 1.14 ± 0.08 >1.00
Umamizyme 200 >1.00 >1.00
Blood pressure
Conclusion
15
• P. palmata contains high protein content of good quality that can be
extracted with good results using enzymatic treatment with xylanse
• Xylanase disrupted the rigid cell wall, resulting in higher protein yield in
the solid phase
• Protein extract from P. palmata is high in essential amino acids and
would therefore be suitable as food and nutraceutical ingredient
• The results indicate that hydrolysis with protease can be a beneficial
method to extract bioactive hydrolysates from P. palmata
Publications
Acknowledgement
17
Málfríður Bjarnadóttir, MatísBjörn Viðar Aðalbjörnsson, MatísAnna Nilsson, MatísGuðmundur Óli Hreggviðsson, MatísÓlafur H. Friðjónsson, Matís
Rasa Slizyte, SINTEF OceanMichael Y. Roleda, NIBIO
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