extraction of proteins from palmaria palmata
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Extraction of proteins from Palmaria palmata
Rósa Jónsdóttir
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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
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Utilisation of Palmaria palmata
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• Palmaria palmata - dulse
• Good nutritional value
• Suitable for human food and animal feed application
• Utilized as food in Iceland for centuries
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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
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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
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• 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
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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of the familyThermoactinomycetaceae according to 16S rRNA sequence analysis and blasting to the NCBI nrdatabase (Matís, unpublished results).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
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Flow chart of the extraction process
Freeze drying Freeze drying
100 µm sieve
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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
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Amino acid composition and protein content of P. palmata
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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
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Amino acid composition and protein content of P. palmata
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• 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)
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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
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Bioactivity of different fractions of P. palmata antioxidant activity
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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
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ACE (angiotensin converting enzyme) inhibition
activity of different fractions of P. palmata
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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
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Conclusion
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• 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
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Publications
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Acknowledgement
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Málfríður Bjarnadóttir, MatísBjörn Viðar Aðalbjörnsson, MatísAnna Nilsson, MatísGuðmundur Óli Hreggviðsson, MatísÓlafur H. Friðjónsson, Matís
Rasa Slizyte, SINTEF OceanMichael Y. Roleda, NIBIO