University of Southern Denmark
Natural blue food color from cyanobacteria Spirulina
platensis
Malwade, Chandrakant Ramkrishna; Roda Serrat, Maria Cinta;
Christensen, Knud Villy;Fretté, Xavier; Christensen, Lars
Porskjær
Publication date:2015
Document version:Final published version
Citation for pulished version (APA):Malwade, C. R., Roda Serrat,
M. C., Christensen, K. V., Fretté, X., & Christensen, L. P.
(2015). Natural blue foodcolor from cyanobacteria Spirulina
platensis. Poster session presented at 6th Nordic Natural
ProductsConference, Visby, Sweden.
http://www.fkog.uu.se/nnpc/NNPC2015-abstractbook.pdf
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Natural Blue Food Color from Cyanobacteria Arthrospira
platensis
Chandrakant R. Malwade, Maria C. R. Serrat, Knud V. Christensen,
Xavier Fretté, Lars P. Christensen
Department of Chemical Engineering, Biotechnology, and
Environmental Technology University of Southern Denmark, Campusvej
55, Odense M, Denmark
[email protected]
MOTIVATION
INTRODUCTION
EXPERIMENTAL
Blue colour is an important part of the food color palette used
in
products such as icecream, confectionaries, chewing gum and
soft
drinks. Available blue colors in the market are chemically
synthesized of which EU have the following: Patent blue V,
Indigotine, Brilliant blue FCF. Recently concerns have been
raised
about the safety of synthetic blue colors [1] and together with
the
growing demand among consumers for natural food colors this
has
increased the need for the development of natural blue
colors.
Composition of Linablue Additives removal from Linablue Cleavage
of Phycocyanobilin (Methanolysis)
Patent blue V (E131)
Indigotine (E132)
Brilliant blue FCF (E133)
Aim of this project is to develop a stable natural blue food
color from cyanobacteria Arthrospira platensis (syn. Spirulina
platensis).
Arthrospira is a blue-green microalgae widely used as dietary
supplement due to its high protein content. Arthrospira extract has
already
been approved by USFDA as a food color, but its lack of
stability is a problem [2]. Phycocyanobilin, responsible for blue
color of the
extract, is obtained in pure form and stabilized through
copigmentation and lake formation. In this project, pure
phycocyanobilin (PCB) is
obtained from a commercial extract of Arthrospira
(Linablue®G1).
Linablue is washed with methanol to
remove additives; methanol selectively
dissolves D-trehalose and trisodium citrate.
Procedure included stirring 50 g Linablue
in 400 mL methanol for 30 min.
Procedure is repeated 5 times; 20 g
additive free Linablue is obtained.
Methanol washing also aid in denaturation
of phycocyanins as depicted below.
10 g washed Linablue boiled in
400mL methanol for 16hrs at 65 °C
Samples are taken at regular interval
from mixture for HPLC analysis
After 16hrs, methanol solution is
analyzed with HPLC and LC-MS
Linablue powder
Methanol wash D-Trehalose & trisodium citrate
Protein in normal state
Denatured protein
Purification of methanolysis mixture
666.02
0,0 1,0 2,0 3,0 4,0 5,0 6,0 7,0 8,0 9,0 10,0
0
25
50
75
100
140
1 - 3,917
2 - 4,583
3 - 6,203
min
mAU
300 400 500 600 700
629,60 349,72
277,52
nm
300 400 500 600 700
368,71
666,02
nm
300 400 500 600 700
364,19
650,98
nm
1 2
3
Cultivation of
Arthrospira platensis
Phycobilisomes (Light harvesting complexes of
phycobiliproteins)
Phycocyanobilin (Chromophore)
Extraction
Cleavage & Purification
C-Phycocyanin (Phycobiliprotein)
PCB
Methanolysis mixture is purified by using flash
column chromatography
RP C-18 column with acetone and water is used as
mobile phase
85mg phycocyanobilin is obtained
0,0 1,0 2,0 3,0 4,0 5,0 6,0 7,0 8,0 9,0 10,0
0
125
250
375
500
625
700 1 - 4,563
min
mAU
200 400 600 800 1000 1200 1400 1600 1800 2000
m/z
0
20
60
80
587.61
1173.45
1760.33
1195.39
40
100 PCB + H+
FUTURE PERSPECTIVES
Physico-chemical properties of phycocyanobilin will be
determined
Further optimization of cleavage and purification process to
obtain pure
phycocyanobilin will be done.
Stabilization and intensification of phycocyanobilin color will
be attempted
through its copigmentation with naturally occurring
molecules.
REFERENCES
1. European Food Safety Authority (2013) EFSA Journal, 11,
2818.
2. Newsome, G. et al. (2014) J. Agric. Food Chem., 62,
6498−6511.