Isochrysis galbana MICROALGA -...
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Factor Optimum Press 300.0 bar Temp 50.0 oC Optimize Desirability Optimum value = 0.662769 Optimum 3.98 % Yield 621.8 mg carotenoids/g extract 284.8 mg chlorophylls/g extract
OPTIMIZATION OF SUPERCRITICAL FLUID EXTRACTION
PROCESSES TO OBTAIN BIOACTIVES FROM
Isochrysis galbana MICROALGA
J.A. Mendiola1, G. Sullini1.2, A. Cifuentes1, E. Ibáñez1 1 Instituto de Investigación en Ciencias de la Alimentación CIAL (CSIC-UAM). C/Nicolás Cabrera 9, 28049 Madrid.
2 Università degli Studi di Messina, Messina, Italia.
MIRACLES WPs and Activities
WP1: Production and Harvesting
WP4: Product development and market assessment
Cell disruption Extraction
Hydrophobic compounds
(lipids)
Hydrophilic compounds (proteins,
carbohydrates)
Food, incl. functional foods, nutraceuticals
Aquaculture / aquafeed
Non-Food applications
WP3: Development of Integrated Biorefinery
Harvesting
WP2: Bio-prospecting, metabolic optimization, cultivation at extreme
locations
New production strains
Industrial strains
NATAC, URDV, DSM
EWOS, SPAROS
BIOPOL, IMENZ, CHIMAR, NATAC, CE
Marke
t assessm
en
t
VFT WU DLO CSIC ET
DNL DSM VFT WU, UT, UHU, VITO, FITO,
TMUC
WU, FCPCT, UIB, UA, UniRes, FITO
Sequential extractions
1st step: SC-CO2 extraction to recover omega-3 and highly lipophilic compounds, Experimental design
2nd step: SC-CO2+ ethanol extraction to obtain extracts rich in other bioactives (more polar).
Isochrysis galbana microalga
Isochrysis galbana is a marine flagellated microalga belonging to the phylum Haptophyta, class Coccolithophyceae, subclass Prymnesiophycidae, order Isochrysidales, family Isochrysidaceae. Its cells are easily assimilated by larval animals because of their small size (4-7 μm) and absence of a tough cell wall. I. galbana is represented by different strains, the most commonly cultured one is the Tahitian strain “T-iso”. Generally, this microalga is characterized by a fast growth rate, wide temperature and salinity tolerance and absence of toxins. As all microalgae, the growth and biochemical composition of I. galbana cultures is dependent on variations of culture conditions (e.g. nutrients, temperature, salinity, and photoperiod).
This Haptophyta is of substantial interest in aquaculture, principally to feed mollusk larvae, as well as fish and crustaceans in the early stages of growth.
Isochrysis galbana is a marine microalga well recognized as a natural rich source of EPA and DHA that are accumulated as oil droplets in prominent lipid bodies in the cell. This unicellular microalgae is investigated also for its high amount of the carotenoid Fucoxanthin.
Supercritical fluid extraction
Foodmics Laboratory activities
Sequential extractions: 1st step optimization
3-level factorial design (32). The studied factors were P (100-300 bar) and T (40-60 ºC) Other factors fixed: 10 g of algal material used with 30 g of sea sand as dispersive agent, 5 L/min CO2(gas) Extraction time optimization: Kinetic study. Extractions performed at different times in the center-point of experimental design (200 bar, 50 ºC). Extraction time fixed at 30 min (75 % extractable material recovered)
40 ºC 50 ºC 60 ºC
200 bar 40 ºC 50 ºC 60 ºC
100 bar 40 ºC 50 ºC 60 ºC
300 bar
Statistical optimization of experimental design for YIELD
Yie
ld %
Pressure (bar)
R2 of mathematical model 93.2% Significant factors: Pressure and P*P Goal: maximize yield Optimum value = 4.51 % Optimum for yield: P= 254.2 bar T= 55.4 ºC
Multiple response optimization: Yield, Carotenoids and chlorophyls Extraction conditions optimized for 1st extraction step to: maximize yield and carotenoids and minimize chlorophylls
• Extraction of polar compounds based on Gas Expanded Liquids
Liquid Gas Expanded Liquid (GXL)
• Extractions performed on the residue from 1st extraction step • Experimental design
• 15-70% Ethanol • Pressure high enough to expand EtOH, but not not to dissolve CO2 (ca. 60-80 bar) • Same temperature as 1st step optimum (50 ºC) to reduce cooling time
Composition of extracts: carotenoids up to 63%, chlorophylls up to 32 %. PolyUnsaturated Fatty Acids (linolenic, DHA…)
detected by GC-MS metabolomic method. Clear trend at 50 ºC: higher pressure, higher
carotenoid concentration.
1st
Ste
p
2n
d S
tep
Composition of extracts: Extracts very rich in chlorophylls Polyunsaturated Fatty Acids (linolenic,
DHA…) detected by GC-MS metabolomic method at lower concentration than 1st step
GC-MS metabolomic method also reported the presence of several carbohydrates
Acknowledgements This work has been financed by an FP7 project: 613588 MIRACLES http://miraclesproject.eu/
CONCLUSIONS
The recovery of bioactive compounds from the microalga Isochrysis galbana has been set up. In the 1st step pure supercritical CO2
was used and extracts rich in carotenoids were obtained. In the 2nd step the application of Gas Expanded Liquid Extraction was studied for the first time on this microalga, providing extracts with very high content in chlorophylls and carbohydrates.