The effect of salinity, nitrate concentration, pH and temperature on eicosapentaenoic acid (EPA) production by the red unicellular alga Porphyridium purpureum
Introduction
The red unicellular alga Porphyridium purpureum is a potential source of eicosapentaenoic acid (EPA) and arachidonic acid (AA), which are essential fatty acids for human nutrition (Radwan, 1991). EPA is effective in preventing blood platelet aggregation and reducing blood cholesterol and AA has been proposed to be a precursor for prostaglandin PGE2 biosynthesis (Cohen, 1988). The fatty acid composition of P. purpureum is highly dependent on environmental conditions, such as salinity, temperature and pH (Cohen, 1988; Cohen et al., 1988; Lee and Tan, 1988; Lee et al., 1989). In nature, Porphyridium occurs in a diverse spectrum of habitats and has been isolated from freshwater, brackish and marine environments, as well as from the surface of moist soils or pots in greenhouses (Vonshak, 1988).
In this work we optimized the conditions for EPA production by P. purpureum using both systematic and stepwise optimization. The EPA production per cell and total EPA production per culture volume were used as optimization parameters instead of production per dry weight, since dry weight as a parameter for biomass is biased during the cultivation due to extracellular polysaccharide synthesis. For the same reason Ahern et al. (1983)also found it better to express production in terms of cell number or culture volume rather than dry weight. The systematic optimization was used to determine the optimum concentrations of sodium chloride and nitrate in the medium. The production pH and temperature were thereafter optimized stepwise.
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Maintenance and selection of algal strains
Five strains of the alga Porphyridium purpureum (Drew et Ross), 1380-1a, 1380-1b, 1380-1c, 1380-1d and 1380-1f, were obtained from the Sammlung von Algenkulturen, Pflanzenphysiologisches Institut der Universität Göttingen. The cultures were maintained in liquid culture on ASW medium (Jones et al., 1963) (27 g l−1 NaCl, 9.9 mM nitrate, pH 7.6). The cultures were subcultured every 3 weeks by transferring cells to fresh medium to obtain 2×105 cells per ml. In order to select a suitable strain for
Optimization of sodium chloride and nitrate concentrations for EPA production
Second order polynomial mathematical models for growth, EPA production and AA production were derived from the experimental results (Table 3) using regression analysis (SAS/Stat program package). The equation for growth (Y1, cell number per ml) as a function of sodium chloride (x1) and nitrate (x2) concentration is:where R2=0.81***; ***P<0.001; **P<0.01 and *P<0.05. R2 is the percentage of the total variation in the
Discussion
It is known that the C20-polyunsaturated fatty acids, including EPA, are mainly associated with chloroplast and membrane constituents such as galactolipids and phospholipids. Therefore factors that promote the biosynthesis of cell polar lipids and/or increase membrane fluidity also promote the biosynthesis of these fatty acids (Radwan, 1991). It has been demonstrated in several studies that high EPA concentrations are closely related to good growth conditions and to high photosynthetic activity
Conclusions
Using a combination of systematic and stepwise optimization we were able to increase the EPA production per cell from the value of 0.2 pg per cell to a value of 1.5 pg per cell. The volumetric production increased from the 40 μg per 50 ml in the ASW medium at 25°C to 298 μg per 50 ml in production medium (41.8 g l−1 NaCl, 12.1 mM nitrate) at 8°C. These optimum conditions were also tested in bioreactors. In bioreactors the EPA production per cell remained on the same level as in the shake flasks
Acknowledgements
The skillful technical assistance of Ms Leena Toivonen is gratefully acknowledged.
References (13)
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- Cohen, Z., 1988. Production of eicosapentaeonic and arachidonic acid by the red alga Porphyridium cruentum. In:...
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