The effects of environmental variables upon the lipid class and fatty acyl composition of a marine microalga: Nannochloropsis oculata (Droop) Eustigmatophyceae. Hibberd

Abstract

Detailed analyses of the lipid class and fatty acid composition were carried out for the marine microalgal species Nannoch/oropsis oculata (Droop) (CCAP strain no. 849/1) of the division Eustigmatophyceae (Hibberd). The alga was grown in batch and continuous culture using a novel culturing apparatus, the cage culture turbidostat, the construction of which is detailed in full. The total lipid extract yielded by the alga varied in a growth-phase dependent manner within the range 25 % to 80 % of the lyophilised cell mass. Of this between 40 % and 70 % was recovered as fatty acid methyl esters (FAME) upon transesterification. The total fatty acid composition of N. oculata consisted mainly of 16:0, 16:1 and 20:5(n-3), these three fatty acids often accounting for greater than 80 % of the total fatty acid mass. Between 9 % and 50 % of the mass of total FAME was accounted for by 20:5(n-3), the balance being accounted for by variations in the relative proportions of 16:0, 16:1, 18:1, 18:2 and 20:4. During periods of low cellular division rate, such as the lag- and stationary-phases, the proportion of polyunsaturated fatty acids (PUFA) (mainly 20:5(n-3» decreased. The total fatty acids became increasingly saturated as higher proportions of shorter chain length fatty acids accumulated, mainly in triacylglycerols (TAO). Increased cellular proportions of total lipid resulted from TAO accumulation which occurred on account of preferential partitioning of carbon into TAO biosynthesis whilst cellular division was suspended. The fatty acid composition of the TAO was more saturated at high synthesis rate and vice-versa at lower rates. The galactolipids, monogalactosyldiacylglycerol (MODO) and digalactosyldiacyl glycerol (DODO) were rich in 20:5(n-3) during exponential cell division containing up to 77 % and 53 % 20:5(n-3) respectively. Phosphatidylcholine (PC) was the only cl~s to contain significant proportions of CIS fatty acids during exponential growth, thus implicating its involvement in the acyl chain elongation reactions between the Cl6 and <;0 fatty acids. Culture incubation temperature in the range 5 °C to 25°C did not influence the fatty acid composition of N. oculata. The effect of temperature upon culture dynamics at the lower culture incubation temperatures gave an apparent decrease in the PUFA content of the total fatty acid at a given point on the cultures growth curves. By expressing the data in tenns of culture doubling periods during the exponential-phases of growth it was found that temperature had no real effect upon fatty acid unsaturation or chain length. at either the total or the individual lipid class FAME level after the cells had passed through five doubling periods. Increasing the culture medium salinity from one quarter to one and a half times that of normal seawater decreased the un saturation and chain length of the fatty acids at both total and individual lipid class levels. The change resulted from the progressive accumulation of 18:1 and 18:2"at the expense of 20:5. Variation of salinity did not affect the dynamics of the cultures in the same respect as temperature in that a lag-phase was not observed on the cultures growth curves. However. such a phase was evident in the fatty acid profile of the cells in the period following inoculation. The 'effects of culture illumination intensity in the range 45 Jill m-2 sec-I to 170 Jill m 2 sec-! were examined under continuous culture conditions using the cage culture turbidostat Accumulation of saturated TAG by the cells at the higher illumination intensities gave an apparent decrease in the rate of PUFA biosynthesis. The polar lipid classes were found to be more highly unsaturated at higher illumination intensities. At lower illumination intensity TAG accumulation was reduced and the total fatty acid composition was accordingly more unsaturated. The fatty acid composition of the TAG component was more unsaturated but those of the polar lipid classes were less unsaturated than at higher illumination intensity. Increased illumination increased the degree of un saturation of the polar lipid cl~sses. Excess fixed carbon was partitioned into TAG biosynthesis. primarily as 16:0 and 16:1. The net accumulation of this lipid class even at high cell division rates resulted in a low overall unsaturation level. The effects of decreasing nitrate concentration in the range 1.0 mM N03 - to 0.001 mM N03 - had a similar basis to those of illumination in that the changes in the total fatty acid composition were largely governed by the rate of TAO accumulation. At high nitrate concentrations the cellular division rate was relatively high and the proportion of TAO in the total lipid extract was low. Consequently, both total and individual lipid classes contained high proportions of unsaturates, particularly 20:5(n-3). However, when the nitrate concentration was decreased, such that it began to limit the rate of cellular division, TAG accumulated Cursory analyses of the molecular species of the galactolipid classes, MODO and DODO, and phospholipid class PC are presented. The effects of environmental variables are discussed in tenns of the changes which may occur in the growth phase distribution of the cells in asynchronous culture, along with the concommitant changes in the lipid composition of the cells. The potential linkage of the elongation and desaturation reactions with both MODO and PC is also discussed briefly with reference to future research