A 13CO2 Enrichment Experiment to Study the Synthesis Pathways of Polyunsaturated Fatty Acids of the Haptophyte Tisochrysis lutea

The production of polyunsaturated fatty acids (PUFA) in Tisochrysis lutea was studied using the gradual incorporation of a 13C-enriched isotopic marker, 13CO2, for 24 h during the exponential growth of the algae. The 13C enrichment of eleven fatty acids was followed to understand the synthetic pathways the most likely to form the essential polyunsaturated fatty acids 20:5n-3 (EPA) and 22:6n-3 (DHA) in T. lutea. The fatty acids 16:0, 18:1n-9 + 18:3n-3, 18:2n-6, and 22:5n-6 were the most enriched in 13C. On the contrary, 18:4n-3 and 18:5n-3 were the least enriched in 13C after long chain polyunsaturated fatty acids such as 20:5n-3 or 22:5n-3. The algae appeared to use different routes in parallel to form its polyunsaturated fatty acids. The use of the PKS pathway was hypothesized for polyunsaturated fatty acids with n-6 configuration (such as 22:5n-6) but might also exist for n-3 PUFA (especially 20:5n-3). With regard to the conventional n-3 PUFA pathway, Δ6 desaturation of 18:3n-3 appeared to be the most limiting step for T. lutea, “stopping” at the synthesis of 18:4n-3 and 18:5n-3. These two fatty acids were hypothesized to not undergo any further reaction of elongation and desaturation after being formed and were therefore considered “end-products”. To circumvent this limiting synthetic route, Tisochrysis lutea seemed to have developed an alternative route via Δ8 desaturation to produce longer chain fatty acids such as 20:5n-3 and 22:5n-3. 22:6n-3 presented a lower enrichment and appeared to be produced by a combination of different pathways: the conventional n-3 PUFA pathway by desaturation of 22:5n-3, the alternative route of ω-3 desaturase using 22:5n-6 as precursor, and possibly the PKS pathway. In this study, PKS synthesis looked particularly effective for producing long chain polyunsaturated fatty acids. The rate of enrichment of these compounds hypothetically synthesized by PKS is remarkably fast, making undetectable the 13C incorporation into their precursors. Finally, we identified a protein cluster gathering PKS sequences of proteins that are hypothesized allowing n-3 PUFA synthesis.

Углеводы как органический субстрат для микроводоросли Tisochrysis lutea (Haptophyta) в условиях лабораторной культуры

Исследована возможность применения углеводов для оптимизации процесса культивирования микроводоросли Tisochrysis lutea (Haptophyta). Проанализировано влияние D-галактозы, глюкозы и сахарозы в концентрациях 100 и 200 мг·л−1 на динамику численности популяции T. lutea. Установлено, что добавление всех изученных углеводов стимулировало рост микроводоросли, однако наибольший эффект оказала сахароза в концентрации 200 мг·л−1.

A co-cultivation process of Nannochloropsis oculata and Tisochrysis lutea induces morpho-physiological and biochemical variations potentially useful for biotechnological purposes

The biotechnological potential of microalgae has gained considerable importance in many applied fields: biomass production for food and feed, cosmeceutical and pharmaceutical products, energy and phytoremediation. The driving force that inspires the progress in microalgae production is the need for new cultivation systems to obtain simultaneously the maximum yield, reduction of water and nutrients use, and production of economically interesting molecules, such as pigments, fatty acids and polysaccharides. We aim to test, for the first time, the co-cultivation in saline medium of Tisochrysis lutea (Haptophyta) and Nannochloropsis oculata (Ochrophyta) to obtain valuable compounds, i.e. pigments and lipids characteristic of each species, using a single culture process. Mono-cultures of each strain were used as controls. The two strains showed an increase in the concentration of chlorophylls and carotenoids in co-culture. At the end of the experiment, the fatty acid profile was analysed by gas chromatography–mass spectrometry. The lipids in the co-cultivated cell extracts were mainly attributable to N. oculata, which represented 97% of the total cells (ca. 83% of the total biomass) at the end of the experiment. Nevertheless, the ω-3 characteristic of T. lutea (DHA and SDA, absent in N. oculata) was also detectable. Although the co-cultivation of these two phylogenetically different species of microalgae did not show positive effects on the growth and on the total lipid production, however, this process resulted in a reduction of the production costs and a lower consumption of water and nutrients.

A Comparative In Vitro Evaluation of the Anti-Inflammatory Effects of a Tisochrysis lutea Extract and Fucoxanthin

In this study, we compared the effects of a Tisochrysis lutea (T. lutea) F&M-M36 methanolic extract with those of fucoxanthin (FX) at equivalent concentration, on lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. The T. lutea F&M-M36 methanolic extract contained 4.7 mg of FX and 6.22 mg of gallic acid equivalents of phenols per gram. HPLC analysis revealed the presence of simple phenolic acid derivatives. The T. lutea F&M-M36 extract exhibited a potent and concentration-dependent inhibitory activity against COX-2 dependent PGE2 production compared to FX alone. Compared to LPS, T. lutea F&M-M36 extract and FX reduced the expression of IL-6 and of Arg1 and enhanced that of IL-10 and of HO-1; T. lutea F&M-M36 extract also significantly abated the expression of NLRP3, enhanced mir-223 expression and reduced that of mir-146b, compared to LPS (p < 0.05). These findings indicate that T. lutea F&M-M36 methanolic extract has a peculiar anti-inflammatory activity against COX-2/PGE2 and NLRP3/mir-223 that might be attributable to the known anti-inflammatory effects of simple phenolic compounds found in the extract that may synergize with FX. Our data suggest that T. lutea F&M-M36 may serve as a source of anti-inflammatory compounds to be further evaluated in in vivo models of inflammation.