ABSTRACT
The thraustochytrid are marine heterotrophic protists that are widely distributed in the marine world. They are characterized by producing and accumulating great amount of lipids in their cells, especially long chain polyunsaturated fatty acids (LC-PUFA), highlighting the docosahexaenoic acid (DHA, 22:6, n-3), eicosapentaenoic acid (EPA, 20:5, n-3) and arachidonic acid (ARA, 20:4, n-6), as well as pigments of interest for human health and animal nutrition, such as carotenoids. Therefore, the objective of this study was to isolate and characterize three natives isolated of thraustochytrids and assess the potential of the by-products of the manufacture of beer (RB) and protein extraction of Lupine flour (RL) as complex carbon sources to produce biomass, lipid and polyunsaturated fatty acids. Three native strains of thraustochytrid (AS5-B2, IQ81 y VAL-B1), isolated from Chilean coastal waters were morphologically and genetically identified as thraustochytrid. For the determination of biomass production cultures were quantified by gravimetry and the fatty acids quantification and identification were carried out by gas chromatography (GC-FID). Our results show that the culture with any sources of complex carbon used, increased significantly the production of both biomass and total lipids in the strains IQ81 and VAL-B1, compared to glucose as pure carbon source. On the other hand, strain AS5-B2 showed a decrease in the total production of lipids in RB compared to the pure carbon source. For the production of fatty acids, the strains IQ81 and VAL-B1 showed a significant increase in DHA when growing in RB. In conclusion strains IQ81 and VAL-B1 can be used to biotransform industrial waste, such as RB and RL, into a more valuable product such as DHA, EPA, ARA and lipids.
ABSTRACT
Liver alcohol dehydrogenase oxidizes ethanol to acetaldehyde, which is further oxidized to acetate by aldehyde dehydrogenase-2 (ALDH2*1). Individuals who carry a low-activity ALDH2 (ALDH2*2) display high blood acetaldehyde levels after ethanol consumption, which leads to dysphoric effects, such as facial flushing, nausea, dizziness, and headache ("Asian alcohol phenotype"), which result in an aversion to alcohol and protection against alcohol abuse and alcoholism. Mimicking this phenotype may reduce alcohol consumption in alcoholics. RNA interference (RNAi) is a cell process in which a short interfering RNA (siRNA) of 21-25 bp guides the degradation of a complementary target mRNA. Thus, siRNAs may be useful in mimicking the Asian phenotype by inhibiting ALDH2 gene expression. We determined the inhibitory effect of three chemically synthesized siRNAs targeted against rat ALDH2 mRNA in human embryonic kidney cells (HEK-293 cell lines) transfected with a plasmid carrying the rat ALDH2 cDNA. Two of the three siRNAs were active, yielding a 65-75% reduction of ALDH2 activity. Based on the most promising siRNA sequence, three short hairpin RNA (shRNA) genes driven by the human U6 RNA promoter were designed and cloned in a plasmid. After transfection of HEK-293 cells, one of the genes was shown to be active, yielding a 50% reduction of ALDH2 activity. This effect is consistent with a 50% reduction in ALDH2 mRNA, whereas neither beta-actin mRNA nor the interferon-inducible transmembrane protein-1 mRNA levels were affected. This study describes chemically synthesized siRNAs and an endogenously synthesized shRNA, which reduce ALDH2 activity and constitute tools that should be of value for further alcohol research.
