Complementary Analytical Platforms of NMR Spectroscopy and LCMS Analysis in the Metabolite Profiling of Isochrysis galbana.

This study was designed to profile the metabolites of Isochrysis galbana, an indigenous and less explored microalgae species. 1H Nuclear Magnetic Resonance (NMR) spectroscopy and Liquid Chromatography-Mass Spectrometry (LCMS) were used to establish the metabolite profiles of five different extracts of this microalga, which are hexane (Hex), ethyl acetate (EtOAc), absolute ethanol (EtOH), EtOH:water 1:1 (AqE), and 100% water (Aq). Partial least square discriminant analysis (PLS-DA) of the generated profiles revealed that EtOAc and Aq extracts contain a diverse range of metabolites as compared to the other extracts with a total of twenty-one metabolites, comprising carotenoids, polyunsaturated fatty acids, and amino acids, that were putatively identified from the NMR spectra. Meanwhile, thirty-two metabolites were successfully annotated from the LCMS/MS data, ten of which (palmitic acid, oleic acid, α-linolenic acid, arachidic acid, cholesterol, DHA, DPA, fucoxanthin, astaxanthin, and pheophytin) were similar to those present in the NMR profile. Another eleven glycerophospholipids were discovered using MS/MS-based molecular network (MN) platform. The results of this study, besides providing a better understanding of I.galbana's chemical make-up, will be of importance in exploring this species potential as a feed ingredient in the aquaculture industry.

De novo transcriptome analysis of Chlorella sorokiniana: effect of glucose assimilation, and moderate light intensity.

Chlorella can produce an unusually wide range of metabolites under various nutrient availability, carbon source, and light availability. Glucose, an essential molecule for the growth of microorganisms, also contributes significantly to the metabolism of various metabolic compounds produced by Chlorella. In addition, manipulation of light intensity also induces the formation of secondary metabolites such as pigments, and carotenoids in Chlorella. This study will focus on the effect of glucose addition, and moderate light on the regulation of carotenoid, lipid, starch, and other key metabolic pathways in Chlorella sorokiniana. To gain knowledge about this, we performed transcriptome profiling on C. sorokiniana strain NIES-2168 in response to moderate light stress supplemented with glucose under mixotrophic conditions. A total of 60,982,352 raw paired-end (PE) reads 100 bp in length was obtained from both normal, and mixotrophic samples of C. sorokiniana. After pre-processing, 93.63% high-quality PE reads were obtained, and 18,310 predicted full-length transcripts were assembled. Differential gene expression showed that a total of 937, and 1124 genes were upregulated, and downregulated in mixotrophic samples, respectively. Transcriptome analysis revealed that the mixotrophic condition caused upregulation of genes involved in carotenoids production (specifically lutein biosynthesis), fatty acid biosynthesis, TAG accumulation, and the majority of the carbon fixation pathways. Conversely, starch biosynthesis, sucrose biosynthesis, and isoprenoid biosynthesis were downregulated. Novel insights into the pathways that link the enhanced production of valuable metabolites (such as carotenoids in C. sorokiniana) grown under mixotrophic conditions is presented.

Identification and Analysis of microRNAs in Chlorella sorokiniana Using High-Throughput Sequencing.

Chlorella is a popular microalga with robust physiological and biochemical characteristics, which can be cultured under various conditions. The exploration of the small RNA content of Chlorella could improve strategies for the enhancement of metabolite production from this microalga. In this study, stress was introduced to the Chlorella sorokiniana culture to produce high-value metabolites such as carotenoids and phenolic content. The small RNA transcriptome of C. sorokiniana was sequenced, focusing on microRNA (miRNA) content. From the analysis, 98 miRNAs were identified in cultures subjected to normal and stress conditions. The functional analysis result showed that the miRNA targets found were most often involved in the biosynthesis of secondary metabolites, followed by protein metabolism, cell cycle, and porphyrin and chlorophyll metabolism. Furthermore, the biosynthesis of secondary metabolites such as carotenoids, terpenoids, and lipids was found mostly in stress conditions. These results may help to improve our understanding of regulatory mechanisms of miRNA in the biological and metabolic process of Chlorella species. It is important and timely to determine the true potential of this microalga species and to support the potential for genetic engineering of microalgae as they receive increasing focus for their development as an alternative source of biofuel, food, and health supplements.

Abnormal spermatogenesis in mice unable to synthesize ascorbic acid.

Although exposure to environmental toxicants, including endocrine-disrupting chemicals, is thought to be a possible cause of male infertility, the pathogenesis of male reproductive disorders remains unclear. In the present study, we used Gulo-/- mutant mice, which are unable to synthesize ascorbic acid, to study the importance of dietary vitamin C (VC) on spermatogenesis. Regular chow containing approximately 110 mg/kg VC is unable to support the growth of these mutant mice, but a VC supplement in their drinking water (330 mg/L) is able to ameliorate the VC deficiency. Testes of Gulo-/- mutants born from heterozygous mothers without VC supplement (VC-deficient mice) and those born from mothers given a VC supplement (VC-sufficient mice) were examined by morphological and biochemical analyses. Morphological analysis revealed that apoptosis of spermatocytes occurred frequently in VC-deficient mice at 20 days of age. Two-dimensional electrophoresis analysis revealed the specific disappearance of heat-shock protein (Hsp) 70 in the testes of 20-day-old VC-deficient mice. In the present study, the relationship between the apoptosis of spermatocytes and Hsp70 in VC-deficient mice is discussed.