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In this study, we attempted to understand signaling pathways behind lipid biosynthesis by employing a chemical genetics approach based on small molecule inhibitors. Specific signaling inhibitors of MAP kinase or modulators of cAMP signaling were selected to evaluate the functional roles of each of the key signaling pathways in three different microalgal species: Chlamydomonas reinhardtii, Chlorella vulgaris, and Haematococcus pluvialis. Our results clearly indicate that cAMP signaling pathways are indeed positively associated with microalgal lipid biosynthesis. In contrast, MAP kinase pathways in three microalgal species are all negatively implicated in both lipid and carotenoid biosynthesis.

Cultivation is the most expensive step in the production of biodiesel from microalgae, and substantial research has been devoted to developing more cost-effective cultivation methods. Plant hormones (phytohormones) are chemical messengers that regulate various aspects of growth and development and are typically active at very low concentrations. In this study, we investigated the effect of different phytohormones on microalgal growth and biodiesel production in Chlamydomonas reinhardtii and their potential to lower the overall cost of commercial biofuel production. The results indicated that all five of the tested phytohormones (indole-3-acetic acid, gibberellic acid, kinetin, 1-triacontanol, and abscisic acid) promoted microalgal growth. In particular, hormone treatment increased biomass production by 54 to 69 % relative to the control growth medium (Tris-acetate-phosphate, TAP). Phytohormone treatments also affected microalgal cell morphology but had no effect on the yields of fatty acid methyl esters (FAMEs) as a percent of biomass. We also tested the effect of these phytohormones on microalgal growth in nitrogen-limited media by supplementation in the early stationary phase. Maximum cell densities after addition of phytohormones were higher than in TAP medium, even when the nitrogen source was reduced to 40 % of that in TAP medium. Taken together, our results indicate that phytohormones significantly increased microalgal growth, particularly in nitrogen-limited media, and have potential for use in the development of efficient microalgal cultivation for biofuel production.

In this study, proteomic approach was employed to advance our understanding of genes associated with microalgal lipid production. We first generated random mutants of Chlamydomonas reinhardtii, which displayed increased or decreased lipid content, compared to that in the wild-type. Mutants were designated as CR12 and CR48. Then, the changes of protein expression associated with the mutations of Chlamydomonas reinhardtii were analyzed. We could identify proteins that were significantly up-regulated or down-regulated either in the wild-type or CR12 or CR48. Our results will help understand additional genes or pathways directly or indirectly linked to microalgal lipid production., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

A comparative study of Chlamydomonas reinhardtii wild type CC124 and a cell wall-less mutant sta6-1 is described using FACS in conjunction with two different lipophilic fluorescent dyes, Nile Red and BODIPY 505/515. The results indicate that BODIPY 505/515 is more effective for the vital staining of intracellular lipid bodies and single cell sorting than Nile Red. While BODIPY 505/515 stained cells continued to grow after single cell sorting using FACS, Nile Red stained cells failed to recover from sorting. In addition, a comprehensive study was performed to establish a quantitative baseline for future studies for either lipid accumulation and/or microalgal growth by measuring various parameters such as cell count, size, fatty acid contents/composition, and optical/confocal images of the wild type and mutant., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

High-cost downstream process is a major bottleneck for producing microalgal biodiesel at reasonable price. Conventional lipid extraction process necessitates biomass drying process, which requires substantial amount of energy. In this regard, lipid extraction from wet biomass must be an attractive solution. However, it is almost impossible to recover lipid directly from wet microalgae with current technology. In this study, we conceived osmotic shock treatment as a novel method to extract lipid efficiently. Osmotic shock treatment was applied directly to wet Chlamydomonas reinhardtii biomass with water content >99%, along with both polar and non-polar organic solvents. Our results demonstrated that osmotic shock could increase lipid recovery approximately 2 times. We also investigated whether the presence of cell wall or different cell stages could have any impact on lipid recovery. Cell wall-less mutant stains and senescent cell phase could display significantly increased lipid recovery. Taken together, our results suggested that osmotic shock is a promising technique for wet lipid extraction from microalgal biomass and successfully determined that specific manipulation of biomass in certain cell phase could enhance lipid recovery further., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Abstract: We measured the growth of and lipid production by the model microalga Chlamydomonas reinhardtii under different phototrophic, heterotrophic, and mixotrophic conditions to determine the optimal conditions for growth and biodiesel production. In particular, we examined cell growth and yield of fatty acid methyl esters (FAMEs) when C. reinhardtii was cultured in the presence of different organic carbon sources (acetate, glucose, glycerol, and sucrose). C. reinhardtii grew under various conditions, but mixotrophic cultivation was best. The greatest biomass production (2.15gL−1 in 5days) and FAME yield (16.41% of biomass) were observed under mixotrophic cultivation with acetate (10gL−1). As an alternative to acetate, we additionally tested the use of volatile fatty acids (VFAs; acetic, propionic, and butyric acids), which can be inexpensively produced through fermentation of food waste. The highest FAME yield (19.02% of biomass) and biomass production (2.05gL−1 in 5days) were obtained with 5gL−1 of VFAs. This result indicates that VFAs can serve as an inexpensive alternative carbon source for maximizing lipid production in mixotrophic cultivation of C. reinhardtii. [Copyright &y& Elsevier]