Abstract
Alternative energy resources have become an important issue due to the limited stocks of petroleum-based fuel. Microalgae, a source of renewable biodiesel, use solar light to convert CO2 into lipid droplets (LDs). Quantification of LDs in microalgae is required for developing and optimizing algal bioprocess engineering. However, conventional quantification methods are both time and labor-intensive and difficult to apply in high-throughput screening systems. LDs in plant and mammalian cells can be visualized by staining with various fluorescence probes such as the Nile Red, BODIPY, and Seoul-Fluor (SF) series. This report describes the optimization of LD staining in Chlamydomonas reinhardtii with SF probes via systematic variations of dye concentration, staining time, temperature, and pH. A protocol for quantitative measurement of accumulation kinetics of LDs in C. reinhardtii was developed using a spectrofluorimeter and the accuracy of LD size measurement was confirmed by transmission electron microscopy (TEM). Our results indicate that our spectrofluorimeter-based measurement approach can monitor kinetics of intracellular LDs (in control and nitrogen-source-starved Chlamydomonas reinhardtii) accumulation that has not been possible in the case of conventional imaging-based methods. Our results presented here confirmed that an SF44 can be a powerful tool for in situ monitoring and tracking of intracellular LDs formation.
Highlights
IntroductionPetroleum-based fuels have been the main energy source of modern society since the Industrial
Petroleum-based fuels have been the main energy source of modern society since the IndustrialRevolution
This study describes the demonstration of staining conditions using SF44 with Chlamydomonas reinhardtii
Summary
Petroleum-based fuels have been the main energy source of modern society since the Industrial. Numerous governmental and industrial efforts are focused on exploring alternative energy sources that are renewable and carbon neutral [1] One of these alternative energy sources is biodiesel derived from crops including corn, soybean, canola, coconut, and palm oils. These materials cannot entirely replace petroleum as transport fuels due to their relatively low oil yield (L/ha) [1]. Other methods including thin-layer chromatography (TLC), high pressure liquid chromatography (HPLC), and gas chromatography (GC), require dedicated analytical instrumentation [20] These methods are both time- and labor-intensive and difficult to apply in high-throughput screening systems. The optimized protocol is a sensitive and quantitative method for high-throughput screening of intracellular lipid accumulations of microalgae for biodiesel research. LDs and liposome specific fluorescence intensity was monitored at 540 nm with excitation at 450 nm
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