BIODIESEL PRODUCTION POTENTIALS OF MICROALGAE ISOLATED FROM ARID ENVIRONMENTS OF NIGERIA
Biodiesel production potentials of microalgae isolated from arid environments of Nigeria were investigated. Twenty-five strains of microalgae were isolated from Maiduguri, and screened for growth and lipid contents. Each isolate was cultivated in a 600 mL transparent bottle containing 200 mL of sterile BG-11 medium at pH of 7.3. Each bottle was inoculated with one week-old pure culture of the isolate (inoculum ratio = 15%), capped with urethane foam and incubated at room temperature (30±2oC) for 12 days. They were either naturally illuminated (about 4000 lux) or by using 2-feet fluorescent tubes arranged in parallel with maximum light intensity of 5000 lux. The photoperiod was 12 hours per day. The effects of temperature (20~40oC), light intensity (0 ~ 5000 lux), photoperiods, carbon dioxide concentration in the aeration gas (0.03 ~ 10%), pH (6.0 ~ 8.0), salinity (0 ~1.5% NaCl), NaNO3 concentration (0.1~2.0 g/L), and KH2PO4 concentration (0.01 ~ 0.1 g/L) on cell growth and lipid production were investigated under photoautotrophic conditions. The effects of glucose (10 ~ 40 g/L), and glycerol (0.25 ~ 1.0 ml/L) on cell growth and lipid production were investigated under heterotrophic conditions. All the cultures were shaken manually twice every day and samples were taken every two days for analyses. Biomass, chlorophyll and lipid concentrations and fatty acid profiles were determined using standard methods. All the experiments were carried out in triplicates. Data were presented using descriptive statistics. Four strains, which showed relatively high growth rates and high lipid contents, were selected for further studies. Thery were tentatively identified as Oocystis IA1, Chlorella IA7, Chlorococcum KA9 and Botyrococcus YA5. The optimum temperature for biomass production by all the isolates was 30oC while 20oC was the best for lipid accumulation except for Botyrococcus YA5 that was 25oC. Highest biomass concentration was obtained at 1000 lux for Oocystis, 5000 lux for Chlorella, between 3500 and 5000 lux for Chlorococcum and 3500 lux for Botyrococcus. The optimum light intensities for lipid accumulation were 1000 lux for Oocystis, 2000 lux for Botyrococcus and 3500 lux for Chlorella and Chlorococcum. The length of photoperiod for both biomass and lipid accumulation by Oocystis was 14: 10h, whereas for the others it was 18:06h. Growth of Oocystis was proportional to the carbon dioxide level. For lipid content, 1.0% CO2 was the best for Oocystis, Chlorella and Chlorococcum, while for Botyrococcus, it was 5%. The growth of all the isolates were most favoured in medium with initial pH of 7.0 and least at pH 6.0. The optimum pH values for lipid accumulation were 6.0 for Oocystis and Chlorococcum, 7.0 for Botyrococcus and 8.0 for Chlorella. The algae grew best in media without NaCl except Chlorococcum which showed the highest growth in a medium with 0.5% NaCl. Lipid accumulation by Chlorella was optimum between 0.5 and 1.0% NaCl, whereas 0.5% NaCl was the best for Chlorococcum and Botyrococcus. As the nitrate concentrations in the medium increased, biomass concentrations increased and the lipid contents decreased in all the isolates. Increases in K2HPO4 concentrations led to decreases in lipid contents in all the isolates. The four isolates showed dose-dependent heterotrophic growths on glucose and glycerol. Overall, the maximum biomass concentrations obtained for the four isolates under the optimum conditions were 3.02 ± 0.2, 3.90 ± 0.1, 3.59 ± 0.19, and 2.89 ± 0.03 g L-1 cell dry weight for Oocystis, Chlorella, Chlorococcum and Botyrococcus, respectively while their maximum values for chlorophyll were 34.97 ± 4.24, 30.00 ± 1.46, 39.71 ± 2.06, and 32.27 ± 0.7 mg/g cell respectively. Furthermore, the maximum lipid contents and production obtained for Oocystis, Chlorella, Chlorococcum and Botyrococcus were 57 ± 1.15% and 135.28 ± 3.32 mgL-1day-1; 67.23 ± 2.72% and 165.22 ± 3.36 mgL-1day-1; 63 ± 3.32% and 131.76 ± 1.11 mgL-1day-1; 65 2.88% ± and 140.37 ± 2.13mgL-1day-1 respectively. Thus, it can be concluded that Chlorella IA7 was the best candidate for growth and lipid production followed by Botyrococcus YA5 while Chlorococcum KA9 was the best isolate for chlorophyll production. In comparison with heterotrophic cultures, autotrophic cultures gave the highest biomass concentration, lipid content, lipid production, and chlorophyll contents. Six different fatty acids were found in Oocystis IA1 and Chlorococcum KA9 methyl esters whereas; Chlorella IA7 and Botyrococcus YA5 contained five fatty acids each. Fatty acid methyl ester profiles indicated the presence of C16:0, C16:1, C18:0, C18:1, and C18:2. Out of these, oleic acid (C18: 1) was predominant with the relative percentages ranging between 73.3 and 85.6%. Biodiesel properties namely: density (at 15oC), gml-1; viscosity at 40oC; flash point (oC); iodine value (10-4 Pa s); acid value (mg KOHg-1); and saponification value (mg KOHg-1) were within the ASTM standards. The present study suggests that the four isolates, especially Chlorella IA7 andr Botyrococcus YA5 are good candidates for large-scale biodiesel production.