Utilization Of Waste Resources For Low-Cost Algae-Based Biofuel Production And Wastewater Bioremediation

University of Minnesota Ph.D. dissertation.June 2016. Major: Bioproducts/Biosystems Science Engineering and Management. Advisor: Roger Ruan. 1 computer file (PDF); xii, 130 pages.Microalgae have great potential to replace current crop feedstocks for biofuel production. However, the algal biofuel industry is still far from being economically available. This dissertation was inspired by the idea of coupling algae-based biofuel generation and municipal wastewater treatment, and developing a biofuel production system that can recycle its own wastes, and thus significantly improve its environmental friendliness and economic viability. The objectives of this study are to fully utilize wastewater and by-products from biodiesel production process to grow mixotrophic algae strains for simultaneous biomass accumulation, biodiesel production and waste stream nutrient removal. The previous study has shown that Chlorella vulgaris grew well on centrate wastewater, which is the best among all municipal wastewater streams for both algae yield and wastewater nutrient removal. For the large scale algal biomass production, wastewater-borne bacteria are expected to influence the algal biomass accumulation. Therefore, different levels of initial algal inoculums were tested to determine the appropriate algae inoculation levels that would allow algae to compete favorably with the wastewater-borne bacteria and dominate the culture. The effect of algae and wastewater-borne bacteria interaction on the algal biomass accumulation and wastewater nutrients removal were studied. Raw and autoclaved centrate was used as the media for algae growth. The results showed that algae can promote bacterial growth, and the presence of bacteria had a significant influence on algal growth pattern, suggesting symbiotic relationship between algae and bacteria at the initial stage of algae cultivation. Moreover, bacteria could increase the algal growth rate and nutrients removal rate at the initial stage of the cultivation. The maximum algal biomass of 2.01 g/L with 0.1 g/L initial algal inoculums concentration can be obtained during algae cultivation in raw centrate medium. The bacteria community profiles tested during cultivation period showed the abundant and diverse microbial community in the samples. Ten phyla and 37 genera were identified. Phylum Bacteroidetes dominate the culture in the centrate wastewater while genus Prevotella was most abundant accounting for 87.1% of the total sequences. At the end of cultivation, microbial community was dominated by phylum Proteobacteria, Bacteroidetes, and Firmicutes. Four bacteria genus, Acinetobacter, Bacteroidales_norank, Megasphaera, and S24-7_norank, may have a strong influence on algal growth. Lipid-extracted microalgal biomass residues (LMBRs) are the leftover biomass from algae-based biodiesel production. The enzymatic hydrolysates of this waste resource, which contain mainly proteins and carbohydrates, were recycled and used as nutritional sources for microalgal cultivation and lipid production. Effect of temperature and substrate concentration on algae growth and lipid production using hydrolyzed LMBRs were studied. The results showed that C. vulgaris could grow mixotrophically in a wide range of temperatures (20∼35 °C). The optimal temperature for cell growth and lipid accumulation of the mixotrophic cultivation of C. vulgaris was between 25 and 30 °C. The neutral lipids of the algal culture at 25 °C accounted for as much as 82 % of the total lipid content in the microalga at culture Day 8. Fatty acid composition analysis showed that the increase of saturated fatty acids was proportional to the increase in cultivation temperature. The maximum biomass concentration of 4.83 g/L and the maximum lipid productivity of 164 mg/L/day were obtained at an initial total LMBRs hydrolyzed sugar concentration of 10 g/L and an initial total concentration of LMBRs hydrolyzed amino acids of 1.0 g/L but decreased at lower and higher substrate concentrations. Waste glycerol generated from biodiesel production using wastewater scum as oil feedstock was recycled and added into wastewater to provide carbon source, support microalgal lipid production, and nutrient removal of wastewater, and further increased the economic viability of scum-based biodiesel production. Effect of crude glycerol and pretreated glycerol concentration and initial pH on algae growth and lipid production of C. vulgaris were tested. The results showed that nutrient removal was improved and lipid production of C. vulgaris was enhanced with the addition of waste glycerol into the wastewater to improve its C/N ratio. The optimal concentration of pretreated glycerol for C. vulgaris cultivation was 10 g L-1 to achieve the biomass concentration of 2.92 g L-1 and lipid productivity of 163 mg L-1 d-1, and the removal of 100% ammonia and 95% of total nitrogen. Alkaline conditions promoted cell growth and lipid accumulation of C. vulgaris and simultaneously stimulated nutrient removal