Aspergillus caespitosus ASEF14, an oleaginous fungus as a potential candidate for biodiesel production using sago processing wastewater (SWW)

Background Oleaginous microorganisms are sustainable alternatives for the production of biodiesel. Among them, oleaginous fungi are known for their rapid growth, short life cycles, no light requirement, easy scalability, and the ability to grow in cheap organic resources. Among all the sources used for biodiesel production, industrial wastewater streams have been least explored. We used oleaginous fungi to decontaminate sago processing wastewater and produce biodiesel. Results Among the 15 isolates screened for lipid production and starch utilization using the Nile red staining assay and amylase plate screening, three isolates accumulated > 20% (w/w) of their dry cell mass as lipids. The isolate ASEF14 exhibited the highest lipid accumulation (> 40%) and was identified as Aspergillus caespitosus based on the 28S rRNA gene sequencing. The maximum lipid content of 54.4% in synthetic medium (SM) and 37.2% in sago processing wastewater (SWW) was produced by the strain. The Fourier-transform infrared (FTIR) spectroscopy of the fungal oil revealed the presence of functional peaks corresponding to major lipids. Principal component analysis (PCA) of the FTIR data revealed major changes in the fatty acid composition during the transition from the growth phase (Days 1–3) to the lipid accumulation phase (Days 4–7). The fatty acid methyl esters (FAME) analysis of fungal oil from SWW contained 43.82% and 9.62% of saturated and monounsaturated fatty acids, respectively. The composition and percentage of individual FAME derived from SWW were different from SM, indicating the effect of nutrient and fermentation time. The fuel attributes of the SM- and SWW-grown fungal biodiesel (kinematic viscosity, iodine value, cetane number, cloud and pour point, linolenic acid content, FA > 4 double bonds) met international (ASTM D6751, EN 14214) and national (IS 15607) biodiesel standards. In addition to biodiesel production, the strain removed various contaminants such as total solids (TS), total suspended solids (TSS), total dissolved solids (TDS), dissolved oxygen (DO), chemical oxygen demand (COD), biological oxygen demand (BOD), total nitrogen (TN), total phosphorus (TP), and cyanide up to 58.6%, 53.0%, 35.2%, 94.5%, 89.3%, 91.3%, 74.0%, 47.0%, and 53.84%, respectively, from SWW. Conclusion These findings suggested that A. caespitosus ASEF14 is a potential candidate with high lipid accumulating ability (37.27%), capable of using SWW as the primary growth medium. The medium and incubation time alter the FAME profile of this fungus. The physical properties of fungal oil were in accordance with the biodiesel standards. Moreover, it decontaminated SWW by reducing several polluting nutrients and toxicants. The fungal biodiesel produced by this cost-effective method could serve as an alternate path to meet global energy demand.

Potential Assessment of Oleaginous Fungi for Sustainable Biodiesel Production: Screening, Identification and Lipid Production Optimization

The present work, aiming to exploit oleaginous fungi for biodiesel production. Ten fungal strains were isolated from two petroleum polluted soil samples and screened for their abilities to accumulate lipid. Lipid rich three species viz, Aspergillus terreus, Aspergillus niger and Aspergillus flavus were found to be the highest lipid producers. Potential isolates were identified at the species level by morphological (macroscopic and microscopic) examination and molecularly confirmed by using 18S rRNA gene sequencing. Improvement of lipid accumulation by optimization of various parameters of culture conditions. The results reported clearly that the most suitable medium conditions for highest lipid production (38.33%) of Aspergillus terreus as the most potent lipid producer composed of 5% sucrose, 0.5 g/L ammonium nitrate with initial pH 6.0, after seven days of incubation in a static condition. The three promising fungal isolates have been taken for fatty acids analysis by gas chromatograph (GC) after transesterification. Fatty acid methyl esters (FAME) profile indicated the presence of higher saturated fatty acid fractions compared to polyunsaturated fatty acids. The total concentration of fatty acids was 107.98, 38.29, and 37.48 mg/100g of lipid accumulated by A. terreus, A. niger and A. flavus, respectively. Gas chromatograph analysis of A. terreus lipid indicated that oleic acid (C18:1, 18.51%) was the most abundant fatty acid, followed by stearic acid (C18:0, 15.91%) and Myristic acid (C14:0, 14.64%), respectively. Therefore, fatty acid profile of A. terreus has confirmed its potentiality as feedstock for producing lipid for biodiesel manufacturing.

FAMEs Profile of Oil Produced by Oleaginous Fungi Isolated from Fermented Beverage Wastewaters and Soil

Fungal strains isolated from fermented maize (ogi) (PW) and sorghum-based brewery wastewaters (BW) and two soil isolates were evaluated for oleaginicity. The fungal isolates from the wastewater that had lipid content of at least 20% of their biomass were identified by both culture methods and internal transcribed spacer (ITS) 1-5.8S-ITS2 ribosomal DNA sequencing. The isolates were identified to be Aspergillus fumigatus (PW8), Aspergillus flavus (PW10), Candida tropicalis (PW16) and Aspergillus tubingensis (PW3), Trichosporon luoberi (BW7), Aspergillus sp. (BW4) and Candida tropicalis (BW1; BW3). FAMEs composition was determined for the four strains with the highest lipid content by acid-catalyzed transesterification and analyzed by Gas Chromatography-Flame Ionization Detector (GC-FID). Palmitoleic acid was the dominant fatty acid in M. circinelloides and T. reesei, and the best producers of capric and lauric acids were Aspergillus fumigatus and Aspergillus sp. (BW4), respectively. These fatty acids are beneficial in making cosmetics and pharmaceuticals (antimicrobials and dietary supplements). The analysis of the FAMEs profile in the species indicated low amounts or absence of some key long chain fatty acid (LCFA) constituents of biodiesels. Based on the FAMEs profile of M. circinelloides investigated, this strain could hold promise for use as feedstock for biodiesel with genetic engineering and a tailored lipid production favouring enrichment of LCFA. Keywords: Fungal lipids, wastewater, fatty acid methyl ester, GC-FID