Isolation and identification of novel and efficient lipolytic fungi from coconut oil contaminated soi

Abstract

Lipases are biocatalysts that possess remarkable abilities to hydrolyze triacylglycerols, breaking them down into glycerol and fatty acids. These enzymes also catalyze reversible reactions, including esterification, transesterification, and interesterification. The applications of lipases have greatly expanded across various industries, notably in food, pharmaceuticals, cosmetics, detergents, textiles, pulp and paper, leather, medical applications, and many others. Among all living organisms that produce lipases, filamentous fungi are excellent extracellular lipase producers, and fungal lipases have the ability to function at extreme temperatures and pH, are stable in organic solvents, chemo selective, and abundant. Many research studies have been conducted in this area; nevertheless, there are many more lipolytic fungal species to be identified and characterized. Thus, the current study was conducted to isolate and identify the novel lipolytic fungi from coconut oil-contaminated soil. The soil samples were collected from four different areas (A: Normal soil sample from mini Sinharaja area, Faculty of Science, University of Peradeniya, B: Compost sample from Gampaha area, C: Coconut oil-contaminated soil sample from oil mill in Gampaha, D: Coconut oil-contaminated soil sample from oil mill in Kandy). The collected soil samples were cultured in the PDA medium, and fungal species were isolated using streak plate and point inoculation techniques. The fungal strains that were found to be positive by phenol red and Tween 20 tests were subjected to para-nitrophenyl palmitate assay to analyze the lipolytic activity of each species. The selected strains were identified using morphological characteristics and molecular methods. Out of the 12 isolated species, 10 species gave positive results for lipolytic activity, and those were identified based on colony morphology and microscopic view (slide culture technique). Based on the results of the para-nitrophenyl palmitate assay, DNA from three species with the highest lipolytic activity were extracted using the phenol chloroform method and heating method, and ITS regions were sequenced. Through BLAST search and phylogenetic analysis, the most efficient fungal species capable of lipolysis were identified as Rhizopus arrahizus (the most efficient), followed by Aspergillus niger (the second most efficient), and Aspergillus nomius (the third most efficient). In order to establish these fungal species as viable sources of lipases for industrial applications, further enzyme characterization and media optimization studies are necessary.