Screening of Laccase Producing Fungi Using Agro-Wastes under Different Cultural Conditions

Abstract

This study evaluated the potential of fungal isolates for laccase production. Fungi cultures were screened for laccase production on plate assay using 2’ 2’ – azinobis-(-3-ethyl benzthiazoline -6- suphonate) (ABTS), and by submerged fermentation. Result obtained from the plate assay showed the formation of green halo after 2-4 days of incubation due to oxidation of 2’ 2’ – azinobis-(-3-ethyl benzthiazoline -6- suphonate) (ABTS) which is as a result of lignolytic enzymes production Utilization of selected agro-wastes residues (sawdust, plantain and banana peels) for laccase production was evaluated. Fungal isolates were identified based on their cultural characteristics according to standard mycology methods. Light microscopy was performed on cultures and the fungal isolates were identified by their different morphological and colonial characteristics after which they were confirmed using fungal atlas. Total protein content was also determined using the Bradford method Five out of twelve isolates were positive for oxidation of 2’ 2’ – azinobis-(-3-ethyl benzthiazoline -6- suphonate) (ABTS) which signifies laccase enzyme activity and identified as Geotrichum spp, Cephalosporium spp, Trichoderma spp, Trametes spp and Fusarium sppThe highest enzyme activity was observed using Trichoderma spp at 57.1U/l, Trametes spp 51.99 U/l, Fusarium spp had 29.2 U/l. Geotrichum spp and Cephalosporium spp were least at 28.04 U/l and 9.72 U/l respectively. Trichoderma spp was used for further studies. Effect of carbon sources, inoculum size, pH, total protein and shaker speed on laccase production was evaluated. Sawdust gave the highest yield of laccase enzyme 151.17U /l, followed by plantain peel with 62.49U/l of enzyme, banana peel recorded the least of laccase enzyme 54.94 U /l. Protein content obtained from sawdust medium was 45.8mg/ml, plantain peel medium had 29.2 mg/ml while banana peel was the lowest with 16.8mg/ml all at the sixth day of the fermentation process. Optimum pH for sawdust, banana peel and plantain peel was 5.95, 5.94, and 5.83 respectively. Using shaker incubator (150 rpm) with sawdust as carbon source, laccase yield of (310.45U/l) was obtained at pH 6, temperature 25°C and inoculum size of 105 spores per ml. Thus, sawdust can be categorically stated to be safe, cheap and could be suggested for prospective application of higher production of laccase enzyme in various industries.