Isolation and Characterization of Newly Laccase-Producing Endophytic Fungi in Submerged Cultures from Calotropis gigantean Plant Leaves

Abstract

The aim of the study to isolate and identify new laccase sources from an endophytic fungal source that could be used as a weapon for
eliminating and detoxifying contaminants found in wastewater and aquatic habitats. Isolation was done from Calotropis gigantean plant
leaves from different locations of paper mill effluents from the “Raipur” region of Chhattisgarh, India. Positive isolates were obtained
with a dark brown color below and surrounding the fungal colony due to guaiacol oxidation on potato dextrose agar. One potent
endophytic fungal isolate that produces laccase is identified as Aspergillus turcosus by using the internal transcribed spacer (ITS) and
BLAST analysis. After optimization, maximum laccase production was obtained at the following conditions: medium (Czapeck Dox Broth),
carbon source (sucrose), nitrogen source (sodium nitrate), pH (6), activator tannic acid (20 mM), incubation period (35°C) and duration
(8 days) with 3 (8 mM fermentation) disc inoculums. The maximum laccase activity was obtained at 65 U mL-1 in submerged optimized
conditions, which was more than two fold compared to the unoptimized conditions. As estimated by SDS-PAGE, The molecular mass
of the monomer of pure laccase was determined to be 66 kDa. After five days of treatment with the laccase of A. turcosus, the synthetic
dyes phenol red, bromophenol blue, methyl orange, and Congo red lose their colour. Clearance rates for chemical oxygen demand were
59.46 and 48.57%, and phenolic contaminants were 80 and 22.3% in coal and textile effluents during the required treatment periods,
respectively. One novel and potent laccase-producing endophytic fungus was successfully isolated, which can be utilized as a laccaseproducing
source for various industrial applications.