Optimization of laccase from Stenotrophomonas maltophilia E1 by submerge fermentation using coconut husk with its detoxification and biodecolorization ability of synthetic dyes

Abstract

Enzymatic degradation of synthetic dyes holds an immense promise for addressing the environmental concerns associated with the textile and dye industries. This study aimed to isolate bacteria capable of producing laccase enzymes from an anthropogenic environment. Subsequently, viability of utilizing cost-effective agricultural residues as substrates for laccase production was assessed. Response Surface Methodology (RSM) and the One Variable at a Time (OVAT) approach was pursued for the optimization of laccase production, followed by pH and temperature stability, dye degradation and decolorization experiments, toxicological studies on the degraded dye metabolites. In results, laccase-producing bacterial strain was identified as Stenotrophomonas maltophilia strain E1 (S. maltophilia). Among variety of substrates, coconut husk exhibited optimal efficacy. In a statistical optimization study, it was found that S. maltophilia was capable of producing laccase 51.38 IU/mL, i.e., three times higher than the amount of laccase produced by unoptimized medium (16.7 IU/mL), and the enzyme activity was found to be steady at an acidic pH, and a mesophilic temperature range. The laccase obtained from S. maltophilia E1 demonstrated proficient dye decolorization capabilities, achieving a notable 92.1% reduction in Malachite green dye coloration at a concentration of 500 ppm. Gas chromatography-mass spectrometry (GC-MS) analysis of the decolorized derivatives of Malachite green revealed a conversion into a distinct compounds. Moreover, after undergoing laccase treatment, Malachite green exhibited decreased phytotoxic effects on Oryza sativa, pointing to enzymatic detoxification. Collectively, insights gained from the present study will contribute to the development of efficient enzymatic approaches for addressing the environmental pollution caused by synthetic dyes.

Keywords: Stenotrophomonas maltophilia; Biodecolorization; Detoxification; Laccase; Phytotoxicity; Synthetic dyes.

Fig. 1

Screening of isolate E1 form laccase production on A Guaicol B ABTS, and C syringaldazine embedded in agar plates D phylogenetic analysis

Fig. 2

Evaluation of laccase production using agricultural waste residue. Statistical significance; ns > 0.05, *p < 0.05, **p < 0.005, ***p < 0.0005

Fig. 3

Optimization of fermentation parameters by OVAT approach for maximum laccase production. a effect of incubation time, b effect of agitation, c effect of temperature, and d effect of pH on enzyme production. Statistical significance; ns > 0.05, *p < 0.05, **p < 0.005, ***p < 0.0005

Fig. 4

3D surface plots showing the effect of a coconut husk and CuSO₄; b coconut husk and yeast extract; c coconut husk and initial pH; d CuSO₄ and yeast extract; e CuSO₄ and initial pH; f yeast extract and initial pH on laccase production

Fig. 5

Effect of a pH and b temperature on laccase activity and stability. Statistical significance; ns > 0.05, *p < 0.05, **p < 0.005, ***p < 0.0005

Fig. 6

UV–Vis spectrometry analysis of malachite green containing solution before and after treatment with laccase

Fig. 7

The mass spectra of the degradation products of malachite green by laccase. A malachite green (329 m/z), B Michler’s ketone Bis(4-(dimethylamino) phenyl) methanone (267 m/z), C 4-(dimethylamino) benzophenone (223 m/z), D dibenzylmethane (166 m/z), E 4-(dimethylamino) benzaldehyde (145 m/z), F 4-(dimethylamino) phenol (138 m/z)

Fig. 8

A proposed biodegradation pathways inferred from GC–MS analysis of the products generated during the degradation of malachite green by laccase isolated from S. maltophilia E1