Isolation of functional ligninolytic Bacillus aryabhattai from paper mill sludge and its lignin degradation potential

Abstract

Kraft lignin (KL), is the major pollutant in pulp and paper effluent and due to its heterogeneous structure, it is resistant to the depolymerization process. It has drawn much attention from the researcher due to its challenging degradation process. In this study, a KL-degrading bacterium was isolated and screened from paper mill sludge. This bacterium was identified as ligninolytic Bacillus aryabhattai using biochemical and 16SrRNA gene analysis. B. aryabhattai showed maximum activities of lignin peroxidase-LiP (0.74 IU mL^-1) and manganese peroxidase-MnP (9.2 IU mL^-1) on the 4th day, and 5th day, respectively. A total 84% of KL (500 mg L ^-1) reduction was observed after 14 days. The KL bio-degradation was confirmed based on changes in chemical stracture of KL and new metabolites identification using FTIR and GC-MS, respectively. The study concluded that B. aryabhattai maybe becomes a potential biological agent in KL biodegradation and treatment of other lignin-containing industrial effluents.

Keywords: Bacillus aryabhattai; Biodegradation; FTIR; GC–MS; Kraft lignin; SEM.

Fig. 1

A screening test for selecting KL-utilizing bacteria. The preliminarily selected bacteria were grown in culture media (MSM, MSM+KL, and MSM+KL+Glu) for 2 days at 32 °C under shaking (120 rpm) and the OD of bacterial cells was measured at 600 nm. MSM = mineral salt medium, KL = kraft lignin (500 mg L⁻¹), Glu = glucose (0.5%). SL = Serratia liquefaciens, KR = Kocuria rosea, and PP = Pseudomonas putida were included as standard ligninolytic bacteria.

Fig. 2

Quantification of ligninolytic enzymes from B. aryabhattai at different time intervals. Azure B and phenol red were used as indicator dyes for LiP and MnP activity, respectively at the final concentration of 0.01%. (a) LiP activity and (b) MnP activity.

Fig. 3

Growth and KL degradation by B. aryabhattai at a different time interval at 32 °C under shaking (120 rpm) in MSM containing 500 mg L⁻¹ KL.

Fig. 4

SEM images of KL before (a) and after degradation (b) by B. aryabhattai and corresponding EDX (c and d).

Fig. 5

FTIR spectra of KL before (a) and after (b) degradation by B. aryabhattai (14 days).

Fig. 6

GC–MS chromatogram of ethyl acetate extracts of control KL (a) and KL degraded with B. aryabhattai after 14 days. The MS-identified compounds with respect to their retention are listed in Table 1.