| Congresso Brasileiro de Microbiologia 2023 | Resumo: 460-1 | ||||
Resumo:Fungi possess remarkable capabilities for biotransformation of chemical molecules and production of valuable biomolecules. Recent years have witnessed significant advancements in the research of enzymes involved in these processes, largely driven by the principles of green chemistry. Among these enzymes, laccases and peroxidases are prominently found in filamentous fungi, playing crucial roles in the degradation of lignocellulosic materials and recalcitrant compounds. This study aimed to compare the enzymatic profile of laccase and two peroxidases produced by the endophytic fungal strain GK.A3.5, which was isolated from the native tree Guarea kunthiana A. Juss (Meliaceae) in Paraná. The fungal strain was cultivated using solid fermentation with rice straw in a BOD incubator at 28°C for three different time periods: 24 hours (t1), 7 days
(t2), and 15 days (t3). The enzymatic extracts were obtained by washing the fermented material with phosphate buffer (pH 7.4) and subsequent centrifugation. The resulting extract was freeze-dried. Enzymatic activities were assessed using specific substrates: 2,2'-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) for laccase activity, and malonate buffer enriched MnSO4 50mM for manganese peroxidase (MnP), and Veratryl alcohol 10mM for lignin peroxidase (LiP). The enzymatic assays were performed on 96-well microplates using appropriate buffers. Absorbance readings were taken at specific wavelengths using a FLASHScan®530 microplate reader (Analytic Jena AG©). Enzymatic activity was calculated in units (U) expressed as μmol.min −1.L−1. The
calculated enzymatic activity was determined using the equation U.L−1 = (∆A x Vt x Df x 106) / (ɛ x Vs x d x t), where ɛ represents the molar absorptivity coefficient (mol.L−1), that variable to wavelengths. The Vs value used for laccase was 40 μL, while for the peroxidase assays, it was 20 μL. The results indicated robust enzymatic performance. The laccase assay revealed enzymatic activities of 11.99 U.L−1(t1), 5.63 U.L−1 (t2), and 4.52 U.L−1 (t3). For the MnP assay, the activities were 69.64 U.L−1 (t1), 41.05 U.L−1 (t2), and
19.84 U.L−1 (t3). Notably, the LiP assay exhibited varying patterns, with activities of 53.53 U.L−1 (t1), 32.30 U.L−1 (t2), and 122.80 U.L−1 (t3). The ABTS and manganese peroxidase assays displayed high initial production followed by a significant decrease over time. In contrast, the lignin peroxidase activity demonstrated a cyclic pattern, with
the highest activity observed on the first day (t1) and the optimal activity at 15 days (t3). These findings suggest that the chosen substrate provided favorable conditions for peroxidase production. Moreover, in comparison to existing literature, these results may guide future studies focused on the degradation of ligninolytic materials for second-generation alcohol production. The present study provides valuable insights into the comparative enzymatic profile of laccase and peroxidases produced by the endophytic fungal strain GK.A3.5. The findings highlight the significance of enzymatic potential, and
further investigations may explore the utilization of ligninolytic materials for sustainable second-generation alcohol production, based on the promising enzymatic performance observed in this study. Palavras-chave: ABTS, Lignin peroxidase, Manganese peroxidase, Extracellular enzyme, Kinetic enzymatic Agência de fomento:CAPES | |||||