Resumo

Production, characterization and application of alpha-amylase in reducing biofilm of Candida albicans. Geisiany M. de Queiroz¹, Edvânio R. Rodrigues¹, Luan S. Mialick¹, Juliane V. F. da Silva¹, Rodrigo Sorrechia¹, Juliana Cabrini², Ana Cláudia Pavarina², Rosemeire C. L. R. Pietro¹. ¹Department of Drugs and Medicines, School of Pharmaceutical Sciences, UNESP, Araraquara, SP, Brazil. ²Department of Dental Materials and Prosthesis, School of Dentistry, UNESP, Araraquara, SP, Brazil. The alpha-amylases (EC 3.2.1.1) cleave the links of polysaccharides whit 3 or more units of D-glucose. Biofilms are structured microbial communities, connected to a biotic or abiotic surface, encapsulated within a matrix exopolymeric and formed by different species of micro-organisms. Candida albicans is most frequently associated with formation of biofilms causing several manifestations such as oropharyngeal candidiasis and denture stomatitis. The aim of this study was evaluate the effect of alpha-amylase in C. albicans biofilm. Aspergillus oryzae alpha-amylase was obtained by submerged fermentation in bioreactor. The medium used was described by Carlsen and Nielsen, with modifications. The fermentation conditions were: initial pH 6.0, 0.5 vvm aeration rate, 300 rpm agitation rate and temperature at 30 °C for 7 days. The enzyme was recovered by filtration and the protein and enzyme activity was determined. Amylase activity was performed according to Kammoun, using soluble starch as substrate. The enzyme extract was precipitated with ammonium sulfate 80%, lyophilized, characterized by starch gel electrophoresis stained with iodine and SDS-PAGE gel stained with Coomassie Blue. The efficiency in reducing biofilm was observed against C. albicans (ATCC 90028) biofilms of 24 and 48 hours which were placed in contact with the enzyme for 30 minutes. The viability of biofilms was observed by XTT assay. The biochemical characterization confirmed the presence of alpha-amylase with molecular mass about 55 kDa. In biofilms of 24 hours and 48 hours after enzyme contact was observed 72.5% and 88.4% biofilm reduction, respectively. It was possible to conclude that A. oryzae alpha-amylase has great potential in reducing C. albicans biofilm suggesting biotechnological application. Support: CAPES; PADC/FCF-UNESP.