Avaliação da produção de biofloculante por microrganismo isolado da kombucha

Abstract

Flocculation is a process that destabilizes suspended particles in solutions and is widely employed in industry for clarification purposes. This process uses flocculants, which may be inorganic, synthetic organic, or natural. Inorganic and synthetic flocculants are effective and low-cost; however, they generate waste, require high dosages, and may pose risks to human health and the environment. As a result, research has increasingly focused on natural alternatives, such as flocculants produced by microorganisms. A promising source for the bioprospecting of such microorganisms is Kombucha, which contains a diverse array of bacteria and yeasts. In this study, microorganisms were isolated from Kombucha and assessed for their ability to produce bioflocculants using a kaolin suspension for flocculating activity tests. Following the selection of the most promising producers, the isolates were genetically identified through sequencing. Inoculum volume testing was then conducted, followed by an experimental design to optimize cultivation conditions by evaluating pH, temperature, agitation, and production time. Additionally, ideal carbon and nitrogen sources for the bioprocess were investigated. The bioflocculant obtained under optimal cultivation conditions was purified and characterized in terms of total sugar and protein content. Flocculating activity tests were conducted to optimize bioflocculant concentration, cation type, and the pH of the kaolin solution. All experiments were performed in triplicate, and data analysis was conducted using RStudio. Initial results led to the selection of five bacterial and three yeast isolates, among which one bacterium and one yeast exhibited superior flocculating activity. The optimal inoculum volume was 3% (v/v) for the bacterium and 2% (v/v) for the yeast. Since the bacterium was identified as pathogenic, likely due to contamination of the laboratory-cultivated Kombucha, only the results of the yeast isolate were considered in the experimental design. The optimal conditions for bioproduct yield were pH 8.0, temperature of 40°C, agitation at 60 rpm for 96 hours, using sucrose and urea as the best carbon and nitrogen sources, respectively. Purification and characterization results indicated low efficiency, as the total protein content (1.9%) was lower than reported in the literature, and flocculating activity decreased from 77.4% to 54.3% following purification. Due to this, both the purified bioflocculant and the untreated form (supernatant of the culture medium) were used in optimization tests to assess differences in flocculating activity. The best dosage of purified bioflocculant was 0.4 mg/mL. Regarding cation type, no significant difference was observed among barium, calcium, and aluminum chlorides, with the highest activity at pH 12.0 (75.6%). Similarly, for the untreated bioflocculant, the best condition was achieved with calcium chloride and pH 12.0 (86.9%). The results demonstrate that the yeast Cutaneotrichosporon debeurmannianum shows potential for bioflocculant production, achieving significant activity under alkaline conditions. However, the purification and characterization processes must be improved to ensure greater efficiency. Future studies are recommended to compare this bioflocculant with other natural flocculants and to evaluate its application in solutions beyond kaolin, aiming toward commercial viability.