In Vivo Safety Evaluation of Bacteriocin Isolated from Probiotic Lactobacillus

Abstract

The aim of this research was to evaluate the safety of bacteriocin TSU-4 isolated from probiotic lactic acid bacteria Lactobacillus animalis TSU4, isolated from fresh water fish catla catla. The bacteriocin was produced in bulk and purified by a three step method including ammonium sulphate precipitation and chloroform-methanol extraction, followed by cation-exchange chromatography for further purification and finally, reverse-phase HPLC to verify the purity of the bacteriocin. For the safety evaluation study, Immunogenicity and acute and subchronic toxicity of the bacteriocin TSU-4 was evaluated in-vivo in male Swiss-Albino mice. For immunogenicity assay, mice were injected intraperitoneally with the bacteriocin (50 μg/ml) and in combination with freund’s adjuvant. In acute toxicity assay, mice were orally administered with increasing concentration of bacteriocin (50, 100 and 200 mg/kg) sparingly within a period of maximum three hours. For the sub-chronic toxicity evaluation, 0.5 mg/kg/day of the Bacteriocin was orally given to the mice for a period of 21 days. There were no deaths or hypersensitivity reactions observed during the course of acute and sub-chronic toxicity assays. Also, in the antibody titer, there was no significant increase observed. The LD50 value of the bacteriocin was observed to be higher than 200 mg/kg. Serum biochemical tests and Histopathological analysis were also performed for acute and subchronic toxicity groups. No significant change in the serum biochemistry was observed. In the Histopathological studies, acute toxicity group mice treated with 100 mg/kg dose showed mild morphological changes in the kidney and liver but did not lead to disfunctioning of the respective organs. Although, surprisingly there was no damage observed in the mice treated with 200 mg/kg of the bacteriocin. In case of subchronic toxicity group, notable damage and inflammation was observed in both the organs. Hence, bacteriocin TSU-4 can be concluded to cause no remarkable morphological changes in the organs and be a safe and potential candidate for medical as well as industrial application.