Biodegradation potential of bacterial isolates Enterococcus faecalis RBRJ026 and Klebsiella pneumoniae RBRJ024 against benzyl butyl phthalate

Benzyl butyl phthalate (BBP), a high-molecular-weight phthalic acid diester extensively used as a plasticizer, is a ubiquitous contaminant that gained attention due to its endocrine-disrupting properties, toxicity and environmental persistence. In the present study, isolates of gram-positive bacteria Enterococcus faecalis RBRJ026 and gram-negative bacteria Klebsiella pneumoniae RBRJ024 were used to evaluate their BBP degradation potential. Quantitative analysis using ultra high-performance liquid chromatography (UHPLC) revealed that E. faecalis and K. pneumoniae can degrade up to 90% of BBP under various environmental parameters after 48 h of incubation. The optimum temperature and pH of E. faecalis and K. pneumoniae were 33 °C; 5.0 and 35 °C; 7.0, respectively. The optimum inoculum size for degradation by both bacteria was 5% (v/v). It was also studied that both bacteria have survival rates at high salinity conditions. Kinetic experiments revealed the degradation of BBP after 72 h by E. faecalis and K. pneumoniae was best fitted with a first-order kinetic model utilizing initial concentrations ranging from 100 mg L⁻¹ to 500 mg L⁻¹. The growth of bacteria was also measured by determining the optical density (OD 600 ) using double beam UV-vis spectrophotometer after every 12 h for 72 h. In BBP degradation process, major metabolites like mono butyl phthalate (MBP), mono benzyl phthalate (MBzP), phthalic acid (PA) and protocatechuic acid (PCA) were identified using liquid chromatography and mass spectrometry (LC-MS). Furthermore, the degradation pathway of BBP was also studied.