Exploring Biosurfactant Production from Halophilic Bacteria, Isolated from Burgas Salterns in Bulgaria

Biosurfactants produced by halophilic bacteria are gaining attention as eco-friendly and biocompatible alternatives to synthetic surfactants due to their high surface activity, stability under extreme conditions, and intrinsic antimicrobial properties. These amphiphilic biomolecules hold great promise for bioremediation, biomedical, and pharmaceutical applications. In this study, moderately halophilic bacteria capable of biosurfactant production were isolated from saline mud collected at the Burgas solar salterns (Bulgaria). The halophilic microbiota was enriched in Bushnell-Haas (BH) medium containing 10% NaCl amended with different carbon sources. Primary screening in BH liquid medium evaluated the isolates' ability to degrade n-hexadecane while at the same time producing biosurfactants. Thirty halophilic bacterial strains were isolated on BH agar plates supplemented with 2% n-hexadecane, 2% olive oil, or 2% glycerol. Four isolates-BS7OL, BS8OL, BS9GL, and BS10HD-with strong emulsifying activity (E₂₄ = 56%) and reduced surface tension in the range of 27.3-45 mN/m were derived after 7 days of batch fermentation. Strain BS10HD was chosen as the most potent biosurfactant producer. Its phylogenetic affiliation was determined by 16S rRNA gene sequence analysis; according to the nucleotide sequence, it was assigned to Halomonas ventosae. The extract material was analysed by thin-layer chromatography (TLC) and Fourier transform infrared spectroscopy (FTIR). Upon spraying the TLC plate with ninhydrin reagent, the appearance of a pink spot indicated the presence of amine functional groups. FTIR analysis showed characteristic peaks for both lipid and peptide functional groups. Based on the observed physicochemical properties and analytical data, it can be suggested that the biosurfactant produced by Halomonas ventosae BS10HD is a lipopeptide compound.