Integrated Screening and Comparative Analysis of Biosurfactant‐Producing Alkaliphiles From a Mud Volcano

ABSTRACT Biosurfactants from extremophilic bacteria are promising green alternatives to synthetic surfactants, particularly for industrial and environmental processes that demand stability under harsh physicochemical conditions. In this study, alkaliphilic bacterial strains were isolated from a high‐pH mud volcano using Horikoshi I medium (pH 10–10.5) and evaluated for biosurfactant production in hydrocarbon‐supplemented Bushnell Haas medium. A total of 24 isolates were subjected to multi‐assay screening, including drop collapse, oil spreading, emulsification, and surface tension measurements. Drop collapse and oil spreading assays showed strong correlation with surface tension reduction, validating their reliability under alkaline conditions. Five isolates— Exiguobacterium alkaliphilum , Pseudomonas jilinensis , Pseudomonas sp., Pseudomonas fulva , and Alkalihalophilus pseudofirmus —demonstrated high activity, reducing surface tension to 26–30 mN/m, with the lowest value reaching 26.07 mN/m. To enhance comparability, a multi‐dimensional visualization framework (heatmap, grouped bar plots, radar chart) was applied, enabling objective assessment across multiple performance parameters. This study presents the first systematic screening of biosurfactant‐producing alkaliphilic bacteria from a mud volcano, establishing a reproducible workflow for extremophile evaluation. The results highlight microbial candidates with strong potential as resources for future environmental biotechnology, particularly for bioremediation and microbial enhanced oil recovery under alkaline conditions.