In this paper, surface tension measurement, oil-spreading test and blood-plate hemolysis test were
attempted in the screening of demulsifying bacteria. After the comparison to the screening results
obtained in demulsification test, 50 mN/m of surface tension of culture was proposed as a preliminary
screening standard for potential demulsifying bacteria. For the identification of efficient demulsifying
strains, surface tension level was set at 40 mN/m. The detected strains were further verified in demulsi-
fication test. Compared to using demulsification test alone as screening method, the proposed screening
protocol would be more efficient. From the screening, a highly efficient demulsifying stain, S-XJ-1, was
isolated from petroleum-contaminated soil and identified as Alcaligenes sp. by 16S rRNA gene and phys-
iological test. It achieved 96.5% and 49.8% of emulsion breaking ratio in W/O and O/W kerosene emulsion
within 24 h, respectively, and also showed 95% of water separation ratio in oilfield petroleum emulsion
within 2 h. The bio-demulsifier was found to be cell-wall combined. After soxhlet extraction and purifi-
cation through silicon-gel column, the bio-demulsifier was then identified as lipopeptide biosurfactant by
TLC and FT–IR.