The flocculation behaviors of B. licheniformis CGMCC 2876 under different culture conditions were studied in this paper. In the non-bioflocculant producing process, the flocculability of B. licheniformis was enhanced with an increase of ionic strength from 0.001 to 0.3 M, and similar results were observed with the decrease of pH from 7 to 4. The interfacial free energy between cells was 38.8 mJ m⁻², suggesting the cells with hydrophilic surfaces repelled each other. In the bioflocculant producing process, extended DLVO theory was adopted to describe the flocculation behavior. The changes in the flocculation behavior of bacterial cells were primarily attributed to the hydrophobic attraction energy interaction between cells with the lowest value of −104.1 mJ m⁻². Combined with scanning electron microscopy (SEM), transmission electron microscopy (TEM) and X-ray photoelectron spectroscopy analysis, the cell auto-flocculation of B. licheniformis was finally attributed to the hydrophobic polysaccharide secreted as a cell capsule, which led to an increased surface hydrophobicity, thus increasing the flocculation potential.