Diethyl phthalate degradation by the freeze-dried, entrapped Bacillus subtilis strain 3C3

• Cell size alteration is one of the adaptive mechanisms to diethyl phthalate stress.
• Freeze-dried cell bead formula was developed with high efficiency and viability.
• Sucrose incorporation was beneficial as cryoprotectant and bead reinforcing filler.
• DEP treatment by the formulated bead was demonstrated in a packed bed system.

Microbial degradation is the key treatment for diethyl phthalate (DEP) of which the efficacy is subdued by substrate toxicity. DEP-degrading Bacillus subtilis strain 3C3 adopted cell size alteration as one of the adaptive mechanisms in response to DEP stress at high concentrations. Nevertheless, to enhance cell tolerance in the protected environment and to facilitate practical treatment operation, cell entrapment was optimized with the entrapment yield at 89 ± 1% in a modified minimal salt medium-containing alginate matrix and the freeze-dried, entrapped cells were then formulated. Among several compounds tested, incorporation of sucrose proved to be beneficial as a cryoprotectant sustaining cell biodegradation efficiency (97%) and viability (≥90%) during freeze drying, storage under a vacuum condition at low temperatures, rehydration and as an additional matrix filler to reinforce the bead structure. The effective DEP treatment of the formulated, entrapped cells was demonstrated in a packed bed continuous system in which 70% DEP removal at hydraulic retention time (HRT) of 30 min was occurred and was enhanced up to 90% when HRT was increased to 60 min. The work demonstrates an effective preparation and a potential application of the formulated entrapped DEP-degrading cells for DEP treatment.