Combined effects of ultra-high hydrostatic pressure and mild heat on the inactivation of Bacillus subtilis

•6 log bacterial inactivation was achieved by UHHP combined with mild heat.•Permeability change in bacteria membrane increased with pressure.•500 MPa/40 °C caused 82% sub-lethal cells, which retained esterase activity.•500 MPa/60 °C damaged esterase activity, leading to death cell.

Effects of combined ultra-high hydrostatic pressure and mild heat (HPMH) treatments on the food-borne microorganism Bacillus subtilis were evaluated in this study. The B. subtilis culture (counted spores less than 2 log [CFU/mL]) was subjected to 100–500 MPa at 40 °C–60 °C for 15 min at pH 7.0. Treatment with HPMH increased membrane permeability by 10%–89% as determined by the uptake of propidium iodide. Changes in membrane lipids, proteins and DNA were detected in regions 3000 to 2800 cm−1 and 1300 to 900 cm−1 by Fourier transform–infrared spectroscopy (FT-IR). The membrane phospholipid molecules changed from a liquid crystalline state to a gel state, with a decrease in membrane fluidity. HPMH decreased the α-helix content（about 8–22%）while increased the random coil content (about 5–19%) of the cellular protein, which resulted in protein denaturation. Flow cytometry results indicated that HPMH treatment at 60 °C caused 63% damage to the esterase activity of the cells more than HPMH treatment at 40 °C. All of these results revealed the mechanism of HPMH, which is essential for the successful application of high hydrostatic pressure in neutral-pH food processing.