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.