Bacterial spores, among the most resistant organisms on earth, can withstand harsh treatments such as desiccation, chemicals, heat, UV, and γ-radiation. Traditional thermal sterilization methods subject food to temperatures of 120⁰-130⁰C for several minutes, degrading its texture, color, flavor, and nutritional properties. Consequently, there is a push for less severe methods, though nonthermal approaches are ineffective against spore-forming bacteria. Ohmic heating (OH), which passes electricity through food, shows promise by significantly enhancing spore inactivation compared to conventional heating (CH) at the same temperature. However, the precise mechanisms behind OH's effectiveness and the influence of different OH parameters (field strength and frequency) on spore killing are not fully understood.
To investigate this, genetically modified spores of Bacillus subtilis, lacking crucial components such as small acid-soluble proteins (SASP) and essential inner membrane proteins, were used. By comparing the inactivation profiles of these mutants with wild-type spores, the study aimed to identify the components affected by the electrical aspects of OH. Additionally, the effects of varying electric field strength and frequency on spore inactivation were explored. Understanding these mechanisms could lead to designing a less severe inactivation process, preserving food quality.
For accurate comparison between OH and CH, the experimental setup allowed matching temperature histories, eliminating spatial temperature gradients. This setup featured small capillaries as sample holders within a T-shaped OH chamber, ensuring precise heating rates. The process differed from traditional methods by allowing temperature to rise linearly with a constant electric field, followed by immediate cooling, enabling the study of field strength effects without holding time.
Clostridium sporogenes, a surrogate for neurotoxin-producing C. botulinum, was tested first. Results indicated that higher field strengths at specific temperatures significantly increased spore inactivation rates. For example, at 130 ºC, spore inactivation nearly doubled when field strength increased from 30 to 50 V/cm. Similarly, Bacillus subtilis PS533 and its mutants (deficient in SASP, recA, DPA, and IM proteins) showed higher killing rates with increased voltage. CH consistently resulted in less spore reduction compared to OH at the same temperatures. For B. subtilis, no direct relationship between frequency and spore inactivation was observed, with 1kHz yielding the maximum reduction. For C. sporogenes, the maximum killing was achieved at lower frequencies, and the effectiveness decreased as the frequency increased.
The inactivation profiles for SASP-less and DPA-less spores were similar under OH and CH, suggesting these molecules' interaction with the electric field leads to higher spore killing. Results with various inner membrane protein-deficient strains indicated interactions of the electric field with these proteins as well. Therefore, OH accelerates spore killing by increasing inner membrane permeability, dissociating the SASP-DNA complex, and interacting with other core molecules.
These findings enhance the understanding of nonthermal electric field effects and highlight OH's potential as a more efficient spore elimination method without significantly impacting product quality. Utilizing high field strength for spore inactivation offers dual benefits of effective killing in shorter durations, reducing processing time, enhancing food quality, and improving processing plant efficiency, leading to energy, time, and labor savings in the food processing sector.