Impact of acid and alkaline hydrolyses used in gelatin production on the inactivation variability of aerobic and anaerobic bacterial spores

The present study aimed to assess the variability in the inactivation of aerobic and anaerobic spore-forming bacteria during acid and alkaline hydrolyses commonly used in gelatin production. For this, 12 strains previously identified as highly resistant to gelatin processing were used: three Bacillus cereus sensu stricto, three aerobic mesophiles, three aerobic thermophiles, and three anaerobic mesophiles. Porcine skin and bovine tissue samples were inoculated with spores to achieve 6.0 log₁₀CFU/g. Subsequently, porcine skin samples were treated with approximately 0.41 M sulfuric acid (H₂SO₄) for 24 to 48 h (pH < 1), and bovine tissue samples were treated with 0.25 M sodium hydroxide (NaOH) for 7 or 10 days (pH > 10) to simulate chemical hydrolysis used in gelatin production. Chemical inactivation data were fitted using the first-order, Weibull, and biphasic models, with model fitting performance evaluated by the mean square error, F-test, and lack-of-fit test. The reparameterized Weibull model provided the best overall fit of the acid and alkaline spore inactivation data, outperforming first-order and biphasic models. However, for some strains, the fit was not statistically acceptable. Acid hydrolysis for 24 h resulted in a log₁₀-reduction of spores ranging from 0.6 to 2.0, increasing to 1.0 and 3.8 after 48 h. Alkaline hydrolysis showed a log₁₀-reduction from 0.9 to 3.1 after 7 days and 1.6 to 3.7 after 10 days. Variability analysis of log₁₀-reductions was performed to quantify experimental, biological, and strain sources of variation. The variability in log₁₀-reduction among the strains was much higher than the variability among spore batches of the same strain, while experimental variability among technical replicates was low. In conclusion, these findings provide insight into the impact of chemical hydrolysis applied in gelatin production on inactivation variability of spores. These findings serve the gelatin industry to customize and optimize acid and alkaline hydrolysis, also considering the need to achieve a specified microbial reduction of spores to meet microbial specifications set by regulatory authorities and ensure product safety and quality.