Determination of spore inactivation during thermal and pressure-assisted thermal processing using FT-IR spectroscopy

The efficacy of microbial inactivation techniques is currently tested using time-consuming and labor-intensive plate count methods, which are the principal rate-limiting steps in developing inactivation kinetic parameters for alternative food processing technologies. Fourier transform infrared (FT-IR) spectroscopy combined with multivariate analysis was used to quantify viable spores and identify some biochemical changes in samples treated by autoclaving, pressure-assisted thermal processing (PATP), and thermal processing (TP). Spore suspensions (similar to 10(9) CFU/mL) of Bacillus amyloliquefaciens TMW 2.479 Fad 82, B. amyloliquefaciens TMW 2.482 Fad 11/2, B. sphaericus NZ 14, B. amyloliquefaciens ATCC 49764, and Clostridium tyrobutyricum ATCC 25755 were treated by PATP (121 degrees C and 700 MPa) for 0, 10, 20, and 30 s and by TP (121 degrees C) for 0, 10, 20, and 30 s. The concentrations of spores in treated samples were determined by plating (reference method). Models developed using partial least-squares regression (PLSR) for predicting spore levels in treated samples had correlation coefficients (r) of > 0.99 and standard errors of cross-validation ranging between 10(0.2) and 10(0.5) CFU/mL. Changes in dipicolinic acid (DPA) and secondary structure of proteins were found to occur during inactivation of spores by PATP and TP. FT-IR spectroscopy could rapidly estimate viable bacterial spore levels in PATP- and TP-treated spore suspensions, providing an accurate analytical tool for monitoring the efficacy of sterilization techniques in inactivating spore-forming microorganisms.