Risk analysis of the thermal sterilization process. Analysis of factors affecting the thermal resistance of microorganisms.

A risk analysis was applied to experimental heat resistance data. This analysis is an approach for processing experimental thermobacteriological data in order to study the variability of D and z values of target microorganisms depending on the deviations range of environmental factors, to determine the critical factors and to specify their critical tolerance. This analysis is based on sets of sensitivity functions applied to a specific case of experimental data related to the thermoresistance of Clostridium sporogenes and Bacillus stearothermophilus spores. The effect of the following factors was analyzed: the type of target microorganism; nature of the heating substrate; pH, temperature; type of acid employed and NaCl concentration. The type of target microorganism to be inactivated, the nature of the substrate (reference or real food) and the heating temperature were identified as critical factors, determining about 90% of the alteration of the microbiological risk. The effect of the type of acid used for the acidification of products and the concentration of NaCl can be assumed to be negligible factors for the purposes of engineering calculations. The critical non-uniformity in temperature during thermobacteriological studies was set as 0.5% and the critical tolerances of pH value and NaCl concentration were 5%. These results are related to a specific case study, for that reason their direct generalization is not correct.

A critical analysis of mathematical procedures for the evaluation and design of in-container thermal processes for foods.

Kinetic data on thermal destruction of spoilage and quality factors coupled with the temperature history of a product during a heat sterilization cycle are the basic information needed for the evaluation and the design of thermal processes through a physical-mathematical approach. A critical review on the available physical-mathematical procedures used for thermal process calculations of in-container processed foods is presented. The origin and the limitations of each method are discussed. The equations associated with each method, for internal product temperature predictions, are explicitly given. The relative performance of selected methods under identical processing conditions is illustrated. Several problems associated with thermal process calculations are discussed.

Influence of pH, benzoic acid, EDTA, and glutathione on the pressure and/or temperature inactivation kinetics of mushroom polyphenoloxidase.

The pressure and/or temperature inactivation of mushroom polyphenoloxidase (PPO) in the absence and presence of EDTA, benzoic acid, and glutathione was studied on a kinetic basis. In addition, the effect of pH was evaluated. The temperature stability of PPO at atmospheric pressure increased with increasing pH up to pH 6.5. The pressure stability of PPO at room temperature (25 degrees C) also increased with increasing pH. EDTA slightly increased the thermal stability of the enzyme but did not alter the pressure stability of PPO. Benzoic acid protected the enzyme toward temperature but caused sensitization toward pressure when used at a concentration of 50 mM. Glutathione produced sensitization to both temperature and pressure. It is suggested that the sensitizing effect of glutathione is due to an interaction with a disulfide bond of the enzyme.

High pressure, thermal, and combined pressure-temperature stabilities of alpha-amylases from Bacillus species.

Three different alpha-amylases from Bacillus subtilis, B. amyloliquefaciens, and B. licheniformis, were mutually compared with respect to thermal stability, pressure stability, and combined pressure-temperature stability. Measurements of residual enzyme activity and residual denaturation enthalpy showed that the alpha-amylase from B. licheniformis has by far the highest thermostability and that the two other alpha-amylases have thermostabilities of the same order of magnitude. FTIR spectroscopy showed that changes in the conformation of the alpha-amylases from B. amyloliquefaciens, B. subtilis, and B. licheniformis due to pressure occurred at about 6.5, 7.5, and 11 kbar, respectively. It seemed that, for the enzymes studied, thermal stability was correlated with pressure stability. As to the resistance under combined heat and high pressure, the alpha-amylase from B. licheniformis was much more stable than the alpha-amylases from B. amyloliquefaciens and B. subtilis, the latter two being about equally stable. It appears that under high pressure and/or temperature, B. licheniformis alpha-amylase is the most resistant among the three enzymes studied.

Potential Bacillus subtilis alpha-amylase-based time-temperature integrators to evaluate pasteurization processes.

Thermal inactivation kinetics of Bacillus subtilis alpha-amylase (BSA) in different environmental conditions was studied by performing isothermal experiments. As a response property, residual enzymic activity and residual heat of enzyme deterioration were chosen. A comparison of processing values determined from the read-out of a system with actual integrated processing values revealed the potentials of these systems as time-temperature integrators to be used in the pasteurization domain (temperatures of 70 to 100 degrees C) for target attributes with z-values ranging from 6 to 12 degrees C.

Recent advances in process assessment and optimisation.

After stating the general principle of food preservation, this paper focuses on currently available methods to evaluate quantitatively the integrated time temperature impact during and/or after a thermal preservation process. In this context, both the physical-mathematical approach and the use of time temperature integrators are briefly reviewed and recent evolutions are indicated. Also new trends with regard to thermal process optimisation are highlighted.

Evaluation of the integrated time-temperature effect in thermal processing of foods.

In this review, current methods used to evaluate the integrated impact of time and temperature upon preserving a food product by a heat treatment are considered. After identifying the basic premise any preservation scheme shall meet, the central role of a feasible description for the heat activation kinetics of microorganisms, their spores, and other quality attributes are stressed. Common concepts to quantify a thermal process are presented. Shortcomings of the prevalent evaluation methods are highlighted and attention is given to the development, restrictions, and possibilities of time-temperature-integrators as "new" evaluation tools to measure the impact of a "classical" in-pack heat treatment and more modern heating techniques such as continuous processing of solid/liquid mixtures on foods.

Theoretical Consideration on the Influence of the z-Value of a Single Component Time/Temperature Integrator on Thermal Process Impact Evaluation.

The allowed difference in z-value between a single component time/temperature integrator (SCTTI) and target attribute to measure the impact of a thermal process with a given accuracy was examined theoretically. For isothermal heating profiles, the issue and the degree of over- or underestimation of the actual process-value can be predicted as a function of z-value difference and reference temperature. The closer the processing temperature approaches reference temperature, the larger the allowed difference in z-value. As target attributes are characterized by a higher z-value, the allowed z-value difference between SCTTI and target attribute increases. For non-isothermal heating profiles, over- or underestimation depends in addition on the temperature history of the product. In order to obtain a safe estimate of the impact value, whatever the shape of the time/temperature profile, a new approach is suggested based on a SCTTI with a z-value below the target z-value in combination with the use of a reference temperature above or equal to the maximum processing temperature.

DSC and protein-based time-temperature integrators: Case study of alpha-amylase stabilized by polyols and/or sugar.

Differential scanning calorimetry (DSC) was used as a tool for rapid assay of the thermostability of two Bacillus sp. alpha-amylases and horseradish peroxidase as a function of the concentration of glycerol, sorbitol, and sucrose. In this screening study, the DSC peak temperature proved to be a good measure of protein thermostability. By means of isothermal heating experiments, the kinetics of heat decay of B. amyloliquefaciens alpha-amylase were studied by following the course of the DSC peak area (heat exchange (DeltaH/wt)) as a function of time. The high stability of this enzyme in the presence of polyolic alcohols or carbohydrates allowed working at temperatures as high as 127 degrees C. The results of this study can have particular relevance with regard to research on and development of protein-based time-temperature integrators (TTls) for evaluating heat pasteurization or sterilization treatments of foods or pharmaceutical products. The use of the DSC peak area (DeltaH.wt) as TTI-response was validated in experiments with a time-variable temperature profile. Finally, it was shown how the results of such non-isothermal experiments can even be used for (re-) estimation of the protein decay kinetic parameters (k, E(A)). (c) 1994 John Wiley & Sons, Inc.

Convenience of immobilized Bacillus licheniformis alpha-amylase as time-temperature-integrator (TTI).

For the immobilization of Bacillus licheniformis alpha-amylase to porous glass beads, the performances of three possible linking agents, glutaric dialdehyde, benzoquinone and s-trichlorotriazine were assessed in respect of the protein yield, the enzymic activity and the thermostability of the immobilized enzyme. These three properties are to be evaluated in view of the possible use of the enzyme preparations as time-temperature-integrators (TTIs) for assessing the severity of heat pasteurization or sterilization processes of food or pharmaceuticals. All three linkers improved the enzyme's resistance to irreversible heat inactivation to a similar extent and in each case biphasic inactivation kinetics were observed, whereas the dissolved B. licheniformis alpha-amylase showed a simple first order decay. The immobilization yield, measured as protein per carrier weight, did not differ markedly for the three linkers, although the enzymic activity of the glutaric dialdehyde-linked enzyme was lower than that of the benzoquinone- and s-trichlorotriazine-linked preparations.

The influence of polyalcohols and carbohydrates on the thermostability of alpha-amylase.

The influence of polyhydric alcohols and carbohydrates on the thermostability, i.e., the heat inactivation kinetics, of Bacillus licheniformis alpha-amylase was studied in the temperature range 96 degrees to 130 degrees C. High concentrations (from 9 to 60 weight percent) of glycerol, sorbitol, mannitol, sucrose, or starch can markedly decrease the inactivation rate constant, k, and in the studied cases, this stabilizing effect grows stronger with increasing additive concentration. Statements about stabilization should, however, be specified carefully with respect to temperature, because E(A) is mostly altered likewise. For dissolved enzyme E(A) was almost always decreased in the presence of polyol or carbohydrate, whereas for immobilized enzyme it was augmented in each studied instance. The inactivation of dissolved enzyme can, in all the studied cases, be adequately described as a first-order process. Immobilized enzyme, however, shows biphasic then first-order inactivation kinetics, depending on the additive concentration and temperature.

Thermostability of soluble and immobilized alpha-amylase from Bacillus licheniformis.

In view of a possible application of the alpha-amylase from Bacillus licheniformis as a time-temperature integrator for evaluation of heat processes,(11) thermal inactivation kinetics of the dissolved and covalently immobilized enzyme were studied in the temperature range 90-108 degrees C. The D-values (95 degrees C) for inactivation of alpha-amylase, dissolved in tris-HCl buffer, ranged from 6 to 157 min, depending on pH, ionic strength, and Ca(2+) and enzyme concentration. The z-value fluctuated between 6.2 and 7.6 degrees C. On immobilization of the alpha-amylase by covalent coupling with glutaraldehyde to porous glass beads, the thermoinactivation kinetics became biphasic under certain circumstances. For immobilized enzyme, the D-values (95 degrees C) ranged between 17 and 620 min, depending largely on certain environmental conditions. The z-value fluctuated between 8.1 and 12.9 degrees C. In each case of biphasic inactivation, the z-value of the stable fraction (with the higher D-values) was lower than the z-value of the labile fraction.

Lipase-catalyzed ester exchange reactions in organic media with controlled humidity.

Immobilized lipase activity is studied in organic solvent systems of controlled water content under the influence of a variety of reaction parameters, such as temperature, relative humidity, substrate concentrations, and type of fatty acid used. Control of the amount of water in the reaction system was found to be a valuable tool for the orientation of the reaction process and for the determination of the final reaction products. The properties of the immobilized lipase were studied using the interesterification of triolein and palmitic acid as the model system.

Biohydrogenation of unsaturated fatty acids by a mixed culture of rumen microorganisms.

The biohydrogenation of C-18 unsaturated fatty acids was examined in a mixed culture of microorganisms prepared by inoculating a proper growth medium with a sample of rumen fluid. Some major factors influencing the hydrogenation capacity have been investigated. The age of the mixed culture, the type of inoculum used, the concentration of substrates as well as the presence of sterile rumen fluid in the growth medium were found to be important factors determining biohydrogenation behavior. It could be shown that the mixed microbial culture, which had been grown for about 24 h on a medium similar to that of Bryant and Robinson, contained sterile rumen fluid (10% v/v), and had been inoculated with a sample of the whole untreated rumen content, had the best biohydrogenation capacity. The culture was able to carry out the complete conversion of linoleic and linolenic acid to stearic acid.

Measurement of the glucose permeation rate across phospholipid bilayers using small unilamellar vesicles. Effect of membrane composition and temperature.

Small unilamellar vesicles were used to measure the permeability of saturated phosphatidylcholine bilayers to glucose. The presented method circumvents most of the common restriction of classical permeability experiments. Increasing the fatty acid chain length of the lipids reduced the permeation rate significantly. Raising the temperature above that of the lipid phase transition drastically increased membrane permeability. Arrhenius plots demonstrated the activation energy to be independent of membrane composition and the phase-state of the lipids. The permeation process is discussed in terms of a constant energy to disrupt all hydrogen bonds between permeant and aqueous solvent prior to penetrating the membrane. The magnitude of the permeability coefficient is partly determined by a unfavourable change in entropy of activation on crossing the water/lipid interface. All results indicate that the penetration of the dehydrated permeant into the hydrophobic barrier is the rate-limiting step in the permeation of glucose.

Sensitivity of aflatoxin b1 to ionizing radiation.

Aflatoxin B1 in a 5-micrograms/ml water solution was sensitive to ionizing radiation. Inactivation was assayed by the Ames microsome mutagenicity test and confirmed by thin-layer chromatography. Destruction of aflatoxin B1 had already begun at 2.5 kilograms (kGy; 1 kGy = 0.1 Mrad), but a dose exceeding 10 kGy was necessary for total destruction.

Radiation induced lipid oxidation in fish.

Oxidative rancidity in herring and redfish was studied as a function of the applied irradiation dose, the storage time and storage temperature and the packaging conditions.--Measurements of the TBA (thiobarbituric acid) value and the peroxide value were used to evaluate the degree of oxidation of lipids, and were related with sensory scores.--Especially for the fatty fish species (herring) irradiation accelerated lipid oxidation and induced oxidative rancidity. Irradiation of vacuum-packed herring fillets and subsequent storage at +2 degrees C seems to be an interesting process. For the experiments conducted on a semi-fatty fish (redfish), oxidative rancidity was never the limiting factor for organoleptic acceptability.