A model of non‐isothermal degradation of nutrients,pigments and enzymes

Published isothermal degradation curves for chlorophyll A and thiamine in the range 100–150 °C and the inactivation curves of polyphenol oxidase (PPO) in the range 50–80 °C could be described by the model C(t)/C₀ = exp[?b(T)tⁿ] where C(t) and C₀ are the momentary and initial concentrations, respectively, b(T) a temperature dependent ‘rate parameter’ and n, a constant. This suggested that the temporal degradation/inactivation events of all three had a Weibull distribution with a practically constant shape factor. The temperature dependence of the ‘rate parameter’ could be described by the log logistic model, b(T) = log_e[1 + exp[k(T ? T_c)], where T_c is a marker of the temperature level where the degradation/inactivation occurs at a significant rate and k the steepness of the b(T) increase once this temperature range has been exceeded. These two models were combined to produce a non‐isothermal degradation/inactivation model, similar to one recently developed for microbial inactivation. It is based on the assumption that the local slope of the non‐isothermal decay curve, ie the momentary decay rate, is the slope of the isothermal curve at the momentary temperature at a time that corresponds to the momentary concentration of the still intact or active molecules. This model, in the form of a differential equation, was solved numerically to produce degradation/inactivation curves under temperature profiles that included heating and cooling and oscillating temperatures. Such simulations can be used to assess the impact of planned commercial heat processes on the stability of compounds of nutritional and quality concerns and the efficacy of methods to inactivate enzymes. Simulated decay curves on which a random noise was superimposed were used to demonstrate that the degradation/inactivation parameters, k and T_c, can be calculated directly from non‐isothermal decay curves, provided that the validity of the Weibullian and log logistic models and the constancy of the shape factor n could be assumed. Copyright © 2004 Society of Chemical Industry     

The impact of high pressure and temperature on bacterial spores: inactivation mechanisms of Bacillus subtilis above 500 MPa

Abstract: High‐pressure thermal sterilization (HPTS) is an emerging technology to produce shelf stable low acid foods. Pressures below 300 MPa can induce spore germination by triggering germination receptors. Pressures above 500 MPa could directly induce a Ca⁺²‐dipicolinic acid (DPA) release, which triggers the cortex‐lytic enzymes (CLEs). It has been argued that the activated CLEs could be inactivated under HPTS conditions. To test this claim, a wild‐type strain and 2 strains of Bacillus subtilis spores lacking germinant receptors and one of 2 CLEs were treated simultaneously from 550 to 700 MPa and 37 to 80 C (slow compression) and at 60 to 80 C up to 1 GPa (fast compression). Besides, an additional heat treatment to determine the amount of germinated cells, we added TbCl₃ to detect the amount of DPA released from the spore core via fluorescent measurement. After pressure treatment for 120 min at 550 MPa and 37 °C, no inactivation was observed for the wild‐type strain. The amount of released DPA correlated to the amount of germinated spores, but always higher compared to the belonging cell count after pressure treatment. The release of DPA and the increase of heat‐sensitive spores confirm that the inactivation mechanism during HPTS passes through the physiological states: (1) dormancy, (2) activation, and (3) inactivation. As the intensity of treatment increased, inactivation of all spore strains also strongly increased (up to ?5.7 log₁₀), and we found only a slight increase in the inactivation of one of the CLE (sleB). Furthermore, above a certain threshold pressure, temperature became the dominant influence on germination rate. Practical Application: The continuous increase of high‐pressure (HP) research over the last several decades has already generated an impressive number of commercially available HP pasteurized products. Furthermore, research helped to provoke the certification of a pressure‐assisted thermal sterilization process by the U.S. FDA in February 2009. However, this promising sterilization technology has not yet been applied in industrial settings. An improved understanding of spore inactivation mechanisms and the ability to calculate desired inactivation levels will help to make this technology available for pilot studies and commercialization at an industrial scale. Moreover, if the synergy between pressure and elevated temperature on the inactivation rate could be identified, clarification of the underlying inactivation mechanism during HP thermal sterilization could help to further optimize the process of this emerging technology.     

Extraordinary Heat Resistance of Talaromyces flavus and Neosartorya fischeri Ascospores in Fruit Products

Ascospores of three strains each of Talaromyces flavus, Neosartorya fischeri and Byssochlamys fulva/nivea were analyzed for resistance to thermal inactivation in five fruit-based (blueberry, cherry, peach, raspberry and strawberry) products. D_91°C values for two strains of T. flavus ranged from 2.9–5.4 min; D_88°C values ranged from 7.1–22.3 min. Ascospores of N. fischeri were somewhat less heat resistant; D_91°C values were < 2.0 min and D_88°C were 4.2–16.2 min. Ascospores of Byssochlamys spp. were considerably less heat resistant. The type of fruit product did not appear to substantially influence rates of thermal inactivation. No heat-resistant ascospores of T. flavus or N. fischeri, i.e., ascospores capable of surviving 15 min at 75°C, were formed on fruit products stored at 10°C for 137 days. However, T. flavus and N. fischeri formed ascospores on cherry substrate stored at 25°C within 65 and 137 days, respectively, that survived 15 min at 88°C.     

Evaluation of the strain variability of Salmonella enterica acid and heat resistance

The inherent acid and heat resistances of 60 Salmonella enterica strains were assessed in tryptone soy broth without dextrose acidified to pH 3.0 or heated at 57 °C. A total of 360 inactivation curves were generated. Regarding the acid challenge experiments, the inactivation rate (k_acid), estimated using the log–linear model, ranged from 0.47 to 3.25 h⁻¹. A log–linear model with a “survival tail” was used to describe the thermal inactivation of the strains, and the estimated inactivation rate (k_heat) ranged from 0.42 to 1.33 min⁻¹. The strain variability of k_acid was considerably higher than that of k_heat with the coefficient of variation of this kinetic parameter among the tested strains being 39.0% and 18.3%, respectively. No correlation was observed between the estimated k_acid and k_heat values of the 60 S. enterica strains. Furthermore, no trends among the tested strains related to origin, serotype or antibiotic resistance profile were evident. The present study is the first one to comparatively evaluate the inherent acid and heat resistance profiles of multiple S. enterica strains. Beyond their value in strain selection for use in food safety challenge studies, the collected data should be useful in describing and integrating the strain variability of S. enterica acid and heat resistance profiles in quantitative microbial risk assessment.     

Thermal inactivation kinetics of quality-related enzymes in cauliflower (<Emphasis Type="Italic">Brassica oleracea</Emphasis> var. <Emphasis Type="Italic">botrytis</Emphasis><Emphasis Type="Bold">)</Emphasis>

Thermal inactivation of quality-related enzymes in both cauliflower crude enzyme extracts and fresh tissue samples was studied in temperature range 50–100 °C. For crude enzyme extracts, several parameters, reaction rate constants (k) and activation energy (E _a) as well as decimal reduction time (D) and (z) values, were used to characterize the thermal stability. The rates of inactivation were found to follow first-order inactivation kinetics. Activation energies varied between 101.18 and 208.42 kJ mol⁻¹ with z values of 10.59–24.09 °C. The examined kinetics indicated that lipoxygenase was the most heat resistant followed by peroxidase, polyphenol oxidase, pectin methyl esterase and ascorbic acid oxidase. Furthermore, the obtained results from the blanched fresh tissues indicated that inactivation of lipoxygenase secured disappearing of any other enzyme activities. Therefore, this study recommends using lipoxygenase as an indicator enzyme to optimize the thermal treatments of cauliflower products.     

INACTIVATION OF ESCHERICHIA COLI IN SKIM MILK BY HIGH INTENSITY PULSED ELECTRIC FIELDS

The inactivation of microorganisms is the most important function in the processing of milk and dairy products. Traditionally, this purpose is realized by thermal treatment, but heat produces alterations to flavor and taste in addition to nutrient loss. The high intensity pulsed electric field (PEF) treatment should be a good alternative to heat because demonstrations have shown PEF can reduce the Escherichia coli survival fraction in aqueous solutions and model foods. In this study, PEF treatment was found to inactivate E. coli in skim milk (inoculum 10⁹ CFU/mL) at 15C. The microorganism inactivation satisfied Hülsheger's model following a first order kinetic for both the electric field intensity and number of pulses when skim milk inoculated with E. coli was treated in a static or continuous flow chamber. PEF treatment in a continuous system when the critical electric field (E_c) and minimum number of pulses (n_min) were 12.34 kV/cm and 2.7 at 30 kV/cm and 30 pulses (0.7–1.8 μs pulse width) inactivated more microorganisms than in a static system. It has also been proven that increasing the pulse duration increases the E. coli inactivation. The inactivation of E. coli using PEF is more limited in skim milk than in a buffer solution when exposed to similar treatment conditions of field intensity and number of pulses due to the complex composition of skim milk, its lower electrical resistivity and the presence of proteins.     

A Gompertz Model Approach to Microbial Inactivation Kinetics by High‐Pressure Processing (HPP): Model Selection and Experimental Validation

A recently proposed Gompertz model (GMPZ) approach describing microbial inactivation kinetics by high‐pressure processing (HPP) incorporated the initial microbial load (N₀) and lower microbial quantification limit (N_lim), and simplified the dynamic effects of come‐up time (CUT). The inactivation of Listeria innocua in milk by HPP treatments at 300, 400, 500, and 600 MPa and pressure holding times (t_hold) ≤10 min was determined experimentally to validate this model approach. Models based on exponential, logistic‐exponential, and inverse functions were evaluated to describe the effect of pressure on the lag time (λ) and maximum inactivation rate (μ_max), whereas the asymptote difference (A) was fixed as A = log₁₀(N₀/N_lim). Model performance was statistically evaluated and further validated with additional data obtained at 450 and 550 MPa. All GMPZ models adequately fitted L. innocua data according to the coefficient of determination (R² ≥ 0.95) but those including a logistic‐exponential function for μ_max(P) were superior (R² ≥ 0.97). These GMPZ versions predicted that approximately 597 MPa is the theoretical pressure level (P_λ) at which microbial inactivation begins during CUT, mathematically defined as λ (P = P_λ) = t_CUT, and matching the value observed on the microbial survival curve at 600 MPa. As pressure increased, predictions tended to slightly underestimate the HPP lethality in the tail section of the survival curve. This may be overseen in practice since the observed microbial counts were below the predicted log₁₀ N values. Overall, the modeling approach is promising, justifying further validation work for other microorganisms and food systems.     

Prediction of<Emphasis Type="Italic"> Bacillus subtilis</Emphasis> spore survival after a combined non-isothermal-isothermal heat treatment

Different inactivation kinetics data have been used to predict the number of survivors exposed to a heat treatment and, in consequence, to design thermal processes for the food industry. In this work, spores of an acidophilic strain of Bacillus subtilis were heated under isothermal and non-isothermal conditions. Experimental results obtained after isothermal treatments were analysed using the classical two-step linear regression procedure and a one-step non-linear regression method. Data obtained after non-isothermal treatments were analysed using a one-step, non-linear procedure. Kinetic parameters obtained from isothermal heating were close, either using the two-step linear regression (D₁₀₀=6.5 min) or the one-step non-linear regression (D₁₀₀=6.3 min), although the second method gave smaller 95% confidence intervals. The z values derived from non-isothermal heating were higher than those obtained in isothermal conditions (z=9.3 °C for non-isothermal heating at 1 °C/min versus z=7.7 °C for isothermal heating one step non-linear regression). Results were validated with experimental data obtained after different heat treatments, consisting of a phase of temperature increase at a fixed rate, followed by a holding phase. Non-isothermal methods predicted accurately the number of survivors after the heating ramp, while isothermal methods were more accurate for the holding phase of the treatment. When a temperature profile of a typical heat treatment process applied in the food industry was simulated, all predictions were on the safe side.     

Preliminary study of L‐lysine production by Bacillus species using various agricultural by‐products

The production of lysine by Bacillus megaterium SP‐14 and Bacillus circulans Tx‐22 using agricultural by‐products as carbon and nitrogen sources was assessed. Among the carbon substrates used were potato, sorghum, plantain, millet, yam, cassava, and corn starches, while the nitrogen sources include cowpea, bambara‐nut, cotton seed, groundnut, soybean, and blood meals. The effect of natural nitrogen sources (1.0% w/v) and synthetic nitrogen source (4.0% w/v (NH₄)₂SO₄) on lysine production by the Bacillus strains showed that natural nitrogen sources gave better lysine yields.     

Thermal inactivation of polyphenoloxidase and peroxidase in green coconut (Cocos nucifera) water

Coconut water is an isotonic beverage naturally obtained from the green coconut. After extracted and exposed to air, it is rapidly degraded by enzymes peroxidase (POD) and polyphenoloxidase (PPO). To study the effect of thermal processing on coconut water enzymatic activity, batch process was conducted at three different temperatures, and at eight holding times. The residual activity values suggest the presence of two isoenzymes with different thermal resistances, at least, and a two‐component first‐order model was considered to model the enzymatic inactivation parameters. The decimal reduction time at 86.9 ^°C (D_{86.9 °C}) determined were 6.0 s and 11.3 min for PPO heat labile and heat resistant fractions, respectively, with average z‐value = 5.6 °C (temperature difference required for tenfold change in D). For POD, D_{86.9 °C} = 8.6 s (z = 3.4 °C) for the heat labile fraction was obtained and D_{86.9 °C} = 26.3 min (z = 6.7 °C) for the heat resistant one.     

Inactivation Kinetics of Enzymes by Using Continuous Treatment with Microbubbles of Supercritical Carbon Dioxide

ABSTRACT Enzyme inactivation using a new apparatus for continuous treatment with microbubbles supercritical carbon dioxide (SC-CO₂) was investigated. D value of a-amylase (5.0±1.2 min) subjected to microbubbles of SC-CO₂ treatment (microbubbles-SCT) at 35 °C, 30 MPa was lower than that (227 ± 15.9 min) subjected to heat treatment (HT) at 70 °C. D value of acid protease was reduced by microbubbles-SCT at 50 °C, 30 MPa (15.4 ± 4.1 min), compared to HT at 50 °C (233 ± 15.2 min). The activation energy for the inactivation of acid protease (135 ± 8.3 kJ mol^-1) by microbubbles-SCT was 1 half of that (259 ± 9.0 kJ mol^-1) by HT. These results indicated that continuous treatment with microbubbles of SC-CO₂ was effective for enzyme inactivation.     

UV-C inactivation of Escherichia coli at different temperatures

The influence of treatment parameters (dose and temperature), treatment medium characteristics (absorption coefficient, pH and water activity) and microbiological factors (strain, growth phase and UV damage and repair capacity) on Escherichia coli UV-C resistance has been investigated. UV-C doses to inactivate at 25 °C 99.99% of the initial population (4D) of five strains of E. coli in McIlvaine buffer of pH 7.0 with tartrazine added (absorption coefficient of 10.77 cm⁻¹) were 16.60, 14.36, 14.36, 13.22, 11.18 J/mL for strains E. coli STCC 4201, STCC 471, STCC 27325, O157:H7 and ATCC 25922, respectively. The entrance in the stationary growth phase increased the 4D value of the most resistant strain, E. coli STCC 4201, from 13.09 to 17.23 J/mL. Survivors to UV treatments showed neither oxidative damages nor injuries in cell envelopes. On the contrary, the photoreactivation by the incubation of plates for 60 min below visible light (11.15 klx) increased the dose to 18.97 J/mL. The pH and the water activity of the treatment medium did not affect the UV tolerance of E. coli STCC 4201, but the lethal effect of the treatments decreased exponentially (Log₁₀4D = − 0.0628α + 0.624) by increasing the absorption coefficient (α). A treatment of 16.94 J/mL reached 6.35, 4.35, 2.64, 1.93, 1.63, 1.20, 1.02 and 0.74 Log₁₀ cycles of inactivation with absorption coefficients of 8.56, 10.77, 12.88, 14.80, 17.12, 18.51, 20.81 and 22.28 cm⁻¹. The temperature barely changed the UV resistance up to 50.0 °C. Above this threshold, inactivation rates due to the combined process synergistically increased with the temperature. The magnitude of the synergism decreased over 57.5 °C. An UV treatment of 16.94 J/mL in media with an absorption coefficient of 22.28 cm⁻¹ reached 1.23, 1.64, 2.36, 4.01 and 6.22 Log₁₀ cycles of inactivation of E. coli STCC 4201 at 50.0, 52.5, 55.5, 57.5 and 60.0 °C, respectively.

Industrial relevance

Results obtained in this investigation show that UV light applied at mild temperatures (57.5 to 60 °C) could be an alternative to heat treatments for 5-Log₁₀ reductions of E. coli in liquid foods. Since microbial resistance to UV-C light did not depend on the pH and water activity (a_w) of the treatment media, eventual advantages of UV light for pasteurization purposes will be higher in low a_w foods. E. coli STCC 4201 could be considered as a target when UV light processing of foods.     

The contribution of high pressure carbon dioxide in the inactivation kinetics and structural alteration of myrosinase

High pressure carbon dioxide (HPCD) has been verified to be an efficient way of inactivating enzyme activity. This work investigates the influence of temperature (T), pressure (P), exposure times (t) on the activity of commercial myrosinase (MYR) submitted to HPCD. Results showed that only 1.00% of MYR activity retained at 22 MPa and 65 °C for 5 min. Moreover, the first‐order reaction kinetic data of MYR inactivation as influenced by pressure of HPCD were analysed. With the pressure rising from 8 to 22 MPa at 55 °C, the inactivation rate constant (k) increased from 0.015 to 0.024 min^?1, while the decimal reduction time (D) decreased from 157.2 to 96.1 min. Additionally, a series of exploratory experiments were conducted to investigate the contribution of the HPCD parameters (T, P and CO₂ dissolution), with analysing circular dichroism spectroscopy and tryptophan fluorescence spectra, illustrate that CO₂ dissolution plays a dominant role in MYR inactivation and structural alteration.     

Thermal inactivation kinetics of peroxidase and lipoxygenase from fresh pinto beans (Phaseolus vulgaris)

 Thermal inactivation kinetics of crude peroxidase (POX) and lipoxygenase (LOX) in fresh pinto beans were studied over the temperature range of 55–90°C. The inactivation of both enzymes followed first-order kinetics. The biphasic inactivation curves for POX indicate the existence of several isoenzymes of varying heat stability. In the temperature range of 55–70°C, the activation energies (E _a) of POX were 46.5 kcal·mol^–1 for the heat-labile portion and 37.6 kcal·mol^–1 for the heat-stable portion. On the other hand, the LOX enzyme had an E _a value of 42.26 kcal·mol^–1 at 55–75°C and 49.1 kcal·mol^–1 at 55–90°C. Received: 28 July 1997 / Revised version: 16 October 1997     

Thermal inactivation kinetics of peroxidase and lipoxygenase from fresh pinto beans (Phaseolus vulgaris)

 Thermal inactivation kinetics of crude peroxidase (POX) and lipoxygenase (LOX) in fresh pinto beans were studied over the temperature range of 55–90°C. The inactivation of both enzymes followed first-order kinetics. The biphasic inactivation curves for POX indicate the existence of several isoenzymes of varying heat stability. In the temperature range of 55–70°C, the activation energies (E _a) of POX were 46.5 kcal·mol^–1 for the heat-labile portion and 37.6 kcal·mol^–1 for the heat-stable portion. On the other hand, the LOX enzyme had an E _a value of 42.26 kcal·mol^–1 at 55–75°C and 49.1 kcal·mol^–1 at 55–90°C. Received: 28 July 1997 / Revised version: 16 October 1997     

E‐beam irradiation affects the maximum specific growth rate of Bacillus cereus

When conventional preservative treatments are applied, such as heat or acid, the maximum specific growth rate (μ_max) of survivors is the same as that of untreated cells. However, when new nonthermal technology is applied, the effects of it on the kinetics of the microorganism can be unpredictable. In this sense, Cabeza et al. (2010) reported longer doubling times after irradiating with accelerated electron beam. The aim of this work was to study the effect of electron beam irradiation on the μ_max of Bacillus cereus and compare it with a conventional inactivation treatment (heat). To prove this, μ_max was estimated in ham at 12 °C and in TSB at 22 °C after 0, 2, 3 or 4 log reduction by irradiation; likewise, μ_max was estimated in whole milk at 12 °C and in TSB at 22 °C after the same log reduction using heat treatments. Our findings show that irradiation affected the μ_max of survivor cells. Irradiation intensity was inversely proportional to μ_max, such that greater intensity was associated with lower μ_max. At the same time, growth temperature had an effect on the decrease in μ_max: the radiation‐induced reductions in μ_max were greater at 12 °C than at 22 °C. In summary, E‐beam irradiation decreases the μ_max of B. cereus, while heat treatment does not. This suggests that the shelf life of irradiated foods must be longer than that of heat‐preserved foods after the application of a similar inactivation treatment.     

Kinetics of Thermal Inactivation of Peroxidase and Color Degradation of African Cowpea (Vigna unguiculata) Leaves

Cowpea leaves form an important part of the diet for many Kenyans, and they are normally consumed after a lengthy cooking process leading to the inactivation of peroxidase (POD) that could be used as an indicator for the potential shelf life of the vegetables. However, color degradation can simultaneously occur, leading to poor consumer acceptance of the product. The kinetics of POD in situ thermal (for thermal treatments in the range of 75 to 100 °C/120 min) inactivation showed a biphasic first‐order model, with Arrhenius temperature dependence of the rate constant. The kinetic parameters using a reference temperature (T_ref) of 80 °C were determined for both the heat‐labile phase (k_ref = 11.52 ± 0.95 × 10^?2 min^?1 and E_a of 109.67 ± 6.20 kJ/mol) and the heat‐stable isoenzyme fraction (k_ref = 0.29 ± 0.07 × 10^?2 min^?1 and E_a of 256.93 ± 15.27 kJ/mol). Color degradation (L*, a*, and b* value) during thermal treatment was investigated, in particular as the “a*” value (the value of green color). Thermal degradation (thermal treatments between 55 and 80 °C per 90 min) of the green color of the leaves followed a fractional conversion model and the temperature dependence of the inactivation rate constant can be described using the Arrhenius law. The kinetic parameters using a reference temperature (T_refC = 70 °C) were determined as k_refC = 13.53 ± 0.01 × 10^?2 min^?1 and E_aC = 88.78 ± 3.21 kJ/mol. The results indicate that severe inactivation of POD (as an indicator for improved shelf life of the cooked vegetables) is accompanied by severe color degradation and that conventional cooking methods (typically 10 min/100 °C) lead to a high residual POD activity suggesting a limited shelf life of the cooked vegetables.     

Bacterial resistance after pulsed electric fields depending on the treatment medium pH

The objective was to evaluate and compare the pulsed electric field (PEF) resistance of four Gram-positive (Bacillus subtilis, Listeria monocytogenes, Lactobacillus plantarum, Staphylococcus aureus) and four Gram-negative (Escherichia coli, E. coli O157:H7, Salmonella serotype Senftenberg 775W, Yersinia enterocolitica) bacterial strains under the same treatment conditions. Microbial characteristics such as cell size, shape or type of the cell envelopes did not exert the expected influence on microbial PEF resistance. The most PEF resistant bacteria depended on the treatment medium pH. For instance, L. monocytogenes, which showed the highest PEF resistance at pH 7.0, was one of the most sensitive at pH 4.0. The most PEF resistant strains at pH 4.0 were the Gram-negatives E. coli O157:H7 and S. Senftenberg. A subsequent holding of PEF-treated cells in pH 4.0 for 2 h increased the degree of inactivation up to 4 extra Log₁₀ cycles depending on the bacterial strain investigated. Under these treatment conditions, the most PEF resistant bacterial strains were still the pathogens S. Senftenberg and E. coli O157:H7.

Industrial relevance

The design of appropriate food preservation processes by PEF requires the selection of an adequate target bacterial strain, which should correspond to the most PEF resistant microorganism contaminating food. This study indicates that the pH of the treatment medium plays an important role in determining this target bacterial strain. On the other hand, the combination of PEF and subsequent holding under acidic conditions has been proven to be an effective method in order to achieve a higher level of microbial inactivation.     

COD (GADUS MORHUA) TRYPSIN HEAT INACTIVATION: A REACTION KINETICS STUDY

The heat inactivation of cod (Gadus morhua) trypsin was examined at 40–55C by nonlinear regression (NLR) analysis according to 1st-order, n-th order, or consecutive reaction kinetics. At 40C Cod trypsin was inactivated by a 2nd-order reaction; n = 2 (±0.014) with a pseudo rate constant (k′) of 6.46 (±0.04) × 10^?4 (s^?1). The order of reaction decreased with increasing temperature; n = 1.54 (±0.003) at 45C, n = 0.8 (±0.005) at 50C or n = 0.64 (±0.014) at 55C. At 45–55C the consecutive model also indicated 1st-order inactivation kinetics. SDS-PAGE analysis showed that with progressive heating the concentration of cod trypsin decreased and there was an appearance of lower molecular weight polypeptides. The results are consistent with a heat inactivation process involving trypsin autolysis. A mechanism for cod trypsin heat inactivation is proposed which accounts for the observed 1st-order or 2nd-order reaction kinetics as limiting cases.     

Thermal Stabilities of Peroxidases from Fresh Pinto Beans

Heat stabilities of crude and partially purified soluble (SPOX), ionically bound (IPOX) and total peroxidase (TPOX) from fresh pinto beans were investigated at 55–90°C. Heat inactivation of peroxidase (POX) followed first-order reaction kinetics. Each inactivation curve consisted of two linear parts: initial rapid inactivation (heat-labile) followed by slower inactivation (heat-stable). IPOX showed activation during heat treatment with a highly heat-stable isoenzyme (D₉₀=40 min) which was more heat-stable than SPOX. Activation energies for heat-stable parts of crude IPOX and SPOX were, respectively, 12.1 and 36.4 kcalmol-1 with z values 45.4 and 14.1C°. Heat stable SPOX isoenzymes (D₇₀=22.6) were obtained by 65–95% (NH₄)₂SO₄ precipitation from crude SPOX. Two POX fractions (F1 and F2) were separated from TPOX by ion-exchange chromatography.