Original article: Modelling the effects of food ingredients and pH on high‐pressure processing inactivation of Bacillus cereus spores: a laboratorial study

The combination of high pressure and heat on Bacillus cereus spores in food matrix was investigated with the purpose of achieving a predictive model of microbial inactivation. The high‐pressure processing (HPP) conditions were fixed at 540 MPa and 71 °C for 16.8 min, which were determined as the optimum conditions considering six‐log‐cycle reductions of B. cereus spores. The effects of soybean protein, sucrose, soybean oil and pH on the inactivation of B. cereus spores by HPP were evaluated, and a quadratic predictive model for the effects of food ingredients and pH on the reductions of B. cereus spores by HPP was built using response surface methodology. The experimental results showed that soybean protein, sucrose and pH significantly affected the reductions of B. cereus spores. The predictive model is significant, because the level of significance was P < 0.0001 and the calculated F‐value is much greater than the tabulated F‐value. The adequacy of the predictive model equation was verified effectively using the experimental test data that were not used in the development of the model.     

THERMAL RESISTANCE OF BACILLUS STEAROTHERMOPHILUS SPORES IN DRIED PASTA AT DIFFERENT STAGES OF REHYDRATION

The thermal resistance of Bacillus stearothermophilus spores at different stages of rehydration was examined. Inoculation was achieved by rehydrating pasta samples in water containing 1‐mL spore crop (10⁶ spores/mL); the product was heat sealed in sterile pouches and processed in a laboratory retort. Thermal inactivation of spores was carried out using 10 time intervals at 121C for moisture contents, 70, 90, 105, 115, 125 and 145% dry basis. The death curves obtained consisted of two phases, an initial rapid decline (1.5 to 2.6 log reductions) followed by a slower, linear decrease of survivors from which the decimal reduction time (D value) was calculated. D₁₂₁ values were found to decrease with increasing moisture content and ranged from 4.6 to 6.5 min. The thermal resistance constant (z value) was examined to assess the effect of temperature (110–125C) on lethality. z values decreased with increasing moisture content and ranged from 10.7 to 15.6C. The results of this study demonstrate that temperature and moisture content influence the effectiveness of the intended heat process. For products that are rehydrated during heat treatment, the moisture content of the product is a critical factor. Therefore, if complete rehydration is not achieved, this may result in the heat process being less effective for the destruction of microorganisms, which may have implications in regard to food safety.     

Effect of thermosonication in a batch system on the survival of spore‐forming bacteria

Thermosonication may help reduce bacteria counts responsible for spoilage in dairy products. Vegetative cells and spores of Geobacillus stearothermophilus, Anoxybacillus flavithermus and Bacillus subtilis (spores only) were treated with either heat alone or thermosonication in a batch system from 0 to 120 s in tryptic soy broth and 2% fat milk at 72 and 73 °C. D‐values for vegetative cells were calculated and were reduced after thermosonication. Maximum reduction in vegetative cells after thermosonication was 1 log after 30–45 s and for spores was ≤0.2 log after 120 s, which may not influence dairy product quality in scale‐up systems.     

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     

Optimization and evaluation of heat-shock condition for spore enumeration being used in thermal-process verification: Differential responses of spores and vegetative cells of Clostridium sporogenes to heat shock

To evaluate a heat-shock condition for the enumeration of Clostridium sporogenes spores, a surrogate for C. botulinum spores, we examined the heat tolerance of C. sporogenes spores and vegetative cells exposed to a heat shock at 90°C. From the D values of the spores determined in the temperature range of 113–121°C, z value (±SD) and D_90°C value were estimated to be 10.16±0.90°C and 1,071.52 min, respectively, and the inactivation rates were predicted to be only approximately 2% at 90°C for up to 10 min. Meanwhile, the viable count of spores was significantly higher when activated under a heat-shock condition of 90°C for over 9 min than those activated for shorter time periods. The heat tolerance of vegetative cells was extremely low, showing a D_90°C value (±SD) of 0.21±0.01 min. Finally, 3 different heat-shock conditions were compared: 70°C for 30 min, 80°C for 20 min, and 90°C for 10 min, and the experimental comparative data showed no significant differences in viable spore counts. Consequently, these results support that the heat-shock treatment at 90°C for 10 min is suitable to activate spores and to inactivate vegetative cells of C. sporogenes.     

Heat‐resistance characteristics of ascospores of Eurotium chevalieri isolated from apricot juice

A heat‐resistant fungus was isolated from aseptically packaged apricot pulp. The fungus was identified asEurotium chevalieri. Heat resistance of the fungus was studied at four different temperatures (70, 75, 80 and 83°C) after activation of its ascospores for 30 min at 70°C. D₇₀, D₇₅, D₈₀ and D₈₃ values of ascospores ofEurotium chevalieri were estimated by linear regression (log‐survival vs. heating time) as 118.58, 34.15, 5.50 and 3.77 min, respectively. The z‐value was determined in the same way (regression of log‐D values vs. heating temp.) and was found as 8.23°C.     

Effect of rotation on the heat penetration characteristics of thermally processed tuna in oil in retort pouches

Tuna in oil was processed in retort pouches (20 × 17 cm) to a, F₀ value of 10. Heat penetration characteristics were determined using a rotary retort to an F₀ value of 10 and the results compared with a stationary retort. For studying the changes in heat penetration during rotation, tuna in oil was packed into retort pouches, subjected to different rotation speeds (2, 4 and 6 r.p.m.) and thermally processed to the same F₀ value. For the same F₀ value of 10, although there was a reduction in process time with increasing rotation speed up to 6 r.p.m., the magnitude of the reduction in processing time decreased with increase in speed of rotation. However up to 2 r.p.m. there was a considerable reduction in process time.     

Microbiological Validation of Microwave-Circulated Water Combination Heating Technology by Inoculated Pack Studies

A 915‐MHz Microwave‐Circulated Water Combination (MCWC) heating technology was validated for a macaroni and cheese product using inoculated pack studies. Before the tests, heat resistances of a Clostridium sporogenes (PA 3679) spore crop were determined in neutral phosphate buffer and macaroni and cheese product. Trays of macaroni and cheese products were subjected to 3 processing levels: target process (F₀= 2.4), under target process (F₀= 1.2), and over target process (F₀= 4.8). The inoculated packs were evaluated by count‐reduction method and end‐point method. The microbial results showed that microbial destruction resulting from MCWC heating technology matched the calculated degree of sterilization (F₀ value). This study suggests that the MCWC heating technology has potential in sterilizing packaged foods.     

Estimation of the non-isothermal inactivation patterns of Bacillus sporothermodurans IC4 spores in soups from their isothermal survival data

The isothermal survival curves of the heat resistant spores of Bacillus sporothermodurans IC4, in the range of 117-125 degrees C, were determined in chicken, mushroom and pea soups by the capillary method. They were all non-linear with a noticeable upper concavity and could be described by the equation log(10) [N(t)/N(0)]=-b(T)t(n) with a fixed power, n, of the order of 0.7-0.8. The temperature dependence of b(T) could be described by the equation b(T)=log(e)[1+exp[k(T-T(c))]], where T(c) is the temperature where intensive inactivation starts and k is the slope of b(T) at temperatures well above T(c). They were 121-123 degrees C and 0.2-0.4 degrees C(-1), respectively, depending on the soup. These parameters were used to estimate the survival curves of the spores in two non-isothermal heat treatments using the procedure originally proposed by Peleg and Penchina [Crit. Rev. Food Sci. Nutr. 40 (2000) 159]. The results were compared with experimental survival curves, determined by the direct injection method, in another laboratory. There was a general agreement, although not perfect, between the predicted and experimentally observed survival ratios. Also, the isothermal survival parameters, estimated directly from the non-isothermal inactivation data using the model, were in general agreement with those calculated from the isothermal data. This suggests that the heat inactivation patterns of B. sporothermodurans IC4 spores in soups can be at least roughly estimated using the same general survival model, which has until now only been experimentally validated for vegetative bacterial cells.     

FIRMNESS OF THERMAL PROCESSED ONION AS AFFECTED BY BLANCHING

The pectin methyl esterase enzyme system was shown to be involved in firmness of thermally treated onion in the temperature range 50–70C. Thermal softening of onion at 90 and 100C showed an initial steep negative slope with a shallow negative slope at longer heating time. Low‐temperature blanching at 70C was effective to maintain firm onion tissue exposed to excessive heating. Physical strength of onion was substantially diminished when exposed to a commercial sterilization condition where F₀, extent of thermal sterilization, was 3, and/or more. Blanching in water for 120 min at 70C resulted in a maximum value for the firmness of commercially sterilized onion. Firmness of onion, blanched in calcium brine at a concentration range of 0.0–1.0% (wt) prior to heat treatment, decreased with increasing severity of thermal sterilization treatment. At F₀ = 6, blanching in 0.5% calcium brine resulted in maximum firmness of thermally sterilized onion, approximately 70% of that of raw onion.     

Reliability‐based estimation of the survival of Listeria monocytogenes in chorizos

Reliability analysis is especially important when critical decisions are to be made involving potentially severe adverse consequences such as foodborne illness. Owing to uncertainty associated with the parameters controlling survival of Listeria monocytogenes in chorizo (a Mexican‐style sausage), the time needed to reduce the count by a certain number (n) of logs (t_nD) is probabilistic. In this paper the first‐order second‐moment (FOSM) method based on Taylor series expansion is used to derive the expected value and standard deviation of t_nD as function of the operating conditions (random variables) affecting survival, namely initial water activity (a_w0) of the sausage batter, storage temperature (T) and airflow velocity (F), along with their uncertainties characterised by their means and coefficients of variation. For any given n the derived t_nD probability distribution enables one to determine an estimate of t_nD for any desired level of reliability or confidence level, such as 50% (median value), 95%, 99%, etc. Among the conclusions drawn were: (i) the variability associated with T and F has a minor effect on estimating uncertainty in t_nD, whereas the reliability of t_nD estimation is greatly influenced by the uncertainty in a_w0; and (ii) the uncertainty in a_w0 has the greatest impact when a_w0 of the sausage formulation exceeds 0.90. The approach used and discussed in this paper can be applied to any survival/inactivation study to incorporate the effect of uncertainty in the various extrinsic and intrinsic parameters on the survival kinetics of the pathogen in a food system under evaluation. Copyright © 2006 Society of Chemical Industry     

Evaluation and Quantification of Heat Treatments Applied to Food Preserves

The quantitative study of heat treatments for sterilisation uses the Bigelow model to calculate the sterilising value (F). Calculation of F requires the previous determination of parameters D (decimal reduction time at experimental temperature) and Z (thermal-death time parameter), obtained from the thermal-death kinetics. Herein we compare two different methods, namely the Bigelow model and a predictive-type statistical method, to calculate the sterilisation effect against Bacillus coagulans spores when heat was applied to runner bean preserves (variety: Helda). Samples were subjected to various autoclave treatments at working temperatures (T _ai) of 105, 107, 110, and 115°C for periods from 3 to 35 min. The microorganism used was B. coagulans. Sterilisation achieved by these autoclave treatments was determined by using the equation based on the Bigelow model (n _probe = F ^z _Ti/D _Ti) where n is the fractional concentration of colony-forming units (or some quality factor), F ^z _Ti is F at temperature T _i, and D _Ti is D at temperature T _i. The Bigelow model can be used to obtain Z (thermal-death time parameter), which is needed to calculate the traditional sterilisation factor F, but not to determine the reduction factor n for the heat treatments, particularly when microbial indicators with low decimal reduction times (D) are studied. The thermokinetic parameters for B. coagulans in runner bean solution resulted to be Z = 10.64°C and D ₁₂₁ = 0.0264 min (Af = 1.04). Treatment at 115°C for 20 min resulted in the most efficient sterilisation effect for B. coagulans.     

A Controlled Agitation Process for Improving Quality of Canned Green Beans during Agitation Thermal Processing

This work introduces the concept of a controlled agitation thermal process to reduce quality damage in liquid‐particulate products during agitation thermal processing. Reciprocating agitation thermal processing (RA‐TP) was used as the agitation thermal process. In order to reduce the impact of agitation, a new concept of “stopping agitations after sufficient development of cold‐spot temperature” was proposed. Green beans were processed in No. 2 (307×409) cans filled with liquids of various consistency (0% to 2% CMC) at various frequencies (1 to 3 Hz) of RA‐TP using a full‐factorial design and heat penetration results were collected. Corresponding operator's process time to impart a 10‐min process lethality (F_o) and agitation time (AT) were calculated using heat penetration results. Accordingly, products were processed again by stopping agitations as per 3 agitation regimes, namely; full time agitation, equilibration time agitation, and partial time agitation. Processed products were photographed and tested for visual quality, color, texture, breakage of green beans, turbidity, and percentage of insoluble solids in can liquid. Results showed that stopping agitations after sufficient development of cold‐spot temperatures is an effective way of reducing product damages caused by agitation (for example, breakage of beans and its leaching into liquid). Agitations till one‐log temperature difference gave best color, texture and visual product quality for low‐viscosity liquid‐particulate mixture and extended agitations till equilibration time was best for high‐viscosity products. Thus, it was shown that a controlled agitation thermal process is more effective in obtaining high product quality as compared to a regular agitation thermal process.     

The fate of Clostridium perfringens spores exposed to ozone and/or mild heat pretreatment on beef surfaces followed by modified atmosphere packaging

Clostridium perfringens is a natural contaminant of raw beef products that can proliferate to dangerous cell levels under conditions of temperature abuse. Spores of the bacterium were inoculated onto irradiated London broil beef at levels of 3 log₁₀ spores/g beef. Samples of beef (7.5×10.0×1.0 cm) were treated with aqueous ozone (5 ppm O₃ for 5 min), or heat (60°C for 30 min), or both and then vacuum-packaged to 2 kPa for up to 10 d storage at 37°C, 25°C, or 4°C. Storage at 37°C resulted in increases in viable counts after 1 d to over 7 log₁₀ cfu/g beef, whereas storage at 4°C prevented spore germination and growth for all treatments. At 25°C, heat-treated beef samples reached 6 log₁₀ cfu/g viable counts in 2 d and spores/vegetative cells on control or ozone-treated samples did not germinate or grow through the first day of vacuum-packaged storage. Modified atmospheres with increasing CO₂ concentration were also compared with regard to bacterial survival during beef storage at 25°C. C. perfringens spores remained dormant in control and ozone-treated beef during a 10-d storage at 25°C. Pretreatment with heat increased germination and outgrowth during storage of beef, whereas ozone treatment and no treatment controls were effective in inhibiting spore germination and outgrowth in combination with increasing CO₂ concentrations above 30% or refrigeration. These data support the avoidance of heat in the pretreatment of raw beef.     

Thermal resistance of aerobic spore formers isolated from food products

The heat resistance of aerobic spore formers isolated from dairy products and cocoa powder was examined to give an overview of occurring highly heat‐resistant spores. Experiments were conducted in phosphate buffer at different temperatures for 30 min. Two Geobacillus pallidus strains survived the heat treatment at 125 °C with log reductions of 6.68 and 6.73. Furthermore, the inactivation kinetics of one of these strains (Bacillus amyloliquefaciens) were determined using a modified Arrhenius model. The inactivation followed a reaction order of 1 with a reaction rate constant (k_ref) of 0.085/s at 394 K and an activation energy (E_a) of 209 kJ/mol.     

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.     

The Elimination of Listeria Monocytogenes Attached to Stainless Steel or Aluminum Using Multiple Hurdles

Ready‐to‐eat luncheon meats sliced in retail delis have been found to pose the greatest risk of foodborne illness from Listeria monocytogenes among all ready‐to‐eat foods. Slicers used in delis have many removable parts that are connected with seals and gaskets, with spaces, cracks and crevices that are difficult to clean adequately and may provide a niche for L. monocytogenes survival. Standard cleaning and sanitizing practices used by deli employees may not eliminate Listeria in these niches. Moist heat is known to be more effective against L. monocytogenes than dry heat at the same temperature and time. The study reported here investigated the effects of moist heat combined with quaternary ammonium compounds (5 or 10 ppm), chlorine (10 or 25 ppm) or peracetic acid (10 or 25 ppm) on inactivating L. monocytogenes attached to stainless steel or aluminum coupons cut from commercial deli meat slicer components. All sanitizers when used alone resulted in a 2‐ to 3‐log reduction of L. monocytogenes on stainless steel or aluminum surfaces, while moist heat alone resulted in a 3‐ to 4‐log reduction. When combined with heat the quaternary ammonium was used at 5 ppm, peracetic acid at 10 ppm and chlorine at 10 ppm. When the 2 lethal treatments were combined there was a 5‐ to7‐log reduction as compared to initial inoculation.     

MATHEMATICAL MODEL FOR THE SURVIVAL OF LISTERIA MONOCYTOGENES IN MEXICAN-STYLE SAUSAGE

Survival of Listeria monocytogenes in chorizos (Mexican‐style sausages) was modeled in relation to initial water activity (a_w0) and storage conditions using the Weibull cumulative distribution function. Twenty survival curves were generated from chorizos formulated at a_w0 = 0.85–0.97 then stored under four temperature (T) and air inflow velocity (F) conditions. The Weibull model parameters (α and β) were determined for every curve. Predicted survival curves agreed with experimental curves with R² = 0.945–0.992. Regression models (R² = 0.981–0.984) were developed to relate α and β to operating conditions. The times to one‐ and two‐log reduction in count (t_1D and t_2D) were derived from the Weibull model in terms of α and β. A parametric study revealed that L. monocytogenes survival was most sensitive to a_w0 between 0.90 and 0.95. The inactivation of L. monocytogenes could be maximized with higher T and lower a_w0; however, F did not significantly influence survival.     

Performance evaluation of aluminum test cell designed for determining the heat resistance of bacterial spores in foods

Thermal inactivation kinetic studies are necessary to determine heat resistances of spores in the development of new thermal processes for low-acid shelf-stable products. Most currently available sample holders used for solid and semi solid samples in the kinetic studies take long time to reach the target sample temperature, hence fail to provide isothermal condition. In this research, novel aluminum test cells were developed to facilitate easy loading and unloading samples in a hermetically sealed 1 ml cavity to evaluate the heat resistance of bacterial spores when heated at temperatures above 100 °C. Design of the test cell was governed by minimum come-up time. A finite element model based on the commercial software ‘FEMLAB’ was used to simulate transient heat transfer and finalize the test cell dimensions. Performance of the new test cell was evaluated against capillary and aluminum thermal death time tube methods in characterizing the heat resistance of Clostridium sporogenes PA 3679 spores in a phosphate buffer and mashed potato at 121 °C. D₁₂₁ values of PA 3679 spores in both the phosphate buffer and mashed potato using the new test cells were not significantly different (P>0.05) from those by the capillary tube method. The results indicated that the new test cell is appropriate for studying the inactivation kinetics of bacterial spores in microbial validation of conventional and novel thermal processes for low-acid shelf-stable foods.     

Mechanochemical‐assisted extraction and antioxidant activity of polysaccharides from Ganoderma lucidum spores

Mechanochemical‐assisted extraction (MCAE) method was developed to an effective method for polysaccharides extraction from Ganoderma lucidum spores. The MCAE parameters and the antioxidant activity of polysaccharides were investigated. Through response surface methodology design experiments, the processing conditions were optimised as follows: material/solid reagent (Na₂CO₃) 5 g·g^?1, milling time 20 min, solution/material ratio twenty (mL·g^?1) and extraction time 130 min. Under these conditions, the yield of polysaccharides was (5.92 ± 0.13)%, which was in close agreement with the predicted value. Compared with the heat‐reflux extraction method, the MCAE method had higher extraction yield, shorter extraction time and lower extraction temperature. In addition, the polysaccharides obtained from MCAE exhibit significant antioxidant activity.