Measurement of optical absorbance of Clostridium sporogenes PA 3679 spores as a method for determining their thermal inactivation

 The loss of optical density of Clostridium sporogenes PA 3679 spores after heating at temperatures of 121, 126, 130 and 135°C was studied, together with the relationship between this parameter and the recovery capacity of the heated spores. The results show that the spores suffered a greater loss of optical density when they were subjected to more severe heating. A linear relationship was observed between the loss of viability and the optical absorbance of the spores. A certain parallel was detected between the heat resistance parameters of the spores D_T (decimal reduction time) and z (thermal inactivation coefficient), and the kinetic parameters D_T ^A and z^A, which describe the reduction in absorbance. Received: 14 April 1997     

Clostridium sporogenes PA 3679 and its uses in the derivation of thermal processing schedules for low-acid shelf-stable foods and as a research model for proteolytic Clostridium botulinum

The putrefactive anaerobe Clostridium sporogenes PA 3679 has been widely used as a nontoxigenic surrogate for proteolytic Clostridium botulinum in the validation of thermal processes for low-acid shelf-stable foods, as a target organism in the derivation of thermal processes that reduce the risk of spoilage of such foods to an acceptable level, and as a research model for proteolytic strains of C. botulinum. Despite the importance of this organism, our knowledge of it has remained fragmented. In this article we draw together the literature associated with PA 3679 and discuss the identity of this organism, the phylogenetic relationships that exist between PA 3679 and various strains of C. sporogenes and proteolytic C. botulinum, the heat resistance characteristics of PA 3679, the advantages and limitations associated with its use in the derivation of thermal processing schedules, and the knowledge gaps and opportunities that exist with regard to its use as a research model for proteolytic C. botulinum. Phylogenetic analysis reviewed here suggests that PA 3679 is more closely related to various strains of proteolytic C. botulinum than to selected strains, including the type strain, of C. sporogenes. Even though PA 3679 is demonstrably nontoxigenic, the genetic basis of this nontoxigenic status remains to be elucidated, and the genetic sequence of this microorganism appears to be the key knowledge gap remaining to be filled. Our comprehensive review of comparative heat resistance data gathered for PA 3679 and proteolytic strains of C. botulinum over the past 100 years supports the practice of using PA 3679 as a (typically fail-safe) thermal processing surrogate for proteolytic C. botulinum.     

Theoretical comparison of a new and the traditional method to calculate Clostridium botulinum survival during thermal inactivation

When published isothermal survival data of Clostridium botulinum spores in the range 101–121 °C were plotted in the form of logS(t) vs t relationships, where S(t) is the momentary survival ratio, they were all non‐linear. They had a noticeable upward concavity, in violation of the assumption that sporal inactivation is a process that follows first‐order reaction order kinetics. They could be described by the power law model logS(t) = ? b(T)t ^n(T), where b(T) and n(T) are temperature‐dependent coefficients of the order of 0.1–6 and about 0.4 respectively. These coefficients were used to construct simulated survival curves under different heating regimes with a recently proposed model. The model is based on the assumption that the local slope of the non‐isothermal survival curve, or the momentary inactivation rate, is determined solely by the momentary temperature and survival ratio, which in turn are functions of the population thermal history. The survival curves calculated with this model differ considerably from those produced by the standard method based on the traditional D and Z values. The shortcomings of the standard model are that these values depend on the number of points taken for the regression, and that its predicted survival ratios depend on the selected reference temperature. The differential equation which is proposed to replace it can be solved numerically using a program such as Mathematica®. Its predictions solely depend on the observed survival patterns under isothermal conditions and not on any preconceived kinetic model. Nevertheless, the method still needs verification with experimental non‐isothermal survival data, as has already been done with Listeria and Salmonella cells. © 2001 Society of Chemical Industry     

A novel partitioning method as a possible tool for investigating meat. I.

 This paper describes a practical application of the interfacial protein enrichment method, called three-phase partitioning (TPP), and outlines its significance in the differentiation of multicomponent protein systems, such as homogenates and drips of different meats (pork, beef, chicken, turkey and wild-boar). The results obtained using the single-protein model system are also reported to demonstrate the basic process and some characteristic features of TPP. For meat-protein-partitioning experiments, ammonium sulphate (41% relative saturation) and 23.7% tert-butanol were used at 20 – 25°C. Using this two-liquid system, a characteristic distribution of three phases (including a semi-solid midlayer) was obtained for homogenates of eight different meats following centrifugation. Compressibility, expressed as a ratio of layer thickness obtained by low-speed (200 g) and medium-speed (4500 g) centrifugation, allowed us to distinguish between beef and pork drips. A good correlation was found between layer thickness and storage time for wild-boar samples. The partitioning presented here can be considered as a rapid and simple method for comparison between raw meat samples of different origin. Received: 9 July 1997 / Revised version: 1 October 1997     

Rocket immunoelectrophoresis (RIE) for determination of potentially allergenic peanut proteins in processed foods as a simple means for quality assurance and food safety

 Peanuts are one of the most allergenic foods known. The presence of hidden allergens in processed food for reasons of mislabelling or cross-contamination expose allergic individuals to unpredictable risks, especially since highly sensitized subjects may experience severe anaphylactic reactions. The protection of consumers requires specific and sensitive methods for the detection of trace amounts of potentially allergenic peanut components. A rocket immunoelectrophoresis (RIE) procedure was developed allowing the detection of even spurious contaminations with peanut protein. For precipitation of peanut protein a commercially available antiserum was used. By amplifying precipitates with a secondary immunodetection step, 20 ng/ml peanut protein in chocolate extract, equivalent to 0.0002% peanut in chocolate, could still be detected. Model chocolate spiked with various amounts of peanut was investigated down to 0.001% peanut (10 ppm), the limit of quantitative determination. The method was optimized for detection of peanut in chocolate samples. Non-chocolate samples had to be standardized with a chocolate matrix prior to analysis in order to obtain a uniform response. Cross-reactivities with other food proteins did not occur. The method showed high recoveries of 85–101% for chocolate samples down to 10 ppm peanut and good reproducibility with coefficients of variation of ≤ 5 % for samples of ≥ 15 ppm peanut protein. The applicability of this method in the detection of peanut protein in various food commodities was demonstrated: two unlabelled products and two products which did not have peanut listed as an ingredient were identified as containing peanut protein. In all cases where peanut was listed, peanut protein could be determined. The results of RIE were always confirmed by those of a new cell-mediator-release assay that is based on a rat basophil leukaemia (RBL) cell-line (RBL-2H3), cells that are a functional equivalent to mucosal mast cells. Measuring the release of β-hexosaminidase resulting from cross-linking of basophil-bound peanut-specific immunoglobulin E, the assay mimics a main event of the allergic type-I reaction. The cell assay was adapted for food matrices and peanut could be detected down to 0.01% which additionally demonstrated in vitro that even trace amounts of peanut protein could elicit allergic reactions. Received: 28 May 1997 / Revised version: 11 July 1997     

Rocket immunoelectrophoresis (RIE) for determination of potentially allergenic peanut proteins in processed foods as a simple means for quality assurance and food safety

 Peanuts are one of the most allergenic foods known. The presence of hidden allergens in processed food for reasons of mislabelling or cross-contamination expose allergic individuals to unpredictable risks, especially since highly sensitized subjects may experience severe anaphylactic reactions. The protection of consumers requires specific and sensitive methods for the detection of trace amounts of potentially allergenic peanut components. A rocket immunoelectrophoresis (RIE) procedure was developed allowing the detection of even spurious contaminations with peanut protein. For precipitation of peanut protein a commercially available antiserum was used. By amplifying precipitates with a secondary immunodetection step, 20 ng/ml peanut protein in chocolate extract, equivalent to 0.0002% peanut in chocolate, could still be detected. Model chocolate spiked with various amounts of peanut was investigated down to 0.001% peanut (10 ppm), the limit of quantitative determination. The method was optimized for detection of peanut in chocolate samples. Non-chocolate samples had to be standardized with a chocolate matrix prior to analysis in order to obtain a uniform response. Cross-reactivities with other food proteins did not occur. The method showed high recoveries of 85–101% for chocolate samples down to 10 ppm peanut and good reproducibility with coefficients of variation of ≤ 5 % for samples of ≥ 15 ppm peanut protein. The applicability of this method in the detection of peanut protein in various food commodities was demonstrated: two unlabelled products and two products which did not have peanut listed as an ingredient were identified as containing peanut protein. In all cases where peanut was listed, peanut protein could be determined. The results of RIE were always confirmed by those of a new cell-mediator-release assay that is based on a rat basophil leukaemia (RBL) cell-line (RBL-2H3), cells that are a functional equivalent to mucosal mast cells. Measuring the release of β-hexosaminidase resulting from cross-linking of basophil-bound peanut-specific immunoglobulin E, the assay mimics a main event of the allergic type-I reaction. The cell assay was adapted for food matrices and peanut could be detected down to 0.01% which additionally demonstrated in vitro that even trace amounts of peanut protein could elicit allergic reactions. Received: 28 May 1997 / Revised version: 11 July 1997