Analysis of the variability in the number of viable bacteria after mild heat treatment of food.

Variability in the numbers of bacteria remaining in saline solution and whole milk following mild heat treatment has been studied with Listeria innocua, Enterococcus faecalis, Salmonella enterica serovar Enteritidis, and Pseudomonas fluorescens. As expected, the most heat-resistant bacterium was E. faecalis, while P. fluorescens was the least heat resistant, and all bacteria showed greater thermal resistance in whole milk than in saline solution. Despite the differences in the inactivation kinetics of these bacteria in different media, the variability in the final number of bacteria was affected neither by the species nor by the heating substrate, but it did depend on the intensity of the heat treatment. The more severe the heat treatment was, the lower the average number of surviving bacteria but the greater the variability. Our results indicated that the inactivation times for the cells within a population are not identically distributed random variables and that, therefore, the population includes subpopulations of cells with different distributions for the heat resistance parameters. A linear relationship between the variability of the log of the final bacterial concentration and the logarithmic reduction in the size of the bacterial population was found.

Indirect measurement of the lag time distribution of single cells of Listeria innocua in food.

The distribution of log counts at a given time during the exponential growth phase of Listeria innocua measured in food samples inoculated with one cell each was applied to estimate the distribution of the single-cell lag times. Three replicate experiments in broth showed that the distribution of the log counts is a linear mapping of the distribution of the detection times measured by optical density. The detection time distribution reflects the lag time distribution but is shifted in time. The log count distribution was applied to estimate the distributions of the lag times in a liquid dairy product and in liver paté after different heat treatments. Two batches of ca. 100 samples of the dairy product were inoculated and heated at 55 degrees C for 45 min or at 62 degrees C for 2 min, and an unheated batch was incubated at 4 degrees C. The final concentration of surviving bacteria was ca. 1 cell per sample. The unheated cells showed the shortest lag times with the smallest variance. The mean and the variance of the lag times of the surviving cells at 62 degrees C were greater than those of the cells treated at 55 degrees C. Three batches of paté samples were heated at 55 degrees C for 25 min, 62 degrees C for 81 s, or 65 degrees C for 20 s. A control batch was inoculated but not heated. All paté samples were incubated at 15 degrees C. The distribution of the lag times of the cells heated at 55 degrees C was not significantly different from that of the unheated cells. However, at the higher temperatures, 62 degrees C and 65 degrees C, the lag duration was longer and its variance greater.

[Effectiveness of modified atmospheres against psychrotrophic pathogenic microorganisms in proteinaceous food]. / Eficacia de las atmósferas modificadas frente a los microorganismos patógenos psicrotrofos en alimentos proteicos.

Modified atmosphere packaging (MAP) of proteinaceous raw foods (meat, poultry and fish) extends their shelf-lives. It is well established that modified atmospheres (MA) inhibit the psychotropic aerobic Gram-negative bacteria, the main spoilage microflora of proteinaceous raw foods stored under refrigeration. Several researchers have warned about the possible growth of food poisoning microorganisms on them. Considering the minimal growth temperatures of pathogens, this review only deals with Aeromonas hydrophila, Clostridium botulinum, Listeria monocytogenes and Yersinia enterocolitica. C. botulinum produces its toxin in many different atmospheres, but it is unable to grow at temperatures below 3.3 degrees C, and its production rate of the toxin at temperatures below 4.5 degrees C is very low, to the extent that fish can be spoiled before the toxin is detected. Therefore, the control of the storage temperature of MAP fish seems to be indispensable to assure the absence of botulinal toxin. With regard to the other pathogens, vacuum is the atmosphere that may support more readily its growth; the higher the CO2 concentration in the atmosphere, the lower the growth rate is. Some investigations have shown that the growth rates of the psychotropic pathogens in MAP are lower than those of the spoilage flora. It has been shown also that A. hydrophila and L. monocytogenes growth rates are lower under MA than under aerobic storage. In relation to Y. enterocolitica, more investigations should be carried out in order to clear up its behaviour, because the available data in the literature are still confusing and sometimes even contradictory. In conclusion, there are no evidences that support the concern about MAP of proteinaceous raw foods representing a greater hazard than its conventional storage under air.

Effect of the addition of pronase E on the proteolysis in dry fermented sausages.

The effect of the addition of pronase E at two different concentrations on protein breakdown during the ripening of dry fermented sausage was studied. In all batches, water-soluble, non-protein and 5% phosphotungstic acid soluble nitrogens increased sharply during the first days of ripening, then became stabilized until the end of the process (26th day), and the total volatile nitrogen consistently increased during ripening. The greater the pronase E added the higher were the values reached for all these fractions. The changes in total free amino acids showed a similar pattern to that observed for the phosphotungstic acid soluble nitrogen. Histamine and tyramine progressively increased throughout the ripening. By sensory evaluation, no significant differences between the control batch and the batch with the lowest amount of added pronase E were found, but both batches were significantly different (P < 0·1) from the batch manufactured with the highest concentration of pronase E, which was classed as objectionable by the panellists because of its excessive softness.

Extracellular proteinase from Enterococcus faecalis subsp. liquefaciens. I. Growth and extracellular proteinase production under different culture conditions.

Growth and extracellular proteinase production by Enterococcus faecalis subsp. liquefaciens was studied on several culture media and under different incubation conditions. The organisms grew well and developed extracellular proteinase activity on proteinaceous media, but when it grew on Collins basal medium (lacking of protein), growth was poor and proteinase activity was not detected. The activation energy for growth was estimated to be 116 kJ/mol, the optimum being at 37 degrees C. Proteinase production was not affected by temperature in the range studied (7-45 degrees C). Growth rate was not affected by aeration although a higher amount of microorganisms was observed on shaking the culture during incubation. Likewise, extracellular proteolytic activity was about twice higher in cultures shaken at 2.3 or 3.3 Hz than in those shaken at 0 or 1.3 Hz.

Extracellular proteinase from Enterococcus faecalis subsp. liquefaciens. II. Partial purification and some technological important properties.

An extracellular proteinase from Enterococcus faecalis subsp. liquefaciens has been purified 780-fold by a method including gel filtration on Sephadex G-50 and affinity chromatography with gramicidin J as ligand. Approximately 15% of the original enzyme activity was recovered. A purification of 14,800-fold, with 11.4% yield, may be reached using chromatofocusing as final step in the purification procedure. The molar mass of the enzyme has been estimated to be approximately 30 kDa by Sephadex gel filtration and approximately 26 kDa by SDS-PAGE. The isoelectric point has been found to be 4.6. Maximum enzyme activity of the proteinase has been observed at pH 7.5 and 45 degrees C. The enzyme hydrolyzed bovine serum albumin, alpha-lactoalbumin, beta-lactoglobulin, casein and pork myofibrillar and sarcoplasmic proteins. The extracellular proteinase was very stable; the enzyme maintained its activity in cell-free extracts over a very wide range of temperatures (-25 to 37 degrees C) for at least 2 months. At 12 degrees C, it was stable in the pH range of 5.5 to 8.0.

Study of proteolysis during the processing of a dry fermented pork sausage.

Dry pork sausage was formulated, fermented and dried for 41 days. Gross composition was determined and proteolysis was monitored by assaying the following N fractions: water-soluble nitrogen (WSN), salt-soluble nitrogen (SSN), ultrafiltration (UF) permeates of WSN and SSN, phosphotungstic acid (PTA)-soluble and -insoluble fractions and free amino acids. Meat and sausages, water- and salt-insoluble components, UF-retentates of WSN and SSN were assessed by SDS-PAGE at different stages of ripening. The amount of WSN, WSN permeate, PTA-soluble N and free amino acids increased during processing, while the SSN and PTA-insoluble N decreased. The electrophoretic studies demonstrated that proteolysis of the heavy myosin chain, α-actinin and actin was most prominent. The increased insolubility of meat proteins was confirmed by SDS-PAGE.