Effect of high pressure on structural modifications and enzymatic activity of a purified X-prolyl dipeptidyl aminopeptidase from Streptococcus thermophilus.

PepX aminopeptidase from Streptococcus thermophilus ACA DC 0022, used in Greek Feta cheese manufacturing, was purified. PepX comprises two subunits of equal molecular mass estimated, using SDS-PAGE and native-PAGE electrophoresis, to be 86 kDa. The effects of high pressure processing (100-450 MPa, combined with 20-40 °C) on purified PepX activity and structure were studied. Activation of the enzyme was observed after processing at 100-200 MPa and 20-30 °C. More intense processing conditions led to enzyme inactivation. PepX HP-induced conformational changes were also investigated through application of Circular Dichroism spectroscopy (CD). Pressures up to 200 MPa resulted in a structurally molten globule-like state where PepX maintained its secondary structure but the tertiary structure was substantially affected and enzyme activity increased. Both secondary and tertiary structures were affected severely by higher pressures (450 MPa), which reduced enzyme activity.

Shelf-life extension of gilthead seabream fillets by osmotic treatment and antimicrobial agents.

AIMS: The objectives of the study were to evaluate the effect of selected antimicrobial agents on the shelf life of osmotically pretreated gilthead seabream and to establish reliable kinetic equations for shelf-life determination validated in dynamic conditions. METHODS AND RESULTS: Fresh gilthead seabream (Sparus aurata) fillets were osmotically treated with 50% high dextrose equivalent maltodextrin (HDM, DE 47) plus 5% NaCl and 0·5% carvacrol, 0·5% glucono-δ-lactone or 1% Citrox (commercial antimicrobial mix). Untreated and treated slices were aerobically packed and stored isothermally (0-15°C). Microbial growth and quality-related chemical indices were modelled as functions of temperature. Models were validated at dynamic storage conditions. Osmotic pretreatment with the use of antimicrobials led to significant shelf-life extension of fillets, in terms of microbial growth and organoleptic deterioration. CONCLUSIONS: The shelf life was 7 days for control samples at 5°C. The osmotic pretreatment with carvacrol, glucono-δ-lactone and Citrox allowed for shelf-life extension by 8, 10 and 5 days at 5°C, respectively. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of the study show the potential of adding carvacrol, glucono-δ-lactone or Citrox in the osmotic solution to extend the shelf life and improve commercial value of chilled osmotically pretreated fish products. The developed models can be a reliable tool for predicting the shelf life of fresh or minimally processed gilthead seabream fillets in the real chill chain.

Effect of non-thermal processing by High Hydrostatic Pressure on the survival of probiotic microorganisms: study on Bifidobacteria spp.

High Hydrostatic Pressure (HP) processing has been suggested as an alternative method to improve textural attributes of dairy products. Since, the global market seeks improved functional foods, it is important to investigate whether HP processing can be applied to fermented dairy probiotic products. The inactivation kinetics of Bifidobacterium spp. in a model system of acid pH value under high pressure (100-400 MPa) combined with moderate temperature (20-35°C) was investigated. Bifidobacterium spp. inactivation followed first order kinetics at all pressure-temperature combinations used. Pressure and temperature were found to act synergistically on the viability loss of the bacterium. The corresponding z(T) and z(P) values of inactivation were also estimated and, values of 41.5°C and 93.5 MPa at reference pressure of 200 MPa and reference temperature of 25°C were estimated, respectively. HP treatment of 200 MPa at 20-25°C for 10-15 min, recommended for textural modification, is not detrimental to the viability of the studied probiotic culture and would be suitable for respective fermented probiotic products.

Kinetic study of the combined effect of high hydrostatic pressure and temperature on the activity of Lactobacillus delbrueckii ssp. bulgaricus aminopeptidases.

The effect of high hydrostatic pressure (HHP) (100 to 700 MPa) combined with temperature (20 to 40 degrees C) on the activity of 5 aminopeptidases (PepN, PepX, PepY, PepC, and PepA) of Lactobacillus delbrueckii ssp. bulgaricus ACA-DC 0105, used as starter culture for feta cheese production, was studied. Aminopeptidases PepN, PepX, and PepA were activated at pressures up to 200 MPa, at temperatures up to 40 degrees C. PepY and PepC appeared to be more sensitive to pressure and temperature treatment leading to inactivation for pressures above 100 and 200 MPa, respectively, combined with temperature above 30 degrees C. A multi-parameter equation was used for predicting the activation of PepN, PepX, and PepA aminopeptidases in the pressure and temperature domain. Overall, processing at 200 MPa and 20 degrees C may be selected as the optimum conditions for maximum activation of 4 out of 5 aminopeptidases of L. delbrueckii ssp. bulgaricus. A 20-min treatment at these conditions leads to an average 3-fold increase in activity, which could lead to better and faster maturation of white cheese.

Field evaluation of the application of time temperature integrators for monitoring fish quality in the chill chain.

The applicability of time temperature integrators (TTI) as effective tools of chill chain monitoring was assessed. Validated kinetic models of pseudomonads growth of Mediterranean, marine-cultured chilled gilt-head seabream (Sparus aurata) and full knowledge of the response of suitable enzymatic TTI are the basis of the TTI application algorithm. This scheme was evaluated through a controlled field test of exported fish, from harvest to final consumption. Response of TTI attached on different locations of packages was compared to actual temperature recording. Data that could not be obtained during the actual field test, such as microbiological or sensory tests of fish at intermediate points of the chain, were measured in a replicate laboratory study, simulating the handling of products and the real time-temperature profiles of the field test. The conducted field tests showed the applicability and usefulness of TTI monitoring of the fish chill chain, elucidating also the practical difficulties and limitations, that need to be addressed for expanding TTI use as a reliable management tool.

Development of a Safety Monitoring and Assurance System for chilled food products.

The principles of a novel chill chain management policy, coded Safety Monitoring and Assurance System (SMAS) for the optimisation of the distribution of chilled food products within the chill chain are developed. In this system, a new approach based on actual risk evaluation at important points of the chill chain is used in order to promote products to the next stage of distribution. This evaluation based on product's time-temperature history, variation in product's characteristics (e.g. a(w), pH, etc.), and the use of predictive models for the growth of food pathogens, allows to give priority to products in such a way that risk at consumption time is minimized. The effectiveness of SMAS was evaluated against the First In First Out (FIFO) approach, the current method for food distribution, in a case study on the risk of listeriosis of cooked ham using the Monte Carlo simulation technique. Furthermore, the two approaches were compared for their effect on the quality of the products in terms of remaining shelf life at the time of consumption. The results showed that following the SMAS approach the risk of listerisosis is significantly lower while the spoiled products at the time of consumption are significantly reduced compared to FIFO approach.

Application of shelf life decision system (SLDS) to marine cultured fish quality.

Growth of natural microflora of marine cultured, air-packed, sea bass (Dichentrachus labrax) was studied at isothermal conditions in the 0-15 degrees C range and kinetically modelled using the four-parameter Logistic equation. Sensory shelf life was correlated to pseudomonad population and sensory acceptability was correlated to a pseudomonad level, Ns, of 10(7). The variability of their initial population was quantitatively shown and a conductance-based rapid method specific to sea bass pseudomonad enumeration was established as a practical means of N0 determination, required in shelf life predictions. Kinetic models, shelf life correlations and N0 data were incorporated into the shelf life decision system (SLDS) shown to be an effective tool for marine cultured sea bass chill chain management leading to optimization of quality of the fish at consumer's end.

Development and assessment of an intelligent shelf life decision system for quality optimization of the food chill chain.

The principles of application of a Shelf Life Decision System (SLDS) for the optimization of the distribution of chilled fresh and minimally processed food products are developed. The SLDS integrates predictive kinetic models of food spoilage, data on initial quality from rapid techniques, and the capacity to continuously monitor temperature history of the food product with Time Temperature Integrators (TTIs) into an effective chill chain management tool that leads to an improved narrow distribution of quality at consumption time, effectively reducing the probability of products consumed past shelf life end. The applicability and effectiveness of the SLDS is demonstrated and evaluated based on actual food spoilage and TTI kinetics and chill chain data employing the Monte Carlo simulation method.

Applicability of an Arrhenius model for the combined effect of temperature and CO(2) packaging on the spoilage microflora of fish.

The temperature behavior of the natural microflora on the Mediterranean fish red mullet (Mullus barbatus) was examined as a case study. The growth of the spoilage bacteria Pseudomonas spp., Shewanella putrefaciens, Brochothrix thermosphacta, and lactic acid bacteria was modeled as a function of temperature and the concentration of carbon dioxide in modified atmosphere packaging. Combined models were developed and comparatively assessed based on polynomial, Belehradek, and Arrhenius equations. The activation energy parameter of the Arrhenius model, E(A), was independent of the packaging atmosphere and ranged from 75 to 85 kJ/mol for the different bacteria, whereas the preexponential constant decreased exponentially with the packaging CO(2) concentration. We evaluated the applicability of the models developed by using experimental bacterial growth rates obtained from 42 independent experiments performed with three Mediterranean fish species and growth rates predicted from the models under the same temperature and packaging conditions. The accuracy factor and bias factor were used as statistical tools for evaluation, and the developed Arrhenius model and the Belehradek model were judged satisfactory overall.

Use of time-temperature integrators and predictive modelling for shelf life control of chilled fish under dynamic storage conditions.

A systematic approach for fish shelf life modelling and Time Temperature Integrator (TTI) selection in order to plan and apply an effective quality monitoring scheme for the fish chill chain was developed. The temperature behaviour of the natural microflora of the Mediterranean fish boque (Boops boops) was studied and growth of the specific spoilage bacteria Pseudomonas spp. and Shewanella putrefaciens was modelled and correlated to organoleptic shelf life. Arrhenius and square root functions were used to model temperature dependence of maximum growth rates. Bacterial growth and shelf life models were validated under dynamic storage conditions with independent variable temperature experiments. The response of several TTIs from similar storage experiments was also modelled. The reliability of the TTI monitoring was cumulatively expressed by the error in the TTI derived effective temperature (Teff) for different variable temperature distributions. Teff was directly translated to shelf life of the fish.

Modelling the effectiveness of a natural antimicrobial on Salmonella enteritidis as a function of concentration, temperature and pH, using conductance measurements.

The growth of Salmonella enteritidis in a brain heart infusion medium was monitored using the traditional viable count method and by conductance measurements using a Rabit impedance instrument. Growth curves (log10 cfu ml-1 vs time) at three different concentrations of oleuropein (0, 0.2 and 0.8%), pH values in the range of 5-8 and incubation temperatures from 22 to 42 degrees C were modelled using the Gompertz equation. A good correlation between the maximum growth rate from the viable count method and the maximum slope of the conductance curve from the impedance instrument was established. Based on this correlation, the maximum specific growth rate of Salm. enteritidis was modelled as a function of the oleuropein concentration, initial pH values and the incubation temperature with a quadratic equation, using a new, large dataset of growth measurements by conductance. The developed model was validated by statistical comparison of predicted growth rates with growth rates determined by the viable count method, within the limits of the antimicrobial, pH and temperature domain.