No description available.

The aim of this work was to evaluate the use of enzymes to accelerate the ripening of Cheddar cheese. Addition of preparations of β-D-Galactosidase to the cheese Milk. The ripening of Cheddar cheese produced from lactose-hydrolysed milk (e.g. up to 60% hydrolysis of the lactose) , was only slightly accelerated even though one of the β-D-galactosidase enzymes used contained some proteolytic enzyme(s). The numbers of starter bacteria at the end of the cheesemaking process were higher in cheese made from enzyme-treated milk than from untreated control milk. The level of soluble nitrogen gradually increases in Cheddar cheese as it becomes older. In these studies higher values for soluble nitrogen were observed in cheese made from lactose-hydrolysed milk compared with control cheese made from untreated milk after similar periods of ripening. This was true for both commercial lactase enzyme preparations - one of which was highly purified while the other contained substantial amounts of protease. Cheddar cheese manufactured from the latter enzyme contained the highest level of soluble nitrogen throughout the ripening period and this could be associated with appreciable acceleration of the ripening of the cheese. More extensive hydrolysis of the casein fractions was also evident in 6-month old Cheddar cheese made from milk treated with the lactase containing protease compared with the control and also the cheese made from milk treated with highly purified lactase. Other desirable effects were achieved as the result of lactose hydrolysis of the cheese milk: (i) Reduction in the cheesemaking time. (ii) Greater judge preference for cheese manufactured from lactose-hydrolysed milk. This effect could be due to the slight increase in protein degradation of this cheese. (iii) The increased level of glucose and galactose in the whey could be a desirable feature for further processing, i.e. in the production of a sweet syrup. Addition of a commercial brand of neutral proteinase to the cheese cord Cheddar cheese was manufactured by using direct-to-vat inoculation of concentrated frozen mixed strains of mesophilic starter culture. In preliminary experiments the enzyme was provided by the manufacturer as a powdered chemical and added to the milled curd at a rate of 0.001, 0.002, 0.005 and 0.01% (w/w). Separate lots of cheese were ripened at 10° and 13°C, and the enzyme activity in the cheese as assessed by monitoring the level of soluble nitrogen, hydrolysis of casein and organoleptically. The extent of acceleration of Cheddar cheese ripening depended on the level of enzyme added to the curd; for example, experimental cheese to which 0.001 and 0.01% enzyme had been added, had characteristics at 2 months similar to those of the control cheese at 4 and 8 months respectively. In enzyme-treated cheese there was a greater liberation of the more soluble nitrogenous compounds, and gel electrophoresis showed a high reduction in β- and αs1 -caseins compared with the control. All the enzyme-treated cheese had defects in "body and texture" characteristics, and had mottled, weak body and bitter flavour. The extent of the defect was associated with the enzyme level. The changes brought about by the addition of the enzyme did not increase to any large extent after four to six months ripening. The effect of the higher ripening temperature was enhanced enzyme activity. Follow up near-commercial scale trials the neutral proteinase was supplied by the manufacturer in the form of a coating on salt to enhance homogeneity of mixing with the curd. The addition of Neutrase, 0.002, 0.003 and 0.005% (w/w) to the cheese curd increased the proteolytic activity, i.e. greater liberation of more soluble nitrogen and more extensive casein hydrolysis compared with the control made from untreated curd. The flavour intensity of cheese made from enzyme-treated curd was greater than from untreated curd but the following quality problems were observed: - bitter and unacceptable flavour(s); - open and crumbly texture; - brittle and softer body cheese; - discoloured or mottled. The extent of these defects was related to the amount of enzyme added. In order to overcome these defects in the experimental cheese, the enzymatic activity had to be stopped when the desirable flavour intensity in the cheese had been achieved. Preliminary experiments have indicated that the use of Neutrase- treated curd which results in more rapid ripening and development of cheese flavour may have advantages in the production of processed cheese.

No description available.

No description available.

Adenosine triphosphate (ATP) assay and electrical measuring techniques were assessed for their ability to produce an estimate of total colony count. ATP assay could be used for estimating the microbial flora in foods, provided that non-microbial ATP was removed from the sample first. Linear relationships between log[10] microbial ATP content and log[10] colony count were obtained. Electrical measuring techniques provided an inverse linear relationship between electrical detection time and log[10] colony count. The capacitance element of the impedance signal was found to be superior to the conductance element for the estimation of pure cultures and the flora of two foods, since faster detection times and higher correlations with colony count were obtained. Capacitance detection times at elevated temperatures (35° and 25°C) provided a good estimate of psychrotrophic colony count at 15°C. The use of electrical measuring techniques was also investigated for the rapid detection of salmonellae. Most salmonellae tested could be detected electrically in pure culture by the production of a specific signal from the fermentation of dulcitol or by an inhibition of this signal in the presence of a Salmonella-specific bacteriophage. However, when these two tests were used for artificially contaminated foods, an unacceptably high number of false-negative results was obtained. Therefore, further tests were investigated for their potential in the electrical detection of salmonellae. These were: effect of antiserum, lysine decarboxylation and hydrogen sulphide production. The final section of this thesis was concerned with the use of conventional and rapid microbiological techniques in the analysis of bacterial growth/temperature relationships. A Square Root plot (√r = b(T-T[o])) was found to be preferable to the Arrhenius plot (k = Ae[-muu/RT]) in describing the growth of psychrotrophic food spoilage bacteria at chill temperatures. The Square Root plot was also applicable to mixtures of organisms and to ATP and capacitance detection time data. The relevance of such work to the investigation of food spoilage was discussed.

Microbial thermal inactivation survivor curves (log10 numbers plotted against time) have long been described as maintaining a strictly linear rate of decline. However, much evidence exists which suggests deviation from log-linear kinetics does occur, and that this is not purely the result of experimental procedure as contended by some authors. Here, the shape of inactivation kinetics in Salmonella enteritidis was investigated. A heat challenge method was developed which, as far as could be ascertained, was free from methodological artefacts influencing the shape of survivor curves. High initial cell densities allied to sensitive enumeration resulted in biphasic survivor curves at 60°C. Tailing survivors accounted for approximately 1 in 105 of the initial population and possessed roughly four times the heat resistance. At temperatures 50 to 65°C, the presence of tailing prevented the use of D-values to accurately describe death rates. However, describing survivor curves using a log-logistic model increased data-fit at all temperatures investigated. The biphasic nature of survivor curves was studied closely between 49 and 60°C. It was observed that the extent of tailing was temperature-dependent; as temperature decreased, linearity increased such that at 51°C, survivor curves had no tailing. Studies using S. typhimurium and S. senftenberg 775W revealed similar kinetics. In these salmonellas, survivor curves demonstrated linearity at 54 and 57°C, respectively. The influences of culture age and growth rate on the shape of 60°C-inactivation curves were also investigated. Batch-cultured S. enteritidis cells of various maturities gave rise to survivor curves of differing heat sensitivities. Exponentially growing cells were shown to be the most heat sensitive, while late-stationary phase cells were the only populations to result in non-tailed survivor curves. Carbon-limited continuously cultivated cells demonstrated similar biphasic inactivation kinetics. Predictably, the slowest dilution rate corresponded to the greatest heat resistance. Starved cells produced linear inactivation kinetics that were virtually identical to those of late-stationary phase batch-cultured cells. That tailing in batch cultures was similar to chemostat populations, indicated that possible differences in growth rates in batch-cultured cells could not account for tailing. Furthermore, growth was necessary for tailing to be observed. Investigations into the cause of tailing revealed that these cells were not genetically distinct from the majority population. Instead, it is believed that tailing cells arise following the expression of heat-shock proteins during heating. Partial inhibition of de novo protein synthesis during heating resulted in much reduced levels of tailing. It is proposed that the temperature of inactivation determines the proportion of cells capable of expressing a heat-shock response, such that the temperature at which linearity is achieved corresponds to the point at which all cells are fully heat-shock protected.

Pretreatment of stainless steel (AISI 304 & AISI 316) surfaces with skimmed UHT milk significantly reduced the numbers of attaching Staphylococcus aureus and other Gram positive bacteria, although the same effect was not observed with the Gram negative Pseudomonas fragi and Escherichia coli. Milk constituents with molecular weights below 1000 Daltons did not show the observed effect. Droplet contact angle experiments showed that the stainless steel surface became only marginally more hydrophobic when milk was adsorbed. The attachment of S. aureus onto stainless steel was found to be dependent on the physico-chemical properties of the bacterial surface, the stainless steel surface and of the suspending solution. SDS-polyacrylamide gel electrophoresis showed that proteins are adsorbed onto the stainless steel surface in approximately the same proportions as those in the bulk milk. Of the milk proteins, the caseins were more effective at inhibiting attachment. This was found to be true even when the proteins were used at identical concentrations, rather than in the proportions found naturally in milk, k-Casein was the most effective at inhibiting attachment.

The lactic acid fermentation of cheese whey permeate by Lactobacillus helveticus was studied. Precipitate formation during autoclaving of whey permeate was examined. Precipitation was found to be pH and temperature dependent. Qualitative analysis suggested that the precipitate was a calcium-phosphate complex. Solubilisation was achieved both by acidification and use of the sequestering agent EDTA. Optimisation of L. helveticus growth in whey permeate was carried out using factorial design, as opposed to a traditional univariate approach. Using this technique, the variation of specific growth rate with pH, temperature and stiirer speed was assessed. Cell growth and lactic acid formation in whey permeate containing various supplements, were investigated. Yeast extract was the most effective nitrogen/growth factor supplement. Maximum lactic acid production was achieved in permeate containing yeast extract (0.75% w/v), Tween 80 (0.1% v/v) and sodium acetate (0.05% w/v). Optimisation of lactic acid production in supplemented whey permeate was performed using factorial design. Optimum conditions for both acid formation and cell growth were pH 5.9, temperature 42°C and stirrer speed 200 rpm. Fourier transform infrared spectroscopy was applied to the on line and off line quantitative analysis of lactose and lactic acid during the fermentation process. This technique enabled substrate and product levels to be assessed quickly and simply, with no sample pre-treatment. Continuous culture of L. helveticus in MRS medium and supplemented whey permeate was carried out. Substrate conversion and lactic acid productivity decreased with increasing dilution rate. Maximum productivity corresponded to a dilution rate of 0.3 h" 1, whereas minimum residual substrate occured at a dilution rate of 0.1 h' 1 . Translation of the fermentation process from bench scale (11) to pilot scale (161) appeared to be successful. Completion times, productivity and lactose utilisation compared favourably with bench scale results.

Chiroptical, rheological and thermodynamic studies have been undertaken to investigate temperature-induced changes in the ý molecular organisation of gelatin. From the results obtained, a unified model for gelation and melting has been developed, and tested using Monte Carlo computer simulation. The temperature at which gelatin gels are formed has a major influence on the properties of the resulting network, with higher curing temperatures conferring increased thermal stability. In particular, gels formed by sequential curing at two different temperatures show biphasic melting. This is explained in terms of a temperature-dependence of helix length within the junction zones of the gel, and quantified by considering end-effects in the thermodynamics of helix stability. Measurements of 'initial slope' kinetics, performed over a broad concentration range, showed first-order kinetics at low gelatin concentrations, while at higher concentrations a second-order process was also evident. The results are interpreted as triple-helix nucleation at metastable 'hairpin turns' in one chain (bringing two chain segments into close proximity) together with a third strand from either the same chain (first order) or a different chain (second order). From simple geometric considerations, the maximum length of intermolecular helices ( which contribute to the gel network) is greater than that of twasted 9 intramolecular structures, giving a qualitative explanation of the increased strength of gels formed by precuring at higher temperatures (where only long helices are stable) over those quenched directly to low temperature. Monte Carlo simulation incorporating an initial assumption that helix propagation is rapid and proceeds to geometric limits gave unrealistic helix lengths and simulated melting profiles, and was replaced by the assumption that cis-trans isomerisation of peptide bonds is the controlling factor in helix propagation. Using the latter assumption, most aspects of the observed behaviour were successfully reproduced using program variables set within realistic ranges or, where possible, fixed at experimentally-determined values. In particularg the co-operativity of the simulated melting process was critically dependent on the value of a parameter x (the number of triplet units within each helix incapable of participating in bonding, due to end-effects), with a value of x=1 giving the best fits with experiment (consistent with accepted bonding patterns for the collagen triple helix). Other key parameters were the midpoint temperature for melting of the parent collagen, which gave best agreement when set at 37-38"C, and t6e proportion of cis peptide residues present in disordered gelatin chains, with an optimum lower limit of 0.15. Using these values, the simulation reproduced, with excellent precision, the helix fraction and melting profile of gels formed over a wide range of quench temperatures, and gave an acceptable approximation to the form of reaction progress curves obtained for helix formation. The biphasic melting of samples held at intermediate temperature before final quenching was also modelled realistically.

No description available.