In many countries of the world and in some parts of the United States milk is produced which has a high bacterial contamination. Such milk of undesirable quality is frequently delivered to factories engaged in the manufacture of cheddar cheese. This milk commonly contains large numbers of lactic acid-producing bacteria or other types of microorganisms which cause objectionable flavors and textural defects in the cheese. The improvement of the quality of milk supply under some conditions is a matter of great difficulty so that the manufacture of inferior quality milk into cheese is a problem often encountered. In the United States pasteurization of milk is used to reduce the bacterial content and give the cheese maker control over the manufacturing process. Public health officials favor pasteurization as a protection against pathogens; however, in many areas of the world pasteurization is not available. Although pasteurization of milk for cheddar cheese offers certain advantages such as destruction of pathogenic bacteria which may be present, and control of certain undesirable fermentations, experience has shown that pasteurized milk cheese develops flavor slowly and, even with extended ripening, does not have as satisfactory a flavor as good raw milk cheese. The slow ripening usually is attributed to the destruction by heat of certain essential bacteria and enzymes normally present in milk. Pasteurization, however, destroys many enzymes indigenous to milk as well as some beneficial organisms; consequently, cheese made from pasteurized milk ripens more slowly than cheese made from raw milk. For years, leading dairy technologists have been laboring assiduously but quite unsucessfully to produce cheese free from undesirable organisms yet comparable in flavor and in the rapidity of ripening to the best quality of raw milk cheese. Pursuant to these objectives a number of methods such as replenishing the enzymes in milk destroyed by pasteurization, the use of select ripening cultures, and the use of mixtures of various percentages of raw and pasteurized milk have been tried but without complete success. These objectionable features of pasteurization led to interest in another method such as the treatment of milk with edible hydrogen peroxide to control fermentation by means of its germicidal and inhibitory action. This comparative study was conducted to determine the effect of the germicidal properties of hydrogen peroxide in treating raw milk for cheddar cheese making in relation to the flora, quality, and ripening of the cheese. This study was concerned with the remedial measures which can be applied to milk to overcome some defects in the cheese. The antiseptic and germicidal properties of hydrogen peroxide are well known. A study involving the use of hydrogen peroxide and catalase has many possibilities in the dairy industry, and the practical aspects of this problem are numerous. Some phases are herewith indicated: 1. If hydrogen peroxide could be used to improve the general quality of cheddar cheese, it would be a boon to the industry and should have a value in the manufacture of cheddar cheese for shelf curing purposes, canning, processing, and for natural ripening in transparent packages. 2. It was believed that the use of hydrogen peroxide and catalase would increase the safety of raw milk cheese. (Kernsman, 1934, found that 0.1 percent of hydrogen peroxide killed E. coli, E. typhi and staphilococcus.) 3. If hydrogen peroxide could be used for destroying organisms harmful in milk and thus for preventing undesirable fermentation, yet leave intact more of the natural enzymes than is possible in accepted pasteurization procedures, the cheese treated with hydrogen peroxide and catalase might ripen faster than pasteurized-milk cheese and have a finer and more pronounced flavor. 4. If approved by public health authorities in the United States, treating milk with hydrogen peroxide would be a simple method of reducing bacterial content in small communities and rural areas. Such procedure would be very practical in preventing growth of bacteria in milk produced under unsanitary conditions. 5. If the use of hydrogen peroxide could be proved practicable, a beneficial program in most countries and especially in the Middle East where dairy equipment and pasteurizers are not readily available and where the production of unsanitary milk predominates might be established. 6. Since this process does not require special equipment it might prove economical and might become, in the future, a useful method of reducing the bacterial content of milk and preserving some of the natural characteristics of the raw milk for cheese making.

Residual rennin in cheese whey and curd was measured by using a special sensitive substrate. The substrate was made by reconstituting 6 g NDM in 500 ml of buffer containing 0.2 M CaCl2, 0.5 M cacodylic acid and 0.2 M triethanolamine at pH 5.7. Cheese curd was blended into a 1:7 slurry (1 part curd, 7 parts water), and 1.67% sodium chloride was added to the why and slurry to liberate residual rennin from casein. The residual rennin in cheese whey and slurry were determined simultaneously with an identical sample containing known rennin activity. Samples with known activity were prepared by destroying the residual rennin in unknown samples after which a known amount of rennin was added back to a standard. The examine the effectiveness of this method for measuring rennin activity in whey or slurry, a recovery test was developed to measure rennin activities in the whey and curd made by centrifuging rennet-coagulated milk. The average total recovery from 15 replications was 101.6 ± 2.4%. It was found that pH was a main factor affecting rennin distribution between whey and curd. The amount in the curd increased with decreasing pH at setting. Adding 0.02% CaCl2 to milk was of little effect. The results showed that 68.1 6 ± 6.6% of the residual rennin was found in Cheddar cheese whey after dipping and 17.2 6 ± 2.6% of the residual rennin was in the curd after milling.

Cheddar Cheese samples (good cheese, weak cheese, cheese made with pasteurized milk, cheese made with heat-shocked milk, cheese from production plant A, cheese from production plant B, cheese made with adjunct culture, and cheese made without adjunct culture), were evaluated during the ripening stage. Proteolysis was studied by a fractionation scheme, resulting in an insoluble fraction analyzed by urea polyacrylamide gel electrophoresis (Urea-PAGE), and a soluble fraction which was further investigated through water soluble nitrogen (WSN), trichloroacetic acid soluble nitrogen (TCA-SN) and phosphotungstic acid soluble nitrogen (PTA-SN) analyzed by total Kjeldahl nitrogen content (TKN). Reversed phase high performance liquid chromatography (RP-HPLC) was used to study the peptide profile of the water soluble fraction. Lipolyisis was studied by levels of individual free fatty acids determined through gas chromatography-flame ionization detection (GC-FID) after isolation employing solid phase extraction (SPE). Volatile sulfur compounds were studied using head space solid phase micro-extraction (SPME) coupled with gas chromatography-pulsed flame photometric detection (PFPD). It was found that Urea-PAGE is capable to differentiate samples according their age, but cannot discriminate samples regarding the treatment assessed, quality or origin of the samples. However, measurements of total Kjeldahl Nitrogen (TKN) of the WSN, TCA-SN, and PTA-SN fractions, and the principal component analysis of the RP-HPLC peptide profile of the WSN fraction, revealed differences in the rate and pattern of proteolysis for each one of the manufacturing cases. Good cheese, cheese produce in plant TCCA, cheese made in plant CRP with adjunct culture isolated from plant TCCA cheese, and cheese made with heat-shocked milk developed higher level of total nitrogen for the WSN, TCA-SN and PTA-SN fractions, indicating that primary and secondary proteolysis were faster for these samples. This is supported by a PCA model with three principal components that account for the 80-83% of the variability of the data from the RP-HPLC peptide profile analysis, which discriminates the samples according to age and manufacturing practice. In addition, FFA profiles demonstrated higher levels of low and medium chain free fatty acids for good cheese, cheese produce in plant TCCA, cheese made in plant CRP with adjunct culture, and cheese made with heat-shocked milk samples, which suggest faster lipolysis during ripening. The Volatile Sulfur Compounds (VSC) analysis showed higher levels of DMS and MeSH and lower levels of H2S, suggesting faster catabolism of sulfur containing amino acids in good cheese, cheese produce in plant TCCA, cheese made in plant CRP with adjunct culture, and cheese made with heat-shocked milk. / Graduation date: 2013

Thesis (D. Tech.) -- Central University of Technology, Free State, 2010 / Branded dairy products have lately become a global trend. As a result of this, the origin of the milk used in the manufacturing of branded cheeses must be declared by the producer, since it is known that these products are highly adulterated with foreign milk. In South Africa, branded Ayrshire Cheddar cheese has become highly popular due to its unique organoleptic properties and in light of claims that it ripens much faster than cheese made from other milk (not including Ayrshire). This study was therefore directed to investigate the unique properties of branded Ayrshire Cheddar cheese versus Cheddar cheese manufactured from a mixture of other breeds’ milk (not including Ayrshire milk) and to establish a catabolite profile for each cheese type. The outlay of the thesis was constructed into six chapters each with its own outcomes. The first chapter focused on the variations between the two Cheddar cheese batches (produced from Ayrshire and other breeds’ milk) with regards to organic acid, selected chemical parameters and starter microbiotia. In the following three chapters mathematical models were developed that would predict organic-; fatty and amino acid fluxtuations respectively in the cheese made from Ayrshire and other milk. In the last chapter two artificial neural networks were designed with the two starter organisms, Lactococcus lactis and Streptococcus thermophilus as variable indicator respectively. Thirty-two cheese samples of each batch (pure Ayrshire (4) / mix breed with no Ayrshire (4)) were ripened and samples were analysed under the same conditions on the following days after production: 2, 10, 22, 36, 50, 64, 78, and 92. In the subsequent chapters, the following analysis were done on each day of analysis: organic acid by means of high performance liquid chromatography (HPLC); fatty acids by means of Gas Chromatography Mass Spectometry (GCMS); amino acids by means of GC-MS; microbial analysis by means of traditional methods, total DNA extraction and polymerase chain reaction (PCR); and standard chemical analysis for moisture, NaCl and pH. In the first research chapter, the minimum and maximum (min/max) values, standard deviations and proposed rel X values of organic acids were evaluated in Ayrshire and the mixed-breed Cheddar cheese, and showed that isovaleric acid is the organic acid with the least variation relative to concentration in both cheeses and it was assumed that this organic acid is the most effective indicator of cheese uniformity. Clear differences in organic acids, chemical variables and starter micro-organisms were also evident in the two cheese batches. Results obtained from the regression models which was defined for each organic -; amino - and fatty acid by means of mathematical equations can be used by the manufacturer to achieve i.e. the selection of cheese for specialist lines, the early exclusion of defective cheeses, and the establishment of brand origin (Ayrshire vs. mixed-breed Cheddar cheeses). The regression graphs also illustrate unique flux patterns in Ayrshire and the mixed-breed in terms of organic -, fatty -, and amino acid content. In the last chapter, the discrimination between the two batches was respectively done via artificial neural network (ANN) modelling of Lactococcus lactis and Streptococcus thermophilus as indicator organisms. The ANN consisted of a multilayered network with supervised training arranged into an ordered hierarchy of layers, in which connections were allowed only between nodes in immediately adjacent layers. The construction thereof allowed for two output nodes, connected to an input layer consisting of two nodes to which the inputs were connected. In both cheeses the results from the ANN showed acceptable classification of the cheeses based on the counts of L. lactis and S. thermophilus.

Changes in cheese physical properties during aging are related to proteolysis by coagulant type, culture enzymes, and non-starter lactic acid bacteria (NSLAB). Storage temperature also affects aging rate. Cultures are important for flavor development , but less is understood about their role in melting and textural properties. Our objective was to make Cheddar cheese using different cultures, to age it at 6 and 13°C, and measure physical and proteolytic properties over 12 mo to determine whether changes in texture and melting correlated with the extent of proteolysis that occurred during aging. Cheese was manufactured using Lactococcus lactis starter culture either alone or combined with one or both of Lac Lc. Lactis or Lactobacillus helveticus adjunct cultures . Three replicates of cheese were made using 1500 lb of milk. Cheese composition was 35.5 ± 1.0% moisture, 52.5 ± 2.5% FDB, 1.65 ± 0.05% salt, and pH 5.2 ± 0.1. All cheeses were initially stored at 6°C, then half moved to l3°C after 21 d. Texture profile analysis was performed using 25% and 60% compression and melting measured using a Meltmeter at 65°C. The data were analyzed based on culture and temperature over 12-mo storage time. The overall hardness decreased, while the cohesiveness decreased for all treatments. Extent of melting was significantly correlated with hardness (r = 0.62), cohesiveness (r = 0.40), and inversely with adhesiveness (r = 0.24). Correlations with adhesiveness and cohesiveness were not linear. Proteins were extracted from cheese at 1 wk, 1, 2, 4, 6, 9, and 12 mo of aging using 500 mM sodium citrate solution containing 1% sodium chloride. Purified extracts were then applied to a high-performance liquid chromatography CS reverse phase column and large hydrophobic peptides and protein peaks monitored at 214 nm. Melting was inversely correlated with the amount of intact ɑs1-caserienm remaining in the cheese (r = -0.54) and directly correlated with what was thought to be ɑs1-casein (f 24 - 199) (r = 0.56).

This study was conducted to evaluate several forms of equations for predicting Cheddar cheese yields based on the fat and protein content of milk and moisture content of cheese. Production and quality control data from a Cheddar cheese plant for one entire year was used. This included the pounds of milk that went into each vat of cheese, yield of cheese from each vat, cheese moisture from each vat, and fat and protein percentages of the milk. Seven models were derived to predict the yield of Cheddar cheese. The seven models were statistically fitted to the data by applying the Marquardt non-linear least squares method of iteration. These were compared with the commonly used Van Slyke and Price formula, with casein estimated as a percentage of total protein. The differences among the eight models were small.

The use of slower acid-producing starter bacteria for the production of lower fat Cheddar cheese has lead to milder flavor Cheddar cheeses that lack intense Cheddar notes. The metabolism of methionine leads to the production of methanethiol, which is one of the desirable Cheddar cheese flavor compounds. The influence of NaCl and reduced pH was determined for aminopeptidase, lipase/ esterase, and methanethiol-producing capability in selected lactic acid bacteria and brevibacteria in simulated cheese-like conditions. The activity of each enzyme decreased with NaCl addition and pH reduction to approximate a Cheddar cheese environment (5% NaCl and pH 5.2). The mechanism for methanethiol production by the starter and adjunct bacteria was also investigated. Different enzyme systems were found to be responsible for methanethiol production in starter lactococci, lactobacilli, and brevibacteria. In the lactococci, enzymes that acted primarily on cystathionine were responsible for methanethiol production from methionine. Lactobacilli also contained cystathionine-degrading enzymes, but these enzymes have properties different from the lactococcal enzymes. Brevibacterium linensBL2 lacked cystathionine-degrading enzymes, but was capable of the direct conversion of methionine to methanethiol. L-Methionine γ-lyase from B. linens BL2 was purified to homogeneity, and was found to catalyze the α, γ elimination of methionine resulting in the production of methanethiol, α-ketobutyrate, and ammonia. Characterization of the pure enzyme demonstrated that it is pyridoxal phosphate dependent, which is active at salt and pH conditions existing in ripening Cheddar cheese. The addition of either B. linens BL2 or L-methionine γ-lyase to aseptic cheese curd slurries increased methanethiol and total volatile sulfur compound production. In an attempt to increase methanethiol production and Cheddar cheese flavor in reduced-fat Cheddar cheese, B. linens BL2 was added as a starter adjunct to 60% reduced-fat cheese. Sensory evaluation of the cheese indicated that B. linens BL2 improved the flavor of 60% reduced-fat Cheddar cheese. This suggests that the addition of B. linens BL2 is an alternative to the addition of lactic acid bacteria to improve Cheddar cheese flavor via the metabolism of methionine.

A Spiral Plater and a Microtiter system were used to isolate and evaluate cultures for a paired strain culture program. Bacteriophage and temperature sensitivity data of 43 Streptococcus cremoris strains were introduced into a computer cluster program to pair the least similar strains. Selected pairs were challenged with phage. Resistant mutants were developed. Characteristics of proteinase positive and proteinase negative variants were examined. Proteinase positive isolates produced more changes in pH, cell mass and more generations in milk than their counter-parts. Paired proteinase negative cultures produced more change in pH and cell mass and more generations in milk than single strains. Whey based medium under pH control was superior to commercial internal pH control medium for proteinase negative culture propagation. Proteinase negative isolates achieved 90% of the cell mass obtained by their counterparts in nonfat dry milk-yeast medium. Proteinase negative starter culture endured significantly higher phage titers than proteinase positive cells. Proteinase negative variants sustained activity comparable with phage-free controls when challenged for seven cycles with high phage titers. Proteinase positive cells had impaired activity after the second cycle. Pairing of proteinase positive strains was advantageous for phage protection. Erythromycin, streptomycin and penicillin adversely affected the activity of both cell types, yet proteinase positive cells were significantly more inhibited. Pairing neither variant enhanced activity. Cheddar cheese was exclusively manufactured with 2% inoculum of proteinase negative cultures compared to 1.5% usage of the proteinase positive paired strains. Cheese quality and cheese making times were normal. Over 4200 consecutive vats of Cheddar cheese were made in 1982 employing one pair of proteinase positive culture. Acid control and cheese quality were improved. The cheese making times were more uniform. Smaller inocula volumes could successfully be used for bulk starter in cheese plants utilizing pH controlled starter propagation. A needle/syringe system for inoculating starter tanks provided better protection against contamination during inoculation.

Graduation date: 1993

Thesis (Ph.D.)--University of Western Sydney, 2007. / A thesis submitted to the University of Western Sydney, College of Health and Science, Centre for Plant and Food Science, in fulfilment of the requirements for the degree of Doctor of Philosophy. Includes bibliographical references.