Acid aspartyl proteinase of Penicillium roqueforti was purified from culture filtrates using FPLC ion-exchange chromatography on Mono Q and size exclusion chromatography on Superdex 75. The purity obtained allowed us to produce, using rabbits, a specific antiserum which was then used to develop a sandwich ELISA. The test was sensitive (detection limit, 0·25 ng/ml), reproducible (CV, 3·1–6·9%) and closely correlated with enzyme activity measurement (r=0·995, P<0·001). This ELISA should be a valuable tool for monitoring acid aspartyl proteinase level in culture filtrates and for determining the enzyme activity[ratio ]enzyme protein ratio of acid aspartyl proteinase obtained after genetic recombinations or site-directed mutagenesis.

The concentrations of seven types of microorganism in the milk produced by farms using high quality milking procedures were monitored monthly. The most commonly encountered bacterium was Pseudomonas, whose concentration varied greatly between samples, but lactococci, lactobacilli and yeasts were also present at more stable concentrations. Staphylococcus aureus and β-glucuronidase-positive Escherichia coli were occasionally detected. Listeria monocytogenes was found in the samples from only one farm, while Yersinia enterocolitica was never detected. There were seasonal variations in the concentrations of lactobacilli and yeasts over the 2 years monitored. The changes in certain bacteria (Lactococcus, Lactobacillus) on any given farm were very similar from one year to the next. The microbiological characteristics of the milk from the farms examined seemed to be fairly constant. The profiles of some bacteria used for cheesemaking were relatively stable over time. It thus seems that the milk provided by different producers could be typed.

Mineral acid casein, lactic casein and rennet casein curds were prepared either in the laboratory or on pilot-scale equipment. The water-holding capacities of mineral acid casein curd particles prepared in the laboratory were relatively constant for particle sizes >4 mm both after sieving and after centrifugation; however, they increased dramatically at smaller particle sizes to a maximum of 7·2 g water/g dry curd after sieving and 4·3 g water/g dry curd after centrifugation at 4060 g for the size fraction that passed a 0·85 mm screen. The mechanical strength of the centrifugally compressed curd as assessed by a penetrometer probe also decreased dramatically with decrease in particle size. This variation of mineral acid casein curd properties with particle size was attributed partly to variation in particle pH at the instant of curd formation because of imperfect mixing of acid and skim milk, i.e. localized pH gradients. Lactic casein and rennet casein curds and curd formed by acidification of skim milk with glucono-δ-lactone also produced similar but smaller variations in curd properties with particle size. These curd particles are all formed by break up of a quiescent gel and the variation in curd properties with particle size was attributed to variations in mechanical strength in the original milk gel, i.e. weak areas of the gel break up more easily, resulting in small, weak curd particles.

Two methods were compared for the determination of free fatty acids (FFA) from acetic to long-chain acids in samples with a large excess of triacylglycerols (TG) (1[ratio ]200, w/w), such as cheese and other dairy products. In method 1, after fat extraction, FFA were separated from TG by aminopropyl-bonded phase chromatography, injecting the fraction containing FFA directly into the gas chromatograph. In method 2, extracted fat was treated with tetramethylammonium hydroxide, the methyl ester derivatives being formed in the injector. Cheese samples and standard mixtures of FFA and TG in different proportions were analysed by both methods. The cheese sample contained 2·4 times more FFA when analysed by method 2 as compared with the result obtained with method 1. The composition of the standard mixtures analysed by method 1 closely reflected that of the original mixture and gave 90–100% recovery of FFA, regardless of their chain length and the ratio of FFA[ratio ]TG (1[ratio ]1 or 1[ratio ]200, w/w). The composition of samples with a FFA[ratio ]TG ratio of 1[ratio ]200 (w/v) was severely distorted (as compared with the original composition of the sample) when analysed by method 2. Varying recoveries of FFA were also obtained, the largest differences being found for the shorter-chain components. We conclude that the FFA fraction should be separated from the TG fraction before derivatization and chromatographic analysis, particularly for samples in which the FFA represent a minor fraction of the TG.

The effects of ruminant whey and its purified fractions on neutrophil chemotaxis and superoxide production in sheep were studied. Both colostral whey and milk whey were found to inhibit chemotaxis regardless of whether they were autologous or homologous, but the inhibitory effects were abolished by washing neutrophils with culture medium before their use in the chemotaxis assay. Colostral whey and milk whey also inhibited the chemotactic activity of zymosan-activated serum. Whey fractions of various degrees of purity such as lactoferrin, lacto-peroxidase, lactoferrin–lactoperoxidase, α-lactalbumin, bovine serum albumin and whey protein concentrate were then studied. While none of these proteins showed any effects on chemotaxis, lactoferrin–lactoperoxidase and whey protein concentrate were found to have an enhancing effect on superoxide production in a dose-dependent manner. Our results provide information on the modulatory role of ruminant milk proteins in inflammatory responses and warrant future investigation.

The possible hypocholesterolaemic properties of milk and fermented milk products have been investigated in groups of albino rats given a basal diet, basal diet plus cholesterol, and basal diet plus cholesterol together with whole milk or standard or bifidus yogurt. The yogurts were fortified with skim milk powder, condensed whey or lactose-hydrolysed condensed whey. After 30 d, triacylglycerols, total cholesterol, HDL-cholesterol and LDL-cholesterol were measured in serum. Whole milk and ordinary yogurt had no hypocholesterolaemic effect, but standard yogurt containing lactose-hydrolysed condensed whey and all bifidus yogurts lowered serum cholesterol. In general, yogurts changed HDL-cholesterol little, but tended to raise triacylglycerols. There was marked lowering of LDL-cholesterol in rats given either type of yogurt fortified with whey proteins. This study has demonstrated in a rat model that bifidus yogurts and yogurts fortified with whey proteins can reduce total and LDL-cholesterol, and suggests that if they have the same effect in human subjects they have potential value in cholesterol-lowering diets.

Microstructural and textural characteristics of rennet curd made from rehydrated non-fat dried milk standardized to 40 g fat/l were compared with those of non-fat curd made with capsule-forming non-ropy lactic culture. Microstructure examination using confocal scanning laser microscopy indicated that the presence of a capsule-forming culture produced an open casein network with large pores. Distribution of fat globules was observed within the undisturbed casein structure after staining with a fluorescent lipophilic dye which was added to the milk before coagulation. Bacterial capsules and milk fat reduced curd tension and firmness of rennet curd to similar levels.

The effect of a range of milk heat treatments on the rheological properties, at small and large deformations, of acid skim milk gels was investigated. Gels were made from reconstituted skim milk heated at 75, 80, 85 and 90°C for 15 or 30 min by acidification with glucono-δ-lactone at 30°C. Gels were also made from skim milk powder (SMP) samples that had been given a range of preheat treatments during powder manufacture. Heating milks at temperatures [ges ]80°C for 15 min increased the storage moduli (G′) compared with unheated milk and produced gels with G′ in the range 300–450 Pa. Acid gels made from high-heat or medium-heat SMP had higher G′ than gels made from low-heat or ultra-low-heat SMP. Cooling gels to low temperatures resulted in an increase in G′. The yield stress of gels slightly decreased with mild heat treatments of milk, and then increased again to a maximum, finally decreasing slightly with very high heat treatments of milk. The strain at yielding decreased markedly with increasing heat treatment of milk, making these gels brittle and easier to fracture. We propose that denatured whey proteins aggregated with casein particles during the acidification of heated milk and were responsible for most of the effects observed in this study.

The whey proteins of the cow, goat and sheep have previously been fractionated on an analytical scale by reversed-phase HPLC (De Frutos et al. 1992), anion-exchange FPLC (Andrews et al. 1985; Manji et al. 1985; Laezza et al. 1991) and gel permeation FPLC (Andrews et al. 1985; Hill & Kakuda, 1990). Anion-exchange and gel permeation FPLC can readily be scaled up for laboratory preparation of whey protein fractions. There is some indication, however, that anion-exchange FPLC does not give complete separation of β-lactoglobulin and α-lactalbumin from the other minor whey protein fractions (Girardet et al. 1989).

In previous work it has been shown that gel permeation FPLC gives a satisfactory fractionation of the whey proteins of the cow (Law et al. 1993), goat (Law & Brown, 1994) and sheep (Law, 1995). In this paper we describe a scaled-up method of gel permeation that can be used for fairly rapid preparation or purification of four main whey protein fractions from the milks of these species.

Some of the textural changes that occur early in cheese maturation may be related to the redistribution of water within the cheese matrix. To examine this, a model cheese curd system was devised and explored. Initially, cheese curd was prepared using starter and chymosin and the curd pH was controlled by varying the draining and salting pH values. The quantity of serum that could be centrifuged from the resultant curd was less for lower pH curd and decreased in volume with time. The curd pH decreased with time. In the protocol finally adopted, milk was acidified with lactic acid and coagulated with Rennilase 46L. After cheddaring, salting and light pressing, the samples of this curd were finely diced and mixed with glucono-δ-lactone to give curd samples with comparable moisture contents, similar casein proteolysis rates but different pH values. The quantity of serum that could be centrifuged from these samples was greater for pH 5·6 curd than for pH 5·2 curd and decreased faster for the lower pH curd. Neither the curd moisture nor the pH changed significantly during curd storage and the casein proteolysis was low. These results for the model curd system are consistent with known water absorption characteristics of casein curd under ‘equilibrium’ conditions and the effects of pH and mineral salts on this absorption. It was concluded that, during the early stages of cheese ripening, there may be a redistribution of moisture within the cheese, related to the basic properties of the protein matrix and the transient effects of curd salting, rather than as a direct consequence of glycolytic and proteolytic changes.

Hydrophobic and hydrophilic peptides in the water-soluble fraction of 20 batches of Hispanico cheese made from pasteurized and raw milk were determined by reversed-phase HPLC, with detection at 214 and 280 nm. Cheese flavour characteristics were evaluated by a sensory panel, and regressions of bitterness scores on levels of hydrophobic and hydrophilic peptides and their ratio were calculated. The best fitting relationship for pasteurized milk cheese was the linear regression of mean panel bitterness scores on hydrophobic peptides at 280 nm (r2=0·791). The determination coefficient for the regression of hydrophobic peptides at 280 nm on panellist bitterness scores (r2=0·356) was lower, owing to individual differences in the perceived intensity of bitterness. In the case of raw milk cheese, the respective determination coefficients were 0·203 for panel scores and 0·034 for individual panellist scores.

Casein phosphopeptides (CPP) which develop in Grana Padano cheese at different ages were isolated by precipitation with Ba2+ and analysed by HPLC. Profiles were complex throughout the period between 4 and 38 months. CPP in a cheese sample 14 months old were identified by a combination of fast atom bombardment–mass spectrometry and Edman degradation. They were found to consist of a mixture of components derived from three parent peptides, β-CNf(7–28)4P, αs1-CNf(61–79)4P and αs1-CNf(7–21)4P. In total, 45 phosphopeptides were identified: 24 from β-CN, 16 from αs1-CN and 5 from αs2-CN. The presence of aminopeptidase activity during cheese ripening was deduced from the presence of a number of CPP of different lengths with the loss of one or more residues from the N-terminus. The longest had C-terminal lysine and seemed to be progressively hydrolysed by carboxypeptidases A and B to shorter peptides. CPP in cheese appeared to be shortened plasmin-mediated products. Moreover, those most resistant to further hydrolysis contained at least three closely located phosphoserine residues. The anticariogenic activity of CPP is also discussed.

An optimum or a high heat treatment of milk before yogurt preparation was applied according to an experimental design with the deliberate aim of making stirred yogurts with varying properties. Images of sections of yogurt samples were obtained using a transmission electron microscope. These were digitized and segmented via a threshold determination to separate the protein structure from the background. Four spatial statistical estimates were determined in order to evaluate their applicability for characterization of yogurt microstructure. The volume content was estimated and compared with the protein content obtained through chemical analysis. Heat treatment had no significant effect on volume content, as expected. The surface area of the protein aggregates was a useful measure to describe the difference between casein networks obtained by an optimum and a high heat treatment. The star volume gave a measure of the average local size of pores in the yogurt gel. The optimum heat treatment resulted in yogurt with small values for the star volume, indicating a less coarse structure of the casein network compared with yogurt prepared from high heat milk. The covariance function was able to differentiate between yogurts made using two types of heat treatment.

Control programmes for contagious mastitis pathogens, primarily Streptococcus agalactiae and Staphylococcus aureus, consisting of milking hygiene, treatment or isolation of infected cows and post-milking teat disinfection are relatively effective (McDonald, 1970; Natzke, 1977; Fox & Gay, 1993). Similar control programmes are often ineffective in the prevention of intramammary infections caused by environmental bacteria (Eberhart, 1977; Smith et al. 1985; Erskine et al. 1991; Smith & Hogan, 1993). This disparity of success relates to the fact that the principal source of environmental mastitis pathogens is the cow's environment, rather than another cow with an intramammary infection (Eberhart, 1977; Smith et al. 1985). Infections of environmental origin are particularly noteworthy because they may remain common in well managed herds, and these infections often cause severe clinical signs and high case fatality rates (Eberhart, 1977; Smith et al. 1985; Erskine et al. 1991; Smith & Hogan, 1993).

Control programmes for environmental mastitis usually include premilking teat disinfection, decreased use of water in udder preparation, increased concentrations of dietary vitamin E and selenium, improved sanitation of the environment and modification of cow behaviour in the post-milking period (Eberhart, 1977; Smith et al. 1984, 1985; Weiss et al. 1990; Erskine et al. 1991; Smith & Hogan, 1993). Environmental infections are thought to result when bacteria lacking specific virulence factors penetrate the teat sphincter (Eberhart et al. 1979; Smith et al. 1985). The bulk of these exposures are thought to occur between milkings. Control of these infections is problematic because the cow's teat sphincter remains open after mechanical milking (McDonald, 1975a). Presumably, cows are predisposed to intramammary infections when the open teat sphincter is exposed when cows lie down. Hence, dairy managers are advised to offer fresh feed to cows immediately after milking to increase the likelihood that the teat sphincter will close at least partly before cows become recumbent.

The purpose of this study was to determine the effect of feed availability on post-milking behaviour in dairy cows. Specifically, the measured dependent variable was the length of time that cows remained standing after leaving the milking parlour.

There is considerable interest in the relationship between milk protein polymorphism and cheesemaking properties of milk (e.g. Jakob & Puhan, 1992, 1995; Jakob, 1993). For example, the B variant of κ-casein is associated with firmer cheese curds, shorter renneting times, a smaller average micelle size, higher milk casein and κ-casein concentrations and lower whey protein concentrations (Jakob, 1994; Horne et al. 1995).

Lipolysis has many effects in dairy technology. Undesirable consequences include deleterious effects on the organoleptic properties of milk and some cheeses; however, it is desirable in some other cheeses.

The usual methods for detecting lipolysis in milk measure the amounts of free fatty acids liberated from triacylglycerols (International Dairy Federation, 1991). Lipase activity is detected by incubation of samples with substrates such as tributyrin (Castberg et al. 1975) or triolein (Nilsson-Ehle & Schotz, 1976; Shirai et al. 1982). A few reports describe methods using synthetic chromogenic substrates, e.g. p-nitrophenyl butyrate (Shirai & Jackson, 1982), β-naphthyl caprylate (McKellar, 1986; McKellar & Cholette, 1986) or 4-methylumbelliferyl oleate (Stead, 1983). Tsakalidou et al. (1992) have developed the detection of esterase activities on electrophoresis gels.

Parmigiano Reggiano is a hard, cooked cheese produced in specific areas of Northern Italy. The raw material is obtained by mixing the partly skimmed evening milk and whole morning milk. The mixture is then heated to 22°C, and natural whey starter is added at 28–30 g/l after 1–2 min, bringing the pH of the mixture to 6·2–6·3. After coagulation, which takes ∼15 min and occurs at 32–33°C owing to the addition of calf rennet powder, the curd is broken up for 2–4 min, cut into fragments and cooked at a temperature raised gradually to 42–44°C and then more quickly to 55–56°C over 10–15 min. The curd is left undisturbed, covered by the whey, for 40–60 min, then removed and placed inside a circular wooden mould. The cheese is held at ∼20°C for 3 d during which it is turned at frequent intervals to facilitate complete whey drainage. The cheese, which now has its typical shape and size, is then salted by immersion in brine (260–280 g NaCl/l at 16–17°C) for 20–24 d. During this period the cheese absorbs 15–18 g NaCl/kg and its weight decreases by 4%. During the subsequent period of ripening (12–24 months) in store rooms at 16–18°C and 85% moisture, the cheese is frequently turned. At the end of ripening the cheese is cylindrical in shape, with a slightly convex side, 0·22–0·24 m high, 0·40–0·45 m diam. weighs 35–36 kg, has 320–330 g fat/kg dry matter and a minutely granulated internal structure with small holes formed by the activity of some heterofermentative lactic acid bacteria.

Milk production in Ireland, New Zealand and Australia is seasonal, with the majority of cows calving in spring. This pattern of production makes the maximum use of grazed grass, and in Ireland >80% of total milk for manufacturing is produced between April and November inclusive. Such a seasonal pattern of production results in a large variation in the gross composition of milk due to stage of lactation effects (Phelan et al. 1982). Some studies have investigated the relationship between milk composition and its processing characteristics (O'Keeffe et al. 1982; Grandison et al. 1984); however, in these studies the effects of diet and lactation stage were not segregated. Kefford et al. (1995) attempted to segregate the effects of diet and stage of lactation and concluded that diet quality (type and quantity) had a larger effect on Cheddar cheese quality than the stage of lactation.

The nutritive value of the diet of cows fed on grazed grass can change owing to changes in grass supply or quality. In the current study, changes in grass supply were achieved by altering the daily herbage allowance (DHA) to the herds. The objective of the current study was to investigate the effect of varying the DHA in the range 16–24 kg grass dry matter (DM), which is typical of the variation in pasture allowance in Ireland in mid lactation, on milk composition and its processing characteristics.

The camel population of Saudi Arabia is ∼607000 out of a world population of 17 million. All are dromedaries (Camelus dromedarius) of four major ecotypes (Elamin & Wilcox, 1992), giving daily milk yields of 3·5–35·0 kg/animal (Knoess, 1977). Camel milk is low in lactose compared with cows' milk (Elamin & Wilcox, 1992). However, levels of potassium, magnesium, iron, copper, manganese, sodium and zinc are higher than in cows' milk (Sawaya et al. 1984; Abu-Lehia, 1987). Farah (1993) found low levels of potassium and phosphorus in Egyptian dromedaries. There is little difference between the milk of dromedaries and that of bactrian camels (Kheraskov, 1961).

In the Gulf region and some southern Mediterranean countries, substantial amounts of camel milk are consumed by the local people. The investigation reported here was undertaken to shed more light on the nutritional contribution made by camel milk.

Lowering the lactose content of milk in vivo has two potential benefits: to develop milk products for lactose intolerant people (Delmont, 1983) and to produce a more concentrated milk, reducing milk storage volume and milking frequency without affecting the overall fat and protein yields. Lactose synthesis in the lactating mammary gland results from the interaction of α-lactalbumin (α-la) with a Golgian UDP-galactosyltransferase (EC 2.4.1.22), and it has been suggested that lactose and α-la contents are directly related (Fitzgerald et al. 1970). Evidence that α-la is the only protein responsible for the in vivo induction of lactose synthesis came recently from knock-out experiments of the relevant gene (Stinnakre et al. 1994; Stacey et al. 1995), which disrupt lactation, as lactose-deprived milk of α-la-deficient mice is highly viscous. However, although the lactose, fat and protein contents of the milk were found to be directly related to the quantity of α-la present (Stinnakre et al. 1994), only in mice homozygous for the α-la null allele was there a significant alteration of milk concentration (Stacey et al. 1995). Stacey et al. (1995) also demonstrated that human α-la could substitute for its mouse counterpart via homologous recombination.

In the present study, we report the generation of transgenic mice producing only bovine α-la by back-crossing animals from four independent bovine α-la transgenic lines with α-la-deficient mice. Our work confirms that foreign α-la can substitute for the mouse protein and suggests a negative correlation between the α-la content and protein and fat concentrations in milk.