Stearoyl-CoA desaturase 1 (SCD1) greatly contributes to the unsaturated fatty acids present in milk and meat of cattle. The SCD1 enzyme introduces a double bond into certain saturated fatty acyl-CoAs producing monounsaturated fatty acids (MUFA). The SCD1 enzyme also has been shown to be active in the bovine mammary gland converting t11 18 : 1 (vaccenic acid) to c9 t11 conjugated linoleic acid (CLA). The objective of this study was to determine any association between the gene expression of SCD1 and occurrence of its products (c9 14 : 1, c9 16 : 1, c9 18 : 1, and c9 t11 18 : 2) in various bovine tissues. Tissue samples were obtained from lactating Holstein cows (n=28) at slaughter, frozen in liquid nitrogen and stored at −80 °C. Total RNA was extracted and converted to complementary DNA for quantitative real time polymerase chain reaction (PCR) analysis of the SCD1 gene. Extracted lipid was converted to fatty acid methyl esters and analysed by GC. Tissues varied in expression of SCD1 gene with mammary, cardiac, intestinal adipose, and skeletal muscle expressing greater copy number as compared with lung, large intestine, small intestine and liver (371, 369, 328, 286, 257, 145, 73, and 21 copies/ng RNA, respectively). Tissues with high mRNA expression of SCD1 contained greater SCD1 protein whereas detection of SCD1 protein in tissues with low SCD1 mRNA expression was very faint or absent. Across tissues, the desaturase indices for c9 18 : 1 (r=0·24) and sum of SCD products (r=0·20) were positively correlated with SCD1 gene expression (P<0·01 for both). Within each tissue, the relationship between SCD1 gene expression and the desaturase indices varied. No correlation was detected between SCD1 expression and desaturase indices in the liver, large and small intestines, lung, cardiac or skeletal muscles. Positive correlations, however, were detected between SCD1 expression and the desaturase indices in intestinal adipose tissue (P<0·02 for all) except 14 : 1, whereas only c9 18 : 1, c9 t11 18 : 2 and sum of all desaturase indices were positively correlated with SCD1 expression in mammary tissue (P⩽0·03). Overall, the relationship between SCD1 gene expression and occurrence of its products seems to be tissue specific.

Colostrum consists of a number of biologically active proteins and peptides that influence physiological function and development of a neonate. The present study investigated the biological activity of peptides released from first day bovine colostrum through in vitro and in vivo enzymatic digestion. This was assessed for proliferative activity using a human intestinal epithelial cell line, T84. Digestion of the protein fraction of bovine colostrum in vitro was conducted with the enzymes pepsin, chymosin and trypsin. Pepsin and chymosin digests yielded protein fractions with proliferative activity similar to that observed with undigested colostrum and the positive control foetal calf serum (FCS). In contrast trypsin digestion significantly (P<0·05) decreased colostral proliferative activity when co-cultured with cells when compared with undigested colostrum. The proliferative activity of undigested colostrum protein and abomasal whey protein digesta significantly increased (P<0·05) epithelial cell proliferation in comparison to a synthetic peptide mix. Bovine colostrum protein digested in vivo was collected from different regions of the gastrointestinal tract (GIT) in newborn calves fed either once (n=3 calves) or three times at 12-h intervals (n=3 calves). Digesta collected from the distal duodenum, jejunum and colon of calves fed once, significantly (P<0·05) stimulated cell proliferation in comparison with comparable samples collected from calves fed multiple times. These peptide enriched fractions are likely to yield candidate peptides with potential application for gastrointestinal repair in mammalian species.

The aim of this work was to study the effect on teat wall thickness and canal length in sheep and goats of overmilking for 2 min (OM+2) and of milking with used (AL; +3000 milkings) and twisted (TL; 45°) liners in sheep and goats, as well as the effect of milking goats with liners designed for sheep (SL, shorter length and diameter than liners for goats). To this end, we performed four experiments in goats and three in sheep, in a Latin square design with two experimental periods. During the experimental period 4 controls were carried out, performing ultrasound scans before and immediately after milking to determine the teat wall thickness (TWT), teat wall area (TWA), teat end area (TEWA) and teat canal length (TCL). OM+2 caused a significant increase in TWT, TWA, TEWA and TCL in goats and in TWA, TEWA and TCL in sheep. Liner features had a strong influence on the variables studied; aged liners caused significant changes in TWT and TCL in goats and in TWT in sheep; twisted liners produced a significant effect on the increase of TWT and TCL in goats, without reaching significance level in sheep; and milking goats with sheep liners led to a significant increase in TWT, TWA, TEWA and TCL. In practice, it is therefore important to avoid overmilking and the use of worn-out liners. It is also necessary to use liners designed for the morphological features of each species, taking special care to carry out periodic liner positioning revisions to ensure the benefits of pulsation on the teat end. Finally, it would be necessary to carry out long-term experiments to study whether the increase in thickness observed in some experiments is sufficient to affect milking efficiency and mammary gland health status.

Sampling approaches following the dairy chain, including microbiological hygiene status of critical processing steps and physicochemical parameters, contribute to our understanding of how Staphylococcus aureus contamination risks can be minimised. Such a sampling approach was adopted in this study, together with rapid culture-independent quantification of Staph. aureus to supplement standard microbiological methods. A regional cheese production chain, involving 18 farms, was sampled on two separate occasions. Overall, 51·4% of bulk milk samples were found to be Staph. aureus positive, most of them (34·3%) at the limit of culture-based detection. Staph. aureus positive samples >100 cfu/ml were recorded in 17·1% of bulk milk samples collected mainly during the sampling in November. A higher number of Staph. aureus positive bulk milk samples (94·3%) were detected after applying the culture-independent approach. A concentration effect of Staph. aureus was observed during curd processing. Staph. aureus were not consistently detectable with cultural methods during the late ripening phase, but >100 Staph. aureus cell equivalents (CE)/ml or g were quantifiable by the culture-independent approach until the end of ripening. Enterotoxin gene PCR and pulsed-field gel electrophoresis (PFGE) typing provided evidence that livestock adapted strains of Staph. aureus mostly dominate the post processing level and substantiates the belief that animal hygiene plays a pivotal role in minimising the risk of Staph. aureus associated contamination in cheese making. Therefore, the actual data strongly support the need for additional sampling activities and recording of physicochemical parameters during semi-hard cheese-making and cheese ripening, to estimate the risk of Staph. aureus contamination before consumption.

To evaluate the interaction between the levels of dietary concentrate and linseed oil (LO) on milk fatty acid (FA) profile, 24 Holstein cows were used in a randomised complete block design based on days in milk, with a 2×2 factorial arrangement of treatments. Within each block, cows were fed one of four experimental diets containing 30% concentrate (LC) or 70% concentrate (HC), without LO (NLO) or with LO supplemented at 3% of dietary dry matter. Milk FA profiles were analysed with a special emphasis on the intermediates of the predominant trans-11, and a putative trans-13 pathways of ruminal biohydrogenation of cis-9, cis-12, cis-15 18:3. Feeding LO increased the concentrations of cis-9, cis-12, cis-15 18:3 and trans-11, cis-15 18:2 in milk fat, and these increases were of a higher magnitude when LO was added in HC as compared with LC diet (interaction of LO by concentrate). A treatment interaction was also observed for the level of trans-11 18:1 which was higher when feeding LO, but for which the increase was more pronounced with the LC as compared with HC diet. The concentrations of cis-15 18:1 and cis-9, trans-11, cis-15 18:3 were higher in cows fed LO, but feeding HC diets decreased milk fat content of cis-15 18:1 and a tendency for a decrease in cis-9, trans-11, cis-15 18:3 was apparent. Feeding LO increased milk fat content of trans-13 18:1 and cis-9, trans-13 18:2, while the concentrations of these two isomers were not affected by the level of dietary concentrates. The isomer cis-9, trans-13, cis-15 18:3 has not been detected in any of the milk samples. In conclusion, interactions were observed between LO and dietary concentrates on the proportions of some intermediates of the trans-11 biohydrogenation pathway. The presence of trans-13 18:1 and cis-9, trans-13 18:2 supports the existence of a trans-13 pathway, but an 18:3 intermediate with a trans-13 double bond has not been identified.

The objective of this study was to estimate heritability and crossbreeding parameters (breed and heterosis effects) of various fatty acid (FA) concentrations in milk fat of New Zealand dairy cattle. For this purpose, calibration equations to predict concentration of each of the most common FAs were derived with partial least squares (PLS) using mid-infrared (MIR) spectral data from milk samples (n=850) collected in the 2003–04 season from 348 second-parity crossbred cows during peak, mid and late lactation. The milk samples produced both, MIR spectral data and concentration of the most common FAs determined using gas chromatography (GC). The concordance correlation coefficients (CCC) between the concentration of a FA determined by GC and the PLS equation ranged from 0·63 to 0·94, suggesting that some prediction equations can be considered to have substantial predictive ability. The PLS calibration equations were then used to predict the concentration of each of the fatty acids in 26 769 milk samples from 7385 cows that were herd-tested during the 2007–08 season. Data were analysed using a single-trait repeatability animal model. Shorter chain FA (16 : 0 and below) were significantly higher (P<0·05) in Jersey cows, while longer chain, including unsaturated longer chain FA were higher in Holstein-Friesian cows. The estimates of heritabilities ranged from 0·17 to 0·41 suggesting that selective breeding could be used to ensure milk fat composition stays aligned to consumer, market and manufacturing needs.

The impact of mastitis on milk value per litre independent of the effect of mastitis on milk volume, was quantified for Ireland using a meta-analysis and a processing sector model. Changes in raw milk composition, cheese processing and composition associated with increased bulk milk somatic cell count (BMSCC) were incorporated into the model. Processing costs and market values were representative of current industry values. It was assumed that as BMSCC increased (i) milk fat and milk protein increased and milk lactose decreased, (ii) fat and protein recoveries decreased, (iii) cheese protein decreased and cheese moisture increased. Five BMSCC categories were examined from ⩽100 000 to >400 000 cells/ml. The analysis showed that as BMSCC increased the production quantities reduced. An increase in BMSCC from 100 000 to >400 000 cells/ml saw a reduction in net revenue of 3·2% per annum (€51·3 million) which corresponded to a reduction in the value of raw milk of €0·0096 cents/l.

The possibilities of using high quality pastures in conjunction with total mixed ration (TMR) during the grazing season have been examined. An experiment with sixteen Holstein cows blocked and randomly assigned to four treatments in a factorial arrangement was conducted in order to evaluate the influence of grazing time of day (day or night) and type of silage (maize or Italian ryegrass) included in the TMR of dairy cows grazing 12 h daily on milk yield, composition and fatty acid profile. The silage type had no effect on the dry matter intake, milk yield and fat and protein proportions. However, cows grazing during the night ate more grass than cows grazing during the day (8·53 vs. 5·65 kg DM/d; P<0·05). No differences were seen between grazing-time with respect to milk production, fat and protein contents. However, the proportion of polyunsaturated fatty acid was higher in milk of dairy cows grazing at night-time than grazing at day-time, especially 18:2n-6 (2·37 vs. 2·12 g/100 g FA respectively, P<0·05) and 18:2cis9trans11 (2·08 vs. 1·74 g/100 g FA respectively, P<0·05).

The aim of this work was to study how machine milking (MM) carried out in suitable conditions affects teat wall thickness and teat canal length and their return after milking to pre-milking conditions, in comparison with other milk extraction methods considered biological referents: lamb suckling (LS), milk removal by catheter (RC) and hand milking (HM). Three Latin square experiments were designed, using 11 ewes in the first experiment (LS), 13 in the second (RC) and 12 in the third (HM). Each of the Latin squares was divided into two periods: in the first, the left gland of each animal was machine milked and the corresponding treatment (LS, RC and HM) was applied to the right gland. Subsequently, in the second period the extraction methods were interchanged. During the experimental period, 4 sampling days were carried out (2 in each experimental period), where ultrasound scans were taken before (B) and immediately after milking (A) and at 1 (1 h), 2 (2 h), 3 (3 h), 4 (4 h), 6 (6 h), 8 (8 h) and 10 (10 h) hours after milking finished. Teat wall thickness (TWT), teat wall area (TWA), teat end area (TEWA) and teat canal length (TCL) were measured in all the ultrasound images. MM increased TWT after milking compared with RC. TWT, TWA, TEWA and TCL were lower (P<0·05) in HM than in MM. No significant differences (P>0·05) were found between LS and MM for any variable. The extraction method affected the recovery time of the variables, with total teat recovery at 6 h after RC and 4 h after HM. In the case of LS, the TEWA and TCL values were recovered sooner, as of 3 h. Teat recovery time after MM was similar to the extraction method with which it was compared in each experiment. Thus, considering the similar increase in wall thickness and their recovery time compared with the reference methods, it was concluded that machine milking, carried out in optimum conditions and respecting the time interval between milkings usually applied on sheep farms (8–12 h), would not affect teat integrity. Moreover, given the variability observed in teat thickness recovery time between the different experiments, further research should be carried out to study which factors intrinsic and extrinsic to the animal may affect the teat wall thickness and recovery time after machine milking.

In order to protect peroral β-galactosidase from being degraded and hydrolyse milk lactose efficiently in the environments of gastrointestinal tract, a double-capsule delivery system composed of enteric-coated capsule and polylactic acid (PLA) nanocapsules (NCs) was developed for encapsulation of β-galactosidase. β-galactosidase-loaded PLA NCs in the size range of 100–200 nm were prepared by a modified w1/o/w2 technique. During the encapsulation process, dichloromethane/ethyl acetate (1 : 1, v/v) as the solvent composition, high-pressure homogenisation (150 bar, 3 min) as the second emulsification method and polyvinyl alcohol or Poloxamer 188 as a stabiliser in the inner phase could efficiently improve the activity retention of β-galactosidase (>90%). Subsequently, the prepared NCs were freeze-dried and filled in a hydroxypropyl methylcellulose phthalate (HP55)-coated capsule. In vitro results revealed that the HP55-coated capsule remained intact in the simulated gastric fluid and efficiently protected the nested β-galactosidase from acidic denaturation. Under the simulated intestinal condition, the enteric coating dissolved rapidly and released the β-galactosidase-loaded PLA NCs, which exhibited greater stability against enzymatic degradation and higher hydrolysis ratio (∼100%) towards milk lactose than the free β-galactosidase. These results suggest that this double-capsule delivery system represents promising candidate for efficient lactose hydrolysis in the gastrointestinal tract.

Lactones are important contributors to the flavour and aroma of milk and dairy products. This study was conducted to evaluate the effects of dietary linseed oil (LO) and forage to concentrate ratio on milk lactone profile. Twenty four Holstein cows were used during a 4-week feeding trial in a randomised complete block design. Cows were fed diets containing 30% (LC) or 70% (HC) concentrate, and 0% (NLO) or 3% LO in a 2×2 factorial arrangement of treatments. Milk lactone profile was evaluated using the solid phase microextraction technique. The highest levels of δ-lactones (δ-6:0, δ-8:0, δ-10:0, and δ-12:0) were found with the LC/NLO diet. These concentrations were then decreased when cows received either a high level of concentrate or supplemental LO, but these effects were not additive (interaction of LO by concentrate, P<0·01). An interaction of LO by concentrate (P<0·01) was also noted on milk γ-12:0 for which the highest concentration was observed when supplementing LO in HC diet, while no effect was apparent when LO was added in LC diet. Moreover, feeding HC increased the level of γ-12:1 in milk as compared with LC, while LO had no effect on this γ-lactone. Finally, γ-12:2 was not detected in any of the milk samples studied. Organoleptic properties of milk were evaluated in a triangle test showing that a significant number of assessors perceived a difference between milk from cows fed LC/NLO as compared with milk from cows fed HC/LO. The sensory evaluation was completed by a ranking test where the intensities of fresh lactic, foreign and global flavours were not different between treatments. In conclusion, feeding LO in HC diet modified milk lactone profile with a shift toward more γ- and less δ-lactones as compared with LC diet not supplemented with LO. A difference was perceived in a triangle test between milk from these two treatments, but the sensory attributes responsible for this difference have not been identified in the current trial.

This work applies culture-independent methods for the characterization of fungal populations (yeasts and moulds) naturally occurring in Sardinian ewe's milk sampled in the Italian areas with the largest dairy production (Sardinia and Lazio regions). Sequences of the D1/D2 variable domains at the 5′ end of the 26S rRNA gene were obtained by amplification of DNA directly isolated from milk, and this allowed identification of a total of 6 genera and 15 species of fungi. Among the 6 identified genera Geotrichum spp., Candida spp., Phaeosphaeriopsis spp., Pestalotiopsis spp. and Cladosporium spp. belong to the phylum of Ascomycota, while Cryptococcus spp. is part of the phylum of Basidiomycota. In particular, two genera (Pestalotiopsis and Phaeosphaeriopsis) and two species (Plectosphaerella cucumerina and Pryceomyces carsonii) have never been reported in dairy ecosystems before. Results provide evidence that several moulds and yeasts, previously described only in ovine cheeses, are transferred directly from raw milk. The knowledge of fungal consortia inhabiting sheep raw milk is a particularly relevant issue because several species are directly involved in cheese making and ripening, determining the typical aroma. On the other hand, spoilage yeasts and moulds are involved in anomalous fermentation of cheese and may be responsible for considerable economic losses and serious risks for consumers' health.

The assumption, that metabolites derived from the activity of the mammary gland epithelial cells reflect changes in milk secretion and its coagulation properties, was tested in dairy cows. The experiment included cows with uninfected udders and cows with one of the glands infected by different bacteria specie. Analysis were carried at the cow level (including all four glands), or at the gland level. High and significant correlations among the concentrations of lactose, glucose, glucose-6-posphate, milk related respiratory index (the ratio between the concentrations of citrate/lactate+malate in milk) and milk-derived glycolytic index (the ratio between glucose-6-phosphate and glucose in milk) and milk clotting parameters were found. The physiological basis for these relations and their ability to predict the deterioration in milk quality in subclinically infected glands and in glands previously clinically infected with Escherichia coli are discussed.

Milk yield and milk qualitative parameters were evaluated in Lacaune ewes on a diet supplemented with extruded linseed, verbascoside and vitamin E. A 98 d-trial was conducted on 44 ewes and started 40±2 d post partum. The animals were divided into four homogeneous groups of eleven animals each; one control group (CON) without extruded linseed and dietary supplements, and the diet of the other three experimental groups was enhanced with extruded linseed (L group), extruded linseed-verbascoside (LVB group), and extruded linseed-verbascoside-vitamin E (LVBE group). All animals individually received an isoenergetic diet, consisting of 700 g concentrated feed and meadow hay ad libitum. Body weight, body condition score, milk yield and milk qualitative parameters were assessed. LVB and LVBE groups resulted in a significant improvement (P<0·05) in milk yield due to the verbascoside supplementation. The extruded linseed supplementation L, LVB and LVBE groups produced a milk fat increase and a better milk fatty acid profile in terms of a higher monounsaturated fatty acid (MUFA), polyunsaturated fatty acid (PUFA) and conjugated linoleic acid (CLA) content and a reduced saturated fatty acid (SFA) content, a lower n-6/n-3 ratio and atherogenic and thrombogenic index. The dietary verbascoside supplementation in the LVB and LVBE group resulted in a better milk quality due to the low cholesterol level and higher vitamin A and E contents, in addition to an increased oxidative stability highlighted by the lower thiobarbituric acid reactive substances (TBARS) level. Thus, the addition of extruded linseed and verbascoside supplements improved milk yield and quality both from a chemical and nutritive point of view.

Alpha-lactalbumin (α-la) is one of the major proteins in whey. When partially hydrolysed with Bacillus licheniformis protease, it produces nanotubular structures in the presence of calcium ions by a self-assembly process. This study presents investigation of α-la protein structure during hydrolysis and nanotube formation using optical spectroscopy. Before spectroscopic measurements, nanotubes were examined with microscopy. The observed α-la nanotubes (α-LaNTs) were in the form of regular hollow strands with a diameter of about 20 nm and the average length of 1 μm. Amide and backbone vibration bands of the Raman spectra displayed remarkable conformational changes in α and β domains in the protein structure during nanotube growth. This was confirmed by the Fourier-transform infrared (FTIR) spectroscopy data. Also, FTIR analysis revealed certain bands at calcium (Ca++) binding sites of COO− groups in hydrolysed protein. These sites might be critical in nanotube elongation.

The aggregation of proteins after heating of calcium-fortified milks has been an ongoing problem in the dairy industry. This undesirable effect restricts the manufacture of calcium rich dairy products. To overcome this problem, a completely new approach in controlling the heat stability of dairy protein solutions, developed in our lab, has been employed. In this approach, high intensity, low frequency ultrasound is applied for a very short duration after a pre-heating step at ⩾70 °C. The ultrasound breaks apart whey/whey and whey/casein aggregates through the process of acoustic cavitation. Protein aggregates do not reform on subsequent post-heating, thereby making the systems heat stable. In this paper, the acid gelation properties of ultrasonicated calcium-enriched skim milks have also been investigated. It is shown that ultrasonication alone does not change the gelation properties significantly whereas a sequence of preheating (72 °C/1 min) followed by ultrasonication leads to decreased gelation times, decreased gel syneresis and increased skim milk viscosity in comparison to heating alone. Overall, ultrasonication has the potential to provide calcium-fortified dairy products with increased heat stability. However, enhanced gelation properties can only be achieved when ultrasonication is completed in conjunction with heating.

This study investigated the antibacterial properties of plasmin, the plasmin hydrolysis of bovine κ-casein and the fractions (named κC1, κC2, κC3, κC4, and κC5) liberated from it using RP-HPLC. The target bacteria were Escherichia coli, Staphylococcus aureus (pathogenic), Lactobacillus casei and Lactobacillus acidophilus (probiotic). Three peptides (kC1, kC3, and kC4) were found to have antibacterial activity, with κC3 peptide being the most active. The plasmin digest of bovine κ-casein proved to be stronger than any of its fractions in terms of antibacterial potential. Measurement of the minimum inhibitory concentration (MIC) showed that Gram-positive bacteria are generally more sensitive to antibacterial activity than Gram-negative bacteria. The MIC of nisin, as a bacteriocin peptide, was also measured. The three antibacterial peptides were identified using LC-Mass. The molecular mass of kC1, kC3, and kC4 corresponded to the f(17–21), f(22–24), and f(1–3) of bovine κ-casein, respectively. It was also found that the positive charge and hydrophobicity of a peptide are not key factors in antibacterial activity. On the whole, the present study demonstrated that the plasmin digest of κ-casein has a high antibacterial potential and can be considered as a natural antibacterial agent in the food chain.

We studied the effects of changes in the milking routine (lack or presence of 30-s prestimulation, 0 or 1, 2 or 4-min delay between preparation and cluster attachment) and environmental perturbation (unusual loud sounds capable of frightening animals just after stall entry or during the course of milking) on milk removal and milking-related behaviour in dairy dromedary camels. A 30-s prestimulation decreased incidence of bimodal milk flow curves and increased occurrence of the best milk ejection patterns with higher milk flow but had limited effect on milk production in our well-trained animals within a good machine milking setting. However, unusual sounds heard from the beginning of milking or even after milk ejection caused inhibition or disruption of milk removal and modification of camels’ behaviour. Milk ejection was significantly delayed (1·58±0·17 min), residual milk increased over 40% of total milk yield and average and peak milk flow rates were significantly lowered when unusual noises were heard from the beginning of milking. These environmental perturbations increased signs of vigilance and the number of attempts to escape the milking parlour. Delaying cluster attachment for over 1 min after the end of udder preparation caused serious milk losses. Up to 62% of total milk was withheld in the udder when the delay reached 4 min. Average and peak milk flow rates also decreased significantly with delayed milking. Signs of vigilance and attempts to escape from the milking parlour appeared when camels waited for over 2 min. After a 4-min delay, camels showed signs of acute stress. Defaecation prior to milk ejection (solid faeces) and rumination during milking can be used to assess camels’ milk ejection during milking. Animal welfare and milking efficiency can be ensured when camels are pre-stimulated, milked in calm conditions and with cluster attachment within a maximum of a 1-min delay after stimulation

The goal of the present study was to examine the suitability of a short pre-stimulation (P) for 15 s followed by a latency period (L) of 30 s before cluster attachment for machine milking. In addition we tested the effect of a periodic reduction of the vacuum under the teat (VR) during the massage phase from 43 kPa to 12–15 kPa on milking characteristics and teat tissue condition. The study was carried out in 9 cows in a cross-over design. Animals were milked twice daily, and each of the 4 treatment combinations was used for six subsequent milkings (P+L vs. continuous P, and standard pulsation vs. VR, respectively). Milk flow was recorded during all experimental milkings. Longitudinal ultrasound cross sections of the teat were performed by B-mode ultrasound after the last milking of each treatment at 0, 5, and 15 min after the end of milking, respectively. None of the evaluated milking characteristics (total milk yield, main milking time, peak flow rate, average milk flow) differed between treatments. Teat measures as obtained by ultrasound cross sections showed no significant difference if individual treatments were compared at the three time points individually. However, teat wall thickness (TWT) tended to be smaller in VR vs. non-VR treatments at 5 min after milking (P=0·05). In conclusion, teat preparation consisting of a short stimulation followed by a latency period represents a similarly efficient pre-stimulation as a continuous pre-stimulation. VR seems to reduce the load on the teat tissue during milking and thus reduces the development of oedema and hence a less pronounced increase of TWT while milking characteristics are similar with or without VR.

Caseinophosphopeptides (CPPs) are multifunctional bioactive peptides containing phosphorylated seryl residues in their sequence. In the present study, method for the production of CPPs from buffalo milk casein was optimised and characterised for their sequence, calcium solubilising and calcium binding activities. Response surface methodology was used to optimise the conditions for hydrolysis of buffalo casein by trypsin to obtain maximum yield of CPPs. The optimum hydrolysis conditions were as follows: hydrolysis pH 7·5, temperature 37 °C, hydrolysis time 7·0 h. Under these conditions, the experimental yield obtained was 10·04±0·24%, which is slightly lower than value predicted by the model. These CPPs were able to solubilise 1·03±0·08 mg la/mg CPPs in presence of excess phosphate and bind 0·935 mg of Ca/mg of CPPs. Eight phosphopeptides i.e. αs1-CN f (37-58) 2P; αs1-CN f (37-58) 3P; αs1-CN f (35-58) 2P; αs1-CN f (35-58) 3P; αs2-CN f (2-21) 4P; αs2-CN f (138-149) 1P; β-CN f (2-28) 4P and β-CN f (33-48) 1P were identified by LC-MS/MS which contained motif for binding of divalent minerals. The sequences of these CPPs differed from that of derived from bovine casein.