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Use of urea-polyacrylamide electrophoresis for discrimination of A1 and A2 beta casein variants in raw cow's milk.

ABSTRACT: Beta-casein (BC) in cow's milk occurs in several genetic variants, where BC A1 (BCA1) and BC A2 (BCA2) are the most frequent. This work deals with a method based on modified polyacrylamide gel electrophoresis using urea PAGE to discriminate BCA1 and BCA2 variants from Holstein Friesian (HF) and genetically selected Jersey A2/A2 (JA2) cow's milk. Two well defined bands were obtained from BC fraction of HF milk, while that of JA2 showed a single band. Proteins from these bands were sequenced by HPLC-quadrupole linear ion trap/mass spectrometry, resulting in BCA1 and BCA2 separation from the BC fraction of HF milk, whereas BCA2 was the only constituent of JA2 fraction. This method represents a feasible and useful tool to on site phenotyping of BC fraction of cow's milk for pharmaceutical and food industries applications.

SUBMITTER: Duarte-Vazquez MA

PROVIDER: S-EPMC5897294 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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Use of urea-polyacrylamide electrophoresis for discrimination of A1 and A2 beta casein variants in raw cow's milk.

Duarte-Vázquez Miguel Angel MA García-Ugalde Carlos Raúl CR Álvarez Blanca Erika BE Villegas Laura Mariana LM García-Almendárez Blanca Estela BE Rosado Jorge Luis JL Regalado Carlos C

Journal of food science and technology 20180302 5

Beta-casein (BC) in cow's milk occurs in several genetic variants, where BC A1 (BCA1) and BC A2 (BCA2) are the most frequent. This work deals with a method based on modified polyacrylamide gel electrophoresis using urea PAGE to discriminate BCA1 and BCA2 variants from Holstein Friesian (HF) and genetically selected Jersey A2/A2 (JA2) cow's milk. Two well defined bands were obtained from BC fraction of HF milk, while that of JA2 showed a single band. Proteins from these bands were sequenced by HP ...[more]

PMID: 29666547

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Project description:BackgroundCows' milk generally contains two types of β-casein, A1 and A2 types. Digestion of A1 type can yield the peptide β-casomorphin-7, which is implicated in adverse gastrointestinal effects of milk consumption, some of which resemble those in lactose intolerance. This study aimed to compare the effects of milk containing A1 β-casein with those of milk containing only A2 β-casein on inflammation, symptoms of post-dairy digestive discomfort (PD3), and cognitive processing in subjects with self-reported lactose intolerance.MethodsForty-five Han Chinese subjects participated in this double-blind, randomized, 2 × 2 crossover trial and consumed milk containing both β-casein types or milk containing only A2 β-casein. Each treatment period was 14 days with a 14-day washout period at baseline and between treatment periods. Outcomes included PD3, gastrointestinal function (measured by smart pill), Subtle Cognitive Impairment Test (SCIT), serum/fecal laboratory biomarkers, and adverse events.ResultsCompared with milk containing only A2 β-casein, the consumption of milk containing both β-casein types was associated with significantly greater PD3 symptoms; higher concentrations of inflammation-related biomarkers and β-casomorphin-7; longer gastrointestinal transit times and lower levels of short-chain fatty acids; and increased response time and error rate on the SCIT. Consumption of milk containing both β-casein types was associated with worsening of PD3 symptoms relative to baseline in lactose tolerant and lactose intolerant subjects. Consumption of milk containing only A2 β-casein did not aggravate PD3 symptoms relative to baseline (i.e., after washout of dairy products) in lactose tolerant and intolerant subjects.ConclusionsConsumption of milk containing A1 β-casein was associated with increased gastrointestinal inflammation, worsening of PD3 symptoms, delayed transit, and decreased cognitive processing speed and accuracy. Because elimination of A1 β-casein attenuated these effects, some symptoms of lactose intolerance may stem from inflammation it triggers, and can be avoided by consuming milk containing only the A2 type of beta casein.Trial registrationClinicalTrials.gov/NCT02406469.

Project description:The control of bacterial growth during milk processing is crucial for the quality maintenance of commercial milk and milk products. During a period of cold storage prior to heat treatments, some psychrotrophic bacteria grow and produce extracellular heat-resistant lipases and proteases that cause product defects. The use of lytic bacteriophages (phages) that infect and kill bacteria could be a useful tool for suppressing bacterial growth during this cold storage phase. In this study, we isolated a Pseudomonas lactis strain and a phage from raw cow's milk. Quantitative characterization of the phage was used to elucidate whether this phage was active under low temperatures and neutral pH and whether it was inactivated during pasteurization. Phage titer determination was possible under conditions ranging from pH 4 to 9 and from 3°C to 25°C; the phage was inactivated under pasteurization conditions at 63°C for 30 min. Furthermore, we showed that this phage reduced viable bacterial cell counts in both skim and whole milk. The results of this study represent the potential uses of phages for controlling psychrotrophic bacterial growth in raw cow's milk during cold storage.IMPORTANCE Suppression of bacterial growth in raw milk under cold storage is crucial for the quality control of commercially supplied milk. The use of lytic phages as low-cost microbicides is an attractive prospect. Due to strict host specificities, phages must be isolated from the raw milk where the host bacteria are growing. We first isolated the P. lactis bacterial strain and then the phage infecting that strain. Partial phage genomic analysis showed that this is a newly isolated phage, different from any previously reported. This study reports a lytic phage for P. lactis, and we have presented evidence here that this phage reduced viable bacterial cell counts not only in rich medium but also in skim and whole milk. As a result, we have concluded that the phage reported in this study would be useful in milk processing.

Project description:A major protein component of cow's milk is ?-casein. The most frequent variants in dairy herds are A1 and A2. Recent studies showed that milk containing A1 ?-casein promoted intestinal inflammation and exacerbated gastrointestinal symptoms. However, the acute gastrointestinal effects of A1 ?-casein have not been investigated. This study compared the gastrointestinal effects of milk containing A1 and A2 ?-casein versus A2 ?-casein alone in Chinese adults with self-reported lactose intolerance.In this randomised, crossover, double-blind trial, with a 3-day dairy washout period at baseline, subjects were randomised to consume 300 mL of milk containing A1 and A2 ?-casein (ratio 58:42; conventional milk) or A2 ?-casein alone; subjects consumed the alternative product after a 7-day washout period. Urine galactose was measured at baseline after a 15 g lactose load. Subjects completed 9-point visual analogue scales for gastrointestinal symptoms (borborygmus, flatulence, bloating, abdominal pain, stool frequency, and stool consistency) at baseline and at 1, 3, and 12 h after milk consumption.A total of 600 subjects were included. All six symptom scores at 1 and 3 h were significantly lower after consuming A2 ?-casein versus conventional milk (all P<0.0001). At 12 h, significant differences remained for bloating, abdominal pain, stool frequency, and stool consistency (all P<0.0001). Symptom scores were consistently lower with A2 ?-casein in both lactose absorbers (urinary galactose ?0.27 mmol/L) and lactose malabsorbers (urinary galactose <0.27 mmol/L).Milk containing A2 ?-casein attenuated acute gastrointestinal symptoms of milk intolerance, while conventional milk containing A1 ?-casein reduced lactase activity and increased gastrointestinal symptoms compared with milk containing A2 ?-casein. Thus, milk-related gastrointestinal symptoms may result from the ingestion of A1 ?-casein rather than lactose in some individuals.NCT02878876 , registered August 16, 2016. Retrospectively registered.

Dataset Information

Use of urea-polyacrylamide electrophoresis for discrimination of A1 and A2 beta casein variants in raw cow's milk.

Publications

Use of urea-polyacrylamide electrophoresis for discrimination of A1 and A2 beta casein variants in raw cow's milk.

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