Bovine κ-casein glycomacropeptide (GMP) is a sialic acid containing glycopeptide, which is considered as a health promoting compound found in cheese whey. The study described in this research communication was undertaken to determine whether GMP with undetectable level of contaminating protein or phenylalanine can be isolated from bovine whey fraction using batch anion exchange technique with chitin as an adsorbent. A soluble whey fraction (SWF) prepared from 1 g whey protein isolate (WPI) was mixed with a slurry of 1 g chitin, and the mixture was incubated at pH 3.0. After incubation, the mixture was filtered, and the residue obtained (containing chitin-GMP complex) was washed with water and eluted stepwise with 0.5 M NaCl and 2.0 M NaCl. Most of GMP (corresponding to 75.8% of total sialic acid recovered) was eluted with 0.5 M NaCl. The recovered GMP accounted for 5.4% dry weight of WPI (or 18.9% dry weight of SWF). Amino acid analysis showed that there was no detectable level of contaminating amino acids including phenylalanine, histidine, arginine and tyrosine in the GMP fraction. It was concluded that the batch anion exchange method with chitin developed in this study can be used for the isolation of high purity GMP from bovine SWF.

Casein glycomacropeptide (CGMP) is a bioactive milk-derived peptide with potential anti-inflammatory effects. Animal studies suggest that CGMP may work by altering gut microbiota composition and enhancing butyrate production. Its effects on intestinal homoeostasis, microbiota and metabolites in humans are unknown. The aim of the present study was to assess both the intestinal and systemic immunomodulatory effects of orally ingested CGMP. We hypothesised that daily oral CGMP intake would reduce high-sensitive C-reactive protein (hsCRP) in healthy adults. In a single-centre limited but randomised, double-blinded, reference-controlled study, we compared the effects of a 4-week intervention of either 25 g of oral powder-based chocolate-flavoured CGMP or a reference drink. We included twenty-four healthy adults who all completed the study. CGMP had no systemic or intestinal immunomodulatory effects compared with a reference drink, with regard to either hsCRP or faecal calprotectin level, faecal microbiota composition or faecal SCFA content. CGMP ingestion did not affect satiety or body weight, and it caused no severe adverse events. The palatability of CGMP was acceptable, and adherence was high. CGMP did not induce or change gastrointestinal symptoms. In conclusion, we found no immunomodulatory effects of CGMP in healthy adults. In a minor group of healthy adults, oral ingestion of 25 g of CGMP during 4 weeks was safe, well tolerated, had acceptable palatability and was without any effects on body weight.

It has been nearly 70 years since the discovery that strict adherence to a diet low in phenylalanine prevents severe neurological sequelae in patients with phenylalanine hydroxylase deficiency (phenylketonuria; PKU). Today, dietary treatment with restricted phenylalanine intake supplemented with non-phenylalanine amino acids to support growth and maintain a healthy body composition remains the mainstay of therapy. However, a better understanding is needed of the factors that influence N balance in the context of amino acid supplementation. The aim of the present paper is to summarise considerations for improving N balance in patients with PKU, with a focus on gaining greater understanding of amino acid absorption, disposition and utilisation. In addition, the impact of phenylalanine-free amino acids on 24 h blood phenylalanine/tyrosine circadian rhythm is evaluated. We compare the effects of administering intact protein v. free amino acid on protein metabolism and discuss the possibility of improving outcomes by administering amino acid mixtures so that their absorption profile mimics that of intact protein. Protein substitutes with the ability to delay absorption of phenylalanine and tyrosine, mimicking physiological absorption kinetics, are expected to improve the rate of assimilation into protein and minimise fluctuations in quantitative plasma amino acid levels. They may also help maintain normal glycaemia and satiety sensation. This is likely to play an important role in improving the management of patients with PKU.

Milk κ-casein-derived bovine glycomacropeptide (GMP) exerts immunomodulatory effects. It exhibits intestinal anti-inflammatory activity in chemically induced models of colitis. However, to validate its clinical usefulness as a nutraceutical, it is important to assess its effects in a model with a closer pathophysiological connection with human inflammatory bowel disease. Therefore, in the present study, we used the lymphocyte-transfer model of colitis in mice and compared the effects of GMP in this model with those obtained in the dextran sulphate sodium (DSS) model. GMP (15 mg/d) resulted in higher body-weight gain and a reduction of the colonic damage score and myeloperoxidase (MPO) activity in Rag1− / − mice with colitis induced by the transfer of naïve T cells. The colonic and ileal weight:length ratio was decreased by approximately 25 %, albeit non-significantly. GMP treatment reduced the percentage of CD4+ interferon (IFN)-γ+ cells in mesenteric lymph nodes (MLN). The basal production of IL-6 by MLN obtained from the GMP-treated mice ex vivo was augmented. However, concanavalin A-evoked production was similar. The colonic expression of regenerating islet-derived protein 3γ, S100A8, chemokine (C-X-C motif) ligand 1 and IL-1β was unaffected by GMP, while that of TNF-α and especially IFN-γ was paradoxically increased. In the DSS model, GMP also reduced the activity of colonic MPO, but it failed to alter weight gain or intestinal weight:length ratio. GMP augmented the production of IL-10 by MLN cells and was neutral towards other cytokines, except exhibiting a trend towards increasing the production of IL-6. The lower effect was attributed to the lack of the effect of GMP on epithelial cells. In conclusion, GMP exerts intestinal anti-inflammatory effects in lymphocyte-driven colitis.

Caseinomacropeptide (CMP), a 7-kDa phosphoglycopolypeptide fragment released from κ-casein during milk renneting, is heterogeneous with respect to post-translational glycosylation. Several studies have reported that CMP has growth-promoting activity on lactic acid bacteria belonging to the genera Bifidobacterium. The aim of this study was to evaluate the effect of glycosylation and sequence variations between bovine and caprine CMP on the growth of two probiotics: Lactobacillus rhamnosus RW-9595-M and Bifidobacterium thermophilum RBL67. The growth-promoting activities of CMP (mixture of glycosylated (gCMP) and non-glycosylated (aCMP) fractions), aCMP and gCMP were measured in a basal minimal culture medium using turbidimetric microplate assay at 37 °C. Supplementation of the culture media at 2 mg/ml significantly improved maximum growth by 1·5 to 1·8 times depending on the strain, the additive (CMP, aCMP, gCMP), and the bovine or caprine origin (P < 0·05). CMP preparations also decreased the time needed to reach the inflexion point of the growth curve and increase the cell density at that time (P < 0·05). The effects of CMP preparations were dose dependent and significantly superior to the effect of bovine β-lactoglobulin added to the culture media. As gCMP and aCMP were as efficient as bovine and caprine CMP (P > 0·1), it was concluded that the presence of oligosaccharides linked to CMP was not essential for growth-promoting activity of CMP.

Sialic acid, being a biologically active compound, is recognised as an important component of milk and milk products. Almost all the sialic acid estimation protocols in milk require prior hydrolysis step to release the bound sialic acid followed by its estimation. The objective of this work was to estimate sialic acid in milk and milk products by fluorimetric assay which does not require a prior hydrolysis step thus decreasing the estimation time. The recovery of added sialic acid in milk was 91·6 to 95·8%. Sialic acid in milk was found to be dependent on cattle breed and was in the range of 1·68–3·93 g/kg (dry matter basis). The assay was further extended to detect adulteration of milk with sweet whey which is based on the detection of glycomacropeptide (GMP) bound sialic acid in adulterated milk. GMP is the C-terminal part of κ-casein which is released into the whey during cheese making. For detection of adulteration, selective precipitation of GMP was done using trichloroacetic acid (TCA). TCA concentration in milk was first raised to 5% to precipitate milk proteins, especially κ-casein, followed by raising the TCA concentration to 14% to precipitate out GMP. In the precipitates GMP bound sialic acid was estimated using fluorimetric method and the fluorescence intensity was found to be directly proportional to the level of sweet whey in adulterated milk samples. The method was found to detect the presence of 5% sweet whey in milk.

Glycomacropeptide (GMP) is the hydrophilic 64-amino acid C-terminal glycopeptide released into cheese whey when κ-casein is cleaved by chymosin. GMP exists as a mixture of different glycoforms due to the carbohydrates sialic acid (N-acetylneuraminic acid, NeuNAc), galactose (Gal), galactosamine and glucosamine attached by O-glycosidic linkages. GMP reportedly stimulates the release of cholecystokinin (CCK), which may promote satiety. The objectives of the present study were to manufacture three glycoforms of GMP, minimally glycosylated GMP (3·5 (sd 0·1) % NeuNAc and 1·5 (sd 0·1) % Gal), glycosylated GMP (12·0 (sd 0·3) % NeuNAc and 4·2 (sd 0·2) % Gal) and a GMP-depleted whey protein concentrate, and to assess the effects of these fractions relative to glucose on CCK, subjective measures of satiety and food intake. In a randomised double-blind acute study, twenty overweight/obese males (56·9 (sd 7·2) years, 97·4 (sd 8·1) kg, 31·5 (sd 3·0) kg/m2) were recruited to consume four 50 g preloads (two GMP preparations, GMP-depleted whey and glucose) containing 895 kJ. Blood samples and subjective measures of satiety were collected before and at 15, 30, 60, 90, 120 and 180 min after the consumption of preload, and CCK levels were measured. A lunchtime meal of hot food was provided from which subjects ate ad libitum until satisfied. Energy and nutrient intakes from the food consumed were calculated. There was no significant difference in CCK levels, subjective measures of satiety or food intake between treatments at the given preload level. These results suggest that the protein fractions at the dose employed do not influence satiety, CCK levels or energy intake at a subsequent meal.

The effect of feed protein type on body composition and growth has been examined. Evidence exists that whey protein concentrate is effective at limiting body fat expansion. The presence of caseinomacropeptide, a mixture of glycosylated and non-glycosylated carbohydrate residues, in particular glycomacropeptide (GMP) in whey protein concentrate may be important for this effect. The influence of whey protein isolate (WPI) and GMP on weight gain and body composition was examined by feeding Wistar rats ad libitum for 7 weeks with five semi-purified American Institute of Nutrition-based diets differing in protein type: (1) casein; (2) barbequed beef; (3) control WPI (no GMP); (4) WPI+GMP at 100 g/kg; (5) WPI+GMP at 200 g/kg. Body composition was assessed, and plasma samples were assayed for TAG, insulin and glucose. Body-weight gain was lower ( − 21 %) on the control WPI diet relative to casein, with a non-significant influence associated with GMP inclusion ( − 30 %), the effect being equivalent at both levels of GMP addition. Renal and carcass fat mass were reduced in the highest GMP diet when compared with WPI (P < 0·05). Plasma insulin was lowered by GMP at the highest addition compared with WPI alone ( − 53 %; P < 0·01). Plasma TAG in the WPI+GMP (200 g/kg) group were lower ( − 27 %; P < 0·05) than the casein and beef groups. In conclusion, GMP appears to have a significant additional influence when combined with WPI on fat accumulation. WPI alone appears to have the predominant influence accounting for 70 % of the overall effect on body-weight gain. Mechanisms for this effect have not been identified but food intake was not responsible.