Goat Milk Protein Research Articles

Production of goat milk protein hydrolysate enriched in ACE-inhibitory peptides by ultrafiltration.

A global process for the production of goat milk hydrolysates enriched in angiotensin converting enzyme (ACE) inhibitory peptides was proposed. Firstly, the protein fractions (caseins and whey proteins) were separated by ultrafiltration through a 0·14μm ceramic membrane. The casein fraction obtained in the retentate stream of the above filtration step was subsequently hydrolysed with a combination of subtilisin and trypsin. After 3h of reaction, the hydrolysate produced presented an IC50 of 218·50μg/ml, which represent a relatively high ACE inhibitory activity. Finally, this hydrolysate was filtered through a 50kDa ceramic membrane until reaching a volume reduction factor of 3. The permeate produced presented an improvement of more than 30% in the ACE inhibitory activity. In contrast, the retentate was concentrated in larger and inactive peptides which led to a decrease of more than 80% in its inhibitory activity. The process suggested in this work was suitable to obtain a potent ACE inhibitory activity product able to be incorporated into food formulas intended to control or lower blood pressure. Moreover, the liquid product could be easily stabilised by spray dried if it would be necessary.

Perioperative and peripartum prevention of venous thromboembolism in patients with hereditary antithrombin deficiency using recombinant antithrombin therapy

Recombinant human antithrombin (rhAT; ATryn), isolated from the milk of transgenic goats, provides an alternative to human plasma-derived antithrombin (AT) concentrate for perioperative and peripartum prophylaxis of venous thromboembolism (VTE) in patients with hereditary AT deficiency. Optimized rhAT dosing algorithms and improved plasma AT monitoring protocol were used in an open-label, single-arm, multinational, pivotal safety and efficacy study that was conducted in patients with hereditary AT deficiency in perioperative and peripartum settings. Loading and maintenance doses were calculated on the basis of pretreatment AT activity levels. Specific dosing regimens were used for pregnant and surgical patients; rhAT was to be given for at least 3 days and for 14 days or less. The primary efficacy end point was the incidence of any thromboembolic event during rhAT therapy or within 7 days of rhAT discontinuation. Safety and AT activity levels were secondary end points. Six surgical and 12 pregnant patients were treated for a median of 3.2 days (range 0.9-14 days). With the optimized dosing regimens, a median of 1 dose adjustment (range 0-6 dose adjustments) was needed to maintain AT activity levels within 80-120% of normal. No confirmed VTEs occurred during treatment or in the subsequent 7 days. Overall, rhAT was well tolerated, but some bleeding complications occurred after rhAT discontinuation and anticoagulation reinstitution. No antibodies to rhAT or goat milk proteins were detected. Perioperative and peripartum prophylactic rhAT therapy in patients with hereditary AT deficiency is well tolerated and effective in preventing VTE.

Effect of heat and transglutaminase on solubility of goat milk protein‐based films

The effect of heat, transglutaminase and combination of heat and transglutaminase treatments on the solubility of films prepared from goat milk casein, goat milk whey proteins and whole goat milk proteins was investigated. Goat milk casein films were less soluble when treated with transglutaminase and combination of heat with transglutaminase compare with heat‐treated caseins alone. Heat treatment was more effective at decreasing the solubility of whey protein films. SDS‐PAGE patterns demonstrated that goat milk caseins were better cross‐linked by transglutaminase, whereas whey proteins were better cross‐linked by heat. The extent of cross‐linking was further enhanced when a combination of heat and transglutaminase was used.

Antioxidant peptides from goat milk protein fractions hydrolysed by two commercial proteases

Goats' milk microfiltration fractions were hydrolysed with subtilisin or trypsin, or both, and tested for 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) radical scavenging capacity, iron chelation capacity, and inhibition of secondary oxidation products formation in liposomes. The retentate treated with subtilisin was most active regarding radical scavenging capacity (SC50 ≈ 4 μg mL−1), while the permeate treated with subtilisin exhibited the best iron chelation capacity (IC50 ≈ 65 μg mL−1) and prevention of secondary lipid oxidation products formation (33% inhibition at 25 μg mL−1). In the retentate hydrolysate various active peptides were identified. Tyrosine seemed fundamental in the ABTS radical scavenging capacity of the peptides, and also to play a role in the inhibition of formation of secondary lipid oxidation products, in which phenylalanine seemed to play the key role. Non-protein compounds in the permeate hydrolysate seemed more important than peptides for the antioxidant activities detected.

EXPERIENCE OF USAGE OF ADAPTED GOAT MILK FORMULA IN NUTRITION OF HEALTHY INFANTS DURING FIRST SIX MONTHS OF LIFE

Background: The authors had a goal to assess clinical efficacy of adapted goat milk formula and its possibilities for usage in nutrition of infants during first six months of life. Patients and methods : Seventy three healthy infants at the age from 1 to 4 months were followed-up, among them 42 — on artificial feeding and received the investigated formula, and 31 — breast-fed (comparative group). The duration of follow-up was 30–40 days. The criteria of efficacy of the investigated formula were: tolerability, dynamics of weight and height and laboratory characteristics (hemoglobin concentration in erythrocytes and equivalent of hemoglobin in reticulocytes, concentrations of blood iron, ferritin, prealbumin, active metabolites of vitamin D3 (25OH) in capillary blood, revealing of specific immunoglobulins E to cow and goat milk proteins with the help of express-method and coprological test), assessment of actual nutrition of infants with calculation of the main macro- and micronutrients, energetic value of the diet and protein efficacy coefficient. Results : The investigated children adapted goat milk formula meets all the modern criteria according to the composition of such type of food products, provides physiological demands of infants in the main nutrients and energy, maintains their normal physical development and normalizes stool in tendency to constipations. Conclusion : The investigated milk formula can be used in nutrition of non-breastfed infants during first 6 months of life.

Comparative Protein Composition Analysis of Goat Milk Produced by the Alpine and Saanen Breeds in Northeastern Brazil and Related Antibacterial Activities

The protein composition of goat milk differs between goat breeds and could present regional trends. The aim of this study was to comparatively analyze the protein composition of goat milk produced by the Alpine and Saanen breeds in northeastern Brazil and to evaluate the antibacterial activity of its protein fractions. SDS-PAGE, 2-DE electrophoresis and RP-HPLC analyses revealed the absence of αs1-casein in the milk of both breeds and no differences between the αs2-casein, β-casein, β-lactoglobulin and α-lactalbumin profiles. The amounts of soluble proteins and β-casein hydrolysis residues were higher in Saanen milk. Only the protein fraction containing the largest amounts of casein (F60–90%) inhibited bacterial growth, with MIC values between 50 and 100 mg/mL. This study describe for the first time three important points about the goat milk protein of two Brazilian goat breeders: absence of α-s1 casein in the protein profile, differences between the milk protein composition produced by goats of Alpine and Saanen breeders and antibacterial activity of unbroken proteins (casein-rich fraction) present in these milk.

P97 ‐ Characteristic of protein sensibilisation in infants

Cow milk protein sensibilisation is common in bottle-fed infants. It is known about immunological cross-reaction for cow and goat milk protein. We examined 85 infants (39 girls, 46 boys), 1.5 – 18 months old, who were fed by artificial milk formulas. Gastrointestinal symptoms of food allergy were diagnosed in 45 (52.9%) children with diarrhea and in 40 (47.1%) children with constipation. The control group consisted of 25 healthy infants of the same age. The level of total IgE, cow and goat milk proteins IgE allergenspecific antibodys in coprofiltrates were measured by immunoenzymometric method using the spectrophotometer “Sunrise” (Belgium), with test-systems “Allergopharma” and “Dr. Fooke” (Germany). The highest rate of sensibilisation to cow milk protein (89%) was found in infants of 1.5-6.0 months old compared to infants of 6.5-12.0 (75%) and 12.5-18 months old (56%). Increased level of total IgE in coprofiltrates was more common in infants of 1.5-5.5 months old, too, compared to infants of other ages (33%, 25% and 22%, respectively). The sensibilisation to goat milk protein was more common in infants of 6.5-12 months old (59%) and 12.5-18.0 months old (45%) compared to younger (1.5-6.0 months old) infants (23%). In infants of any ages sensibilisation to cow milk protein was more frequent than to goat milk proteins. Increasing rate of sensiblisation to goat milk protein in bottle-fed infants more than 6 months old, probably, depends on immunological cross-reaction for cow milk protein.

P41 ‐ Preexposure prophylaxis of infants’ food allergy

Methods We observed 100 “mother and child” pairs, which were divided into two groups. All children had AD, associated to cow milk protein (CMP) allergy. Lactating mothers from both groups were treated by diet therapy with the replacement of the cow milk to the New Zealand goat milk (1 group: 43 “mother and child” pairs) or non-dairy diet (2 group: 57 “mother and child” pairs). The effect of diet therapy was assessed by dynamics of clinical and immunological AD symptoms in infants after 1-3 months of treatment. Immunological effect of the therapy was assessed by the dynamics of levels of allergen-specific IgE and IgG to CMP, casein, b-lactoglobulin and goat milk protein which were measured by uncompetitive immunoenzyme assay on special test-systems from Allergopharma (Germany). The level of IFN-g, IL-2, IL-4, IL-5, IL-12, IL-13 was measured by immunoenzyme ELISA method.

Cross-Desensitization To Goat and Sheep Milk Protein In Cow's Milk Protein Desensitized Patients

Over 90% cow’s milk protein (CMP) allergic patients cross react to both goat’s milk protein (GMP) and sheep’s milk protein (SMP) due to shared kappa-casein protein epitopes. We investigated whether CMP allergic patients who have been successfully fully desensitized in an oral immunotherapy (OIT) program would be able to consume GMP and SMP without an allergic reaction.

Major proteins in goat milk: an updated overview on genetic variability

Milk and dairy products are very important in Mediterranean diet because of their health promoting and organoleptic properties. In many developing countries, goat rearing has a key role in livestock production. What makes goats so popular is their ability to provide high quality food under diverse climatic conditions and resilience to extreme and capricious environments. In the last years, the interest concerning caprine milk has been increasing also to find a new exploitation for local breeds. To promote the goat dairy products there is a clear need to know the quality and the technological aspects of milk produced. That being so, the purpose of this study was to review the available literature on the major goat milk proteins with a particular attention to recent findings on their genetic variability. Moreover, the main effects of different protein variants on milk yield and composition were also discussed.

Preexposure prophylaxis of infants’ food allergy

Background. Till the present time the problem of food allergy remains to be the topical subject and demands further development of therapeutical and prevention programs, including diet therapy for nursing mothers. The work is aimed at clinical and immunological evaluating of diet therapy of mothers whose children have atopic dermatitis on the back of breast feeding only. Methods. There were 100 «mother and child» couples kept under observance, that were divided into two groups. All the children had atopic dermatitis (AD), associated to cow milk protein allergy. Nursing mothers from both the groups were treated by diet therapy according to which the cow milk in diet have been replaced either with goat milk of New Zealand origin (first group had 43 «mother and child» couples) or with nondairy products (second group had 57 «mother and child» couples). Effectiveness of diet therapy of nursing mothers has been evaluated by the dynamic of clinical and immunological symptoms of atopic dermatitis in 1-3 months from treatment beginning. Immunological effectiveness of the treatment has been evaluated by the dynamic of indexes of specific IgE and IgG antibodies to cow milk protein, β-lactoglobulin, casein, soy protein and goat milk protein and cytokines levels. Results. In the course of treatment, that included diet therapy of nursing mothers and antiallergic cure of infants, both the groups showed remission and saved breast feeding. Immunological evaluating of therapeutic intervention’s effectiveness revealed the improvement of total IgE, allergenspecific IgE and IgG antibodies to cow milk protein and its fractions, as well as to soy and goat milk protein in blood of all the children on the back of breast feeding. Moreover, complex therapy conduced decreasing of pro-inflammatory cytokines concentration in blood. Conclusion. These findings prove reasonability of dietary treatement for nursing mothers, whose children have atopic dermatitis, associated associated to cow milk protein allergy.

Spray Drying of Goat Milk Protein Hydrolysates with Angiotensin Converting Enzyme Inhibitory Activity

A goat milk hydrolysate was stabilized by spray drying using different inlet air temperatures (170–230 °C) and feed flow rates (4–12 mL/min) following a central composite experimental design. In order to evaluate the effect of operational conditions on process yield, powder properties (density, moisture, hygroscopicity, and solubility) as well as angiotensin converting enzyme (ACE) inhibitory activity, experimental data were analyzed by response surface methodology. Input variables showed a significant influence on yield, density, and moisture, while hygroscopicity, solubility, and ACE inhibitory activity were not affected. The dried hydrolysate presented an average IC50 value of 273.13 μg/mL, which involved a loss of 25 % in ACE inhibitory activity with respect to the feed. The variations detected in the low molecular weight fractions of the dried hydrolysates could be responsible for the variations in ACE inhibitory activity.

Algorithm of the choice of nutritional therapy in infants with food allergy

Methods We examined 50 formulae fed infants 1-12 months old with food allergy. The examination included a collection of allergy history, clinical evaluation and specific methods – the measurement of the coprofiltrate levels of total IgE and allergen-specific IgE to the cow milk protein and casein. The level of total IgE, cow and goat milk proteins IgE allergen-specific antibodies in coprofiltrates were measured by immunoenzyme method using the spectrophotometer “Sunrise” (Belgium), with testsystems “Allergopharma” and “Dr. Fooke” (Germany). The nutritional therapy of the food allergy in infants consists in replacement of cow milk protein formula on the formula based on the goat milk. The estimation of the efficiency of therapy based on dynamics of the clinical symptoms of the disease and decrease of the serum levels of total IgE and allergen-specific IgE to the cow milk proteins.

Angiotensin I-converting enzyme inhibitory activity of enzymatic hydrolysates of goat milk protein fractions

Casein and whey protein fractions from goat milk were hydrolysed by subtilisin and trypsin, individually and in combination, to release angiotensin converting enzyme (ACE)-inhibitory peptides. Selected hydrolysates were fractionated by size exclusion chromatography (SEC) and further characterised. The highest ACE-inhibitory activity was obtained from the casein fraction hydrolysed by the combination of enzymes. SEC presented 4 fractions with fraction F2 (<2.3 kDa) containing the highest concentration of peptides and the highest activity. F2 contained a number of peptides not previously identified from caprine caseins but with structural similarity to other ACE-inhibitory peptides. The most active fraction in relation to protein content was F4 with IC50 between 9.3 and 5.1 μg mL−1. This fraction contained a compound tentatively identified as WY, an active dipeptide not previously reported from caseins. The high inhibitory capacity of these fractions points towards the advantage of implementing a membrane process to concentrate the most active peptides.

Optimisation of the hydrolysis of goat milk protein for the production of ACE-inhibitory peptides

Goat milk protein was hydrolysed with subtilisin and trypsin. As input variables, temperature was assayed in the interval 45-70 °C for subtilisin and 30-55 °C for trypsin, while the enzyme-substrate ratio varied from 1 to 5%. The effect of the input variables on the degree of hydrolysis and ACE-inhibitory activity (output variables) was modelled by second order polynomials, which were able to fit the experimental data with deviations below 10%. The individual maximum values of the degree of hydrolysis and the ACE-inhibitory activity were found at conflicting conditions of temperature and enzyme-substrate ratio. Since such maximum values could not be reached simultaneously, a bi-objective optimisation procedure was undertaken, producing a set of non-inferior solutions that weighted both objectives.

Proteomic analysis of differentially expressed proteins in caprine milk during experimentally induced endotoxin mastitis

The goal of the current study was to identify proteins in goat milk before and at 18h following intramammary challenge with lipopolysaccharide (LPS). Initial evaluation of protein profiles generated using 2-dimensional gel electrophoresis on skim milk samples from a group of 6 goats collected before challenge and at 18, 24, and 48h after LPS challenge revealed little change in the abundance of casein proteins, and minimal changes in the presence or abundance of the plasma protein serum albumin, which is known to leak into milk during coliform mastitis in dairy cattle. Proteins in baseline milk samples and in milk from the same goats 18h post-LPS challenge were excised from the gels, and peptides were sequenced using nano-flow liquid chromatography coupled with tandem mass spectrometry. Despite the overwhelming presence of casein proteins and β-lactoglobulin, the lower abundance proteins β-2-microglobulin, fatty acid-binding protein, serum albumin, and retinol-binding protein were detected in skim milk samples from healthy goats. Skim milk samples 18h postchallenge were characterized by the sustained presence and abundance of the casein proteins, and by the presence of haptoglobin, serum amyloid A, lactoferrin, cathelicidin-1, and cathelicidin-3. No marked differences in the intensity of the spot corresponding to serum albumin were observed in gels of skim milk samples 18h postchallenge, which could indicate that the breakdown of the blood–milk barrier during endotoxin mastitis may not be as profound in goats as has been observed in dairy cattle. Nonetheless, the occurrence of an inflammatory response was supported by elevated somatic cell counts in the goat milk following inoculation with endotoxin, as well as by the presence of both antimicrobial and acute phase proteins. The results provide information about the composition of proteins in goat milk as well as added knowledge of the host response during endotoxin mastitis in goats.

Good tolerance to goat's milk in patients with recurrent aphthous ulcers with increased immunoreactivity to cow's milk proteins

Recurrent aphthous ulcers (RAU) represent a very common, but poorly understood mucosal disorder. The connection between immunity to cow's milk proteins (CMP) and oral diseases was noted earlier. The goal of this study was to determine the prevalence of the increased levels of serum antibodies to goat's milk proteins (GMP), by enzyme-linked immunosorbent assay (ELISA) test, in subjects who have RAU and proven increased immunity to CMP. Fifty subjects with RAU (36 with proven increased immunity to CMP and 14 without this increased immunity) were included in this research. Levels of serum IgA, IgG, and IgE antibodies to the same quantity of the examined antigens were determined by ELISA. The statistical analysis of data was performed by Wilcoxon rank-sum test and Mann-Whitney test. The levels of serum antifresh cow's milk IgA, IgG, and IgE antibodies were significantly higher than the levels of serum antifresh goat's milk, in subjects with RAU with proven increased immunoreactivity to CMP (P = 0.0003; P < 0.0001; P < 0.0001). These results indicate that patients with RAU with increased immunity to CMP could consider the use of goat's milk as the alternative protein source.

The dynamics of soluble apoptosis markers during diet therapy in infants with atopic dermatitis

Background. To estimate the dynamics of soluble apoptosis markers in infants with atopic dermatitis for updating mechanisms of immunopathogenesis and improvement of diet therapy. Methods. We observed 66 bottle-fed infants aged 1,5—12 months old (boys -47, girls — 19) with atopic dermatitis (AD). The sensibilization to cow milk protein was revealed in all 66 infants. Detected allergen-specific IgG and IgE antibodies to cow milk protein, its fraction and goat milk protein were the reason to include infants into the 1st group and feed with hydrolyzed formula (27 infants). 39 infants in the 2nd group, who were not sensibilized to goat milk protein, were fed by goat milk based formula. Serum levels of soluble apoptosis markers (sCD153, caspase-8, sFas-L, caspase-9 and annexin-5) were measured by immunoenzyme method (ELISA). Results. The activation of signal apoptosis systems in infants with AD with increased levels of sFas-L и sCD153 was revealed. Levels of caspase-8 and caspase-9 were significantly lower than in control group, and reflected the impaired elimination of modified immmunocompetent cells. The level of annexin-5 was significantly lower in infants with AD than in control group. The estimation of the dynamics of investigated parameters during diet therapy showed significant increase of caspase-9 level in both groups. The level of caspase-8 was increased only in infants who were fed by goat milk formula. Levels of sFas-L, sCD153 and annexin-5 during diet treatment did not differ significantly between groups. Conclusion. The results showed that sCD153, caspase-8, sFas-L, caspase-9 and annexin-5 play a role in the realization of allergic inflammation in infants with AD. The diet therapy with goat milk formula promotes more physiological repair of the effectory component of the apoptosis.

Comparative evaluation of allergic sensitization to milk proteins of cow, buffalo and goat

Milk hypersensitivity due to adverse reactions to milk proteins is among the major public concern of food allergies. In present investigation, comparative assessment of humoral and cell mediated immune responses in mice associated with allergenicity of major milk proteins (casein and β-lactoglobulin) of cow, buffalo and goat were carried out. Caseins of all three species of dairy animals appeared to be less immunogenic and allergenic than their corresponding β-lactoglobulins. Cow milk proteins (caseins and β-lactoglobulin) were observed to increase protein specific IgE sensitization as well as lymphocyte proliferation index significantly (P<0.01) as compared to buffalo and goat milk proteins. On the other hand, buffalo and goat milk proteins similar to cow's proteins raised remarkably higher (P<0.01) immunogenic response in terms of IgG levels than the negative control group. The results obtained were correlated with B and T cell epitopes of milk proteins cited in literature after multiple alignment in silico. Thus present study suggests cow milk proteins (β-lactoglobulin and casein) are highly allergenic among the milk proteins of three species of dairy animals.

Production of transgenic goats expressing human coagulation factor IX in the mammary glands after nuclear transfer using transfected fetal fibroblast cells

There are growing numbers of recombinant proteins that have been expressed in milk. Thus one can consider the placement of any gene of interest under the control of the regulatory elements of a milk protein gene in a dairy farm animal. Among the transgene introducing techniques, only nuclear transfer (NT) allows 100% efficiency and bypasses the mosaicism associated with counterpart techniques. In this study, in an attempt to produce a transgenic goat carrying the human coagulation factor IX (hFIX) transgene, goat fetal fibroblasts were electroporated with a linearized marker-free construct in which the transgene was juxtaposed to β-casein promoter designed to secret the recombinant protein in goat milk. Two different lines of transfected cells were used as donors for NT to enucleated oocytes. Two transgenic goats were liveborn. DNA sequencing of the corresponding transgene locus confirmed authenticity of the cloning procedure and the complementary experiments on the whey demonstrated expression of human factor IX in the milk of transgenic goats. In conclusion, our study has provided the groundwork for a prosperous and promising approach for large-scale production and therapeutic application of hFIX expressed in transgenic goats.