Determination Of The Types Of Peptides And The Anti-inflammatory Activities Obtained From Bovine&Caprine Colostrums And Fermented Milk

Recently, the study of the functional ingredients from food has become a popular field ofstudy. The bioactive peptides, especially the small molecular weight peptides that havefunctional characteristics get more and more people’s attention. The bioactive peptides weresmall amino acid sequences encrypted in milk proteins can be released by enzymaticproteolysis or gastrointestinal digestion. These peptides can exert a wide range of effects andthese have been confirmed by many researchers, such as antimicrobial, antihypertensive,antithrombotic, immunomodulatory, and opioid properties, in addition to aiding in themineral absorption of calcium. Inflammatory diseases significantly impact daily life withrespect to human health, there have been few reports of functional peptides from colostrumsand fermented milk with anti-inflammatory activities. The goal of our study was to determinethe types of peptides purified from bovine&caprine colostrums and their fermented productswith different strains of fermentation using an AKTA purifier10and to investigate thepeptide fractions obtained from the milk samples. The anti-inflammatory activities ofpeptides released from the AKTA milk sample fractions were determined using competitiveTNF-α ELISA and Western blot methods. To determine which types of peptides may havebeneficial effects on human health, the samples were all subjected to simulate humangastrointestinal digestion before being purified using an AKTA. Finally, the liquidchromatography-mass spectrometry was applied to the selected optimal anti-inflammatorypolypeptide to determine the amino acid sequence and the molecular weight, which provide areference for the development of the anti-inflammatory functional food. The mainconclusions were as follows:(1) There are more types of peptides purified from the caprine colostrum than the bovinecolostrums. The fermented milk which are fermented by lactobacillus acidophilus andbifidobacterium compound strains obtained the most types of peptides, in addition, thepeptides in caprine fermented milk are more than in bovine. There are23types of peptidespurified from the bovine colostrums and36types of peptides from the caprine colostrums. There are more types of peptides by using the probiotic strains than the ordinary strains.There are3types of peptides purified from the ordinary strains caprine fermented milk and1type from bovine fermented milk. There are2types of peptides purified from thelactobacillus acidophilus strains bovine fermented milk and4types from caprine fermentedmilk. There are2types of peptides purified from the bifidobacterium strains bovine milk and3types from caprine fermented milk.(2)Using the competitive TNF-α ELISA method to detect the anti-inflammatory, all thepeptides purified from the milk samples show the anti-inflammatory function. The meanvalues for the concentration of TNF-α increased from (2.42±0.006)ng/L to (18.03±0.102)ng/Lin all of the bovine and caprine colostrum samples. The mean values for the concentration ofTNF-α increased from (2.18±0.069)ng/L to (14.03±0.081)ng/L in all of the bovine andcaprine fermented samples.(3) We performed western blot experiments to further determine whether these peptideshad anti-inflammatory activities. The result showed that: Peptide C3-1(one in the third daycaprine colostrums) has potential benefits as an anti-inflammatory supplement formaintaining good health and for the possible prevention of inflammatory diseases; PeptideC①-2(one in the lactobacillus acidophilus and bifidobacterium compound strains caprinefermented milk) has potential benefits as an anti-inflammatory supplement for maintaininggood health and for the possible prevention of inflammatory diseases.(4) The liquid chromatography-mass spectrometry was applied to the selected optimalanti-inflammatory polypeptide to determine the amino acid sequence and the molecularweight. From the analysis of TNF-α ELISA and Western blot, peptide B3-1is selected and itsamino acid sequence maybe is FQSEEQQQTEDELQDK, the molecular weight is1983Da,and it maybe released from beta-casein precursor (Bovine). Peptide C3-1is also selected andits amino acid sequence maybe is VYVEELKPTPEGNLEILLQK, the molecular weight is2315Da, and it maybe released from beta-lactoglobulin (Caprine). From the analysis ofTNF-α ELISA and Western blot, peptide B②-1is selected and its amino acid sequencemaybe is HQGLPQEVLNENLLR, the molecular weight is1761Da, and it maybe releasedfrom alpha-S1-casein precursor [contains: Antioxidant peptide](Bovine). Peptide C①-2isalso selected and its amino acid sequence maybe is SPAQTLQWQVLPNTVPAK, themolecular weight is1979Da, and it maybe released from kappa-casein Ovis dalli (Dallsheep).