By Kelapure N.N and Chavan V.R.

Introduction

Recent research has shown that Milk proteins are the primary source of bioactive peptides, which can be encrypted within the amino acid sequence of dairy proteins. These peptides are inactive within the sequence of the parent protein molecule and can be liberated by (1) gastrointestinal digestion of milk, (2) fermentation of milk with proteolytic starter cultures or (3) hydrolysis by proteolytic enzymes. Biological activities associated with such peptides include immunomodulatory, antibacterial, anti-hypertensive and opioid-like properties. On an industrial scale various technologies developed for the production and isolation of bioactive milk peptides. These technologies are based on novel membrane separation and ion exchange chromatographic methods such as electro-membrane filtration, and reversed phase HPLC being employed by the emerging dairy ingredient industry. A variety of naturally formed bioactive peptides have been found in fermented dairy products, such as yoghurt, sour milk and cheese. In particular, antihypertensive peptides have been identified in fermented milk, whey and ripened cheese.  Hence it is beneficial to use these bioactive peptides in the formulation and improvement of future functional foods and as potent drugs with well-defined pharmacological effects. 

  • Milk is considered as a most perfect food.
  • Milk proteins are complex food constituents.
  • Mainly consists of casein (α, β, γ fractions) & whey protein (β-lactoglobulin,α-lactoalbumin).
  • Proteins exert their physiological action either directly or upon enzymatic hydrolysis in vitro or in vivo.
  • Peptides represent a quite heterogeneous class of compounds and their characteristics deeply depend on the amino acidic composition and on the length of the chain.
  • Low molecular weight peptides are also less allergenic than native proteins; therefore, milk protein hydrolysates are commonly utilized to formulate hypoallergenic food for infants.
  • In food matrices containing sugars, some peptides can undergo the Maillard reaction by the heating process, thus modifying the appearance of the product; in milk, lactose and lysine residues in proteins (mainly in caseins) can give rise to a series of undesirable brown pigments and aromatic compounds which weigh upon the colour and the flavour of heated milk.

    Bioactive peptides

    • Dietary proteins provide a rich source of biologically active peptides.
    • Bioactive peptides are defined as the specific protein fragments that have a positive impact on body functions or conditions and may ultimately influence health.
    • Peptides represent a more bioavailable form of essential amino acids.
    • Act as Nutraceuticals.
    • Bioactive peptides can be latent (or encrypted) within the primary or parent proteins.
    • Proteolysis is required for their release and activation to exert a physiological response on the various systems in the body.
    • Bioactive peptides from milk are mainly released from (casein, whey proteins)
    • Bioactive peptides possess antimicrobial, antihypertensive, antioxidative, anticytotoxic, immunomodulatory, opioid, and mineral-carrying properties.

      Production of bioactive peptides

      • Bioactive peptides are inactive within the sequence of the parent protein and can be released in three ways:
      • Enzymatic hydrolysis by digestive enzymes
      • Food processing
      • Hydrolysis by proteolytic microorganisms or their enzymes

      Food Processing

      • Heat or alkali treatment can generate inter-and intramolecular covalent bonds that are resistant to hydrolysis.
      • Lead to the formation of indigestible peptides.
      • It promotes both the formation and absorption of bioactive peptides that do not occur naturally in the precursor protein.
      • Partially hydrolyzed milk proteins are used in hypoallergenic infant formulae and for clinical applications.
      • Cheese contains phosphopeptides as natural constituents and secondary proteolysis during cheese ripening leads to the formation of various ACE inhibitory peptides.

      Microbial fermentation

      • Bioactive peptides can be generated by the proteolytic activities of the strains of starter and non-starter bacteria e.g. Lactobacillus helveticus, Lactobacillus delbrueckii bulgaricus, Lactobacillus plantarum, Lactobacillus rhamnosus, Lactobacillus acidophilus, Lactococcus lactis, Streptococcus thermophilus used in the manufacture of fermented dairy products.
      • The proteolytic system of lactic acid bacteria (LAB) is well characterized.
      • This system consists of a cell wall-bound proteinase and a number of distinct intracellular peptidases, including endopeptidases, aminopeptidases, tripeptidases and dipeptidases.
      • Proteinases cause the degradation of casein into oligopeptides.
      • Longer chain oligopeptides may be a source of bioactive peptides when further degraded by intracellular peptidases.
      • ACE-inhibitory or antihypertensive peptides (VPP, IPP), immunomodulatory, antioxidative, antimutagenic peptides.

      Isolation of bioactive peptides

      • The bioactive peptides can be isolated using Electro-membrane filtration (EMF)
      • Peptides or proteins are strongly charged compounds.
      • EMF combines conventional membrane filtration with electrophoresis.
      • EMF is used for the isolation of positively charged peptides, especially the bioactive peptide αs2-casein fragment f(183–207), from an αs2-casein (αs2-CN) hydrolysate.
      • CMX and AMX ion-exchange membranes used to prevent direct contact between feed and product with the electrodes
      • Ultrafiltration polysulphone membranes with MWCO between 10 kDa and 100 kDa & membrane area of either 0.008 m2 -0.010 m2 used.

           Sources of Bioactive Peptides

      • Milk contains a wide range of proteins that provide protection against enteropathogens or are essential for the manufacture and characteristic nature of certain dairy products.
      • Peptides are in a latent or inactive state within protein molecules but can be released during enzymatic digestion.
      • Biologically active peptides released from caseins and whey proteins contain 3 to 20 amino acids per molecule.
      • Bioactive peptides (BPs) have been identified within the amino acid sequences of native milk proteins. They may be released by proteolysis during gastrointestinal transit or during food processing.

      General processes

      Physiological effects of bioactive peptides

      1. A) Effects on the cardiovascular system
      • Hypertension is one of the major risk factors for cardiovascular disease.
      • 30% of mortality is related to hypertension or its renal, cardiac or cerebral complications
      • ACE is an enzyme that is located in many tissues and plays an important role in blood pressure regulation and in turn hypertension.
      • ACE catalyses the conversion from angiotensin I to angiotensin II, a hormone which results in vasoconstriction, and subsequently in an increase in blood pressure.
      • Significant blood pressure reduction with 95 ml sour milk per day for 4 weeks.

      B) Effects on the immune system

      • Immuno modulator property
      1. alleviate allergic reactions
      2. enhance mucosal immunity
      • modulate lymphocyte proliferation
      • Two peptides, YG and YGG, fragments of bovine k-casein and α-lactalbumin, exert a stimulating effect on the proliferation of human peripheral blood lymphocytes in vitro
      • Antimicrobial property
      • Peptides derived from αs1-, αs2- and κ-casein & whey proteins been considered as potential precursors of bactericidal fragments. Eg. lactoferricin – a fragment of the whey protein lactoferrin.
      • These peptides have been found to be active against a broad range of pathogenic organisms e.g. E-coli, Helicobacter, Listeria, Salmonella and Staphylococcus, yeasts and filamentous fungi.
      • Antimicrobial peptides αs2 -CN f183-207 and f164-179 inhibit growth of Gram-positive and -negative bacteria at MICs ranges from 8 to 95 μmol/l.

      C) Effects on nervous system

      • The peptides that exist in dairy products which play an active role in the nervous system; are known as opioid peptides.
      • ß-casomorphin is the first major opioid peptide derived from ß-
      • Once these peptides are absorbed into the blood, can travel to the brain and various other organs and elicit pharmacological properties similar to opium or morphine.
      • This may be the reason why human neonates generally become calm and sleepy after drinking milk.

      D) Effects on nutritional status and dental health

      • Peptides enhanced mineral uptake
      • The charged side chains, in particular the phosphate groups of amino acids can bind minerals e.g. Ca, Mg, Fe and Zn. Since
      • CPPs can bind and solubilise minerals, they may have value in the prevention of osteoporosis, dental caries, hypertension, anemia and recalcification of tooth enamel.
      • Glycomacropeptide (GMP) derived from κ-casein seems to contribute to the anticaries effect by inhibiting the adhesion and growth of plaque-forming bacteria on the oral mucosa.

      Antioxidant peptides

      • Peptides with antioxidative properties can be released from caseins by hydrolysis with digestive enzymes and by proteolytic LAB in fermented milk.
      • Peptides from αs-casein possess free radical-scavenging activities and inhibit enzymatic and non-enzymatic lipid peroxidation.
      • Hypocholesterolemic peptide (Ile-Ile-Ala-Glu-Lys) from the tryptic hydrolysate of ß-lactoglobulin suppresses the cholesterol absorption in rat
      • Histidine and proline are the most important amino acids in the lipoprotein peroxidation‐inhibitory activity of peptides.
      • Tyrosine and tryptophan show free radical‐scavenging capacity.
      • Ala-Arg-His-Pro-His-Pro-His-Leu-Ser-Phe-Met(k-cn), Val-Lys-Glu-Ala-Met-Ala-Pro-Lys(β-cn),

      Conclusion

      • Bioactive peptides can be incorporated in the form of ingredients in functional and novel foods, dietary supplements and even pharmaceuticals with the purpose of delivering specific health benefits.
      • Many scientific, technological and regulatory issues must, however, be resolved before these substances can be optimally used in products.
      • There is a need to develop novel technologies, such as chromatographic and membrane separation techniques, to enrich active peptide fractions from the hydrolysates of various food proteins
      • It is important to study the technological properties of active peptide fractions and to develop model foods that contain these peptides and retain their activity for a guaranteed period.
      • Bioactive peptides possess very important biological functionalities, including antimicrobial antihypertensive, antioxidative, anticytotoxic, immunomodulatory, opioid, and mineral-carrying activities.

      About the Other

      MGM College of Food Technology, Gandheli, Chatrapati Sambhajinagar.