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SCIENTIFIC AND TECHNICAL JUSTIFICATION OF CONCEPTUAL PROVISIONS OF PROTEOMICS OF DAIRY BUSINESS
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SCIENTIFIC AND TECHNICAL JUSTIFICATION OF CONCEPTUAL PROVISIONS OF PROTEOMICS OF DAIRY BUSINESS
Рубрики: REVIEW
УДК 66 Химическая технология. Химическая промышленность. Родственные отрасли
Khramtsov A G 1
Авторы:
1.  North Caucasus Federal University

Тип:
Статья
DOI:
https://doi.org/10.21179/2308-4057-2016-2-16-31
Страницы:
с 16 по 31
Статус:
Опубликован
Получено:
31.12.2016
Одобрено:
31.12.2016
Опубликовано:
31.12.2016
Классификаторы:
УДК 66 Химическая технология. Химическая промышленность. Родственные отрасли
Язык материала:
английский
Ключевые слова:
milk proteins, casein, whey proteins, amino acids, composition and properties of protein complex of milk, ways of receipt, ways of use
Аннотация и ключевые слова
Аннотация (русский):
The paradigm of formation of science about nitrogen-containing compounds (protein complex) of raw milk - Proteomics - is stated. The monitoring of fractional composition, possibility of extraction, modification and application of the whole protein complex, caseins and serum proteins, their fractions and derivatives allows to consider the traditional and innovative component of dairy business in an absolutely new light, based on nanoclusters and biotechnology. The road map of casein complex of raw milk is considered. The characteristic of the main fractions of casein, from the point of view of modern biotechnology of cheeses and cottage cheese is provided. The characteristic of serum proteins of raw milk in the native and denatured states and after the microparticulation directed into nanotubes is separately considered. The unimproved opportunities for the modernization of technologies of extraction of protein clusters with the receipt of products for import substitution with export orientation are emphasized. For the first time in the logistics of system analysis the problems of controlled proteolysis of albumins of milk - casein and serum proteins, with the receipt of products for clinical nutrition are considered.

Ключевые слова:
milk proteins, casein, whey proteins, amino acids, composition and properties of protein complex of milk, ways of receipt, ways of use
Текст
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INTRODUCTION. STATEMENT OF THE PROBLEM According to [2] the postulates of LACTOOMICS offered [1] and adapted in print as the science about MILK, and to the principles of logistics of dairy business [3], it is advisable to put briefly some reasons down, in general, in respect of innovations and information technologies, about one of the main components of milk - PROTEINS (from Greek protey - the first) - a protein complex (nitrogen-containing compounds) as the anthem of life creation on our planet (according to F. Engels). In phenomenology logistics together with Glycoomics [4] and Lipidomics [5] the term Proteomics is used. Proteins of milk (nitrogen-containing compounds) are present practically in all dairy products - there is no deproteinized milk (fat-free as opposed to fat and delactosylated as opposed to lactose) in nature (in practice) yet. “Pure” protein complex - milk protein concentrates, casein, its modification - caseinates and derivatives (hydrolyzates) - peptides, an amino acid pool; serum proteins as a whole and individually, and also their multiple derivatives are known in the industry in the form of industrial products. Traditional casein, except its designated purpose as food in dairy products, is known from joiner's glue to artificial caviar. I felt warm for long in a jumper of "casein wool" acquired as a memorable souvenir in Poland (1965) - the casein was Polish and the "wool" was Japanese. And all this set of irreplaceable food products, medical supplies and technical semi-finished products - proteins begins with the multicomponent composition of protein complex of raw milk, the aggregate data about the types and fractions of which, according to the information given by A. Tepel [6], with the modern interpretation by V. V. El'chaninov [7, 8, 9, 10], are stated below. At the same time, the postulates, known from biological chemistry and genetics, general in importance, composition, structure and the properties of native natural proteins do not naturally repeat, the emphasis is only on milk proteins which are part of the biosphere, in the light of Lactoomics. OBJECTS AND METHODS OF STUDY The total content (the initial stage of road map of raw milk) of albumins in milk is within the limits of 2.9 … 4.0% (the total content of nonprotein nitrogen is up to 0.035%), which should be considered within Proteomics in practice, especially when monitoring primary raw milk as a commercial product and when treating serum. This indicator determines, along with milk fat, the technology and economy of dairy business. Is completely determined by the level of development of livestock production. The dairy industry correlates de facto with this indicator. Unfortunately, this indicator hasn't been formalized in this branch at the financial, technological and social levels in our country until recently. Table 1 provides the summary data on the content of albumins in a dry residual of raw milk [11]. It follows from the provided data that the protein component in raw milk is significant (at the level of 25.0%) in a solid and considerably differs by types, which naturally determines the status of received products - a complex, casein (without fractionation), whey proteins (generally, without fractionation). Table 2 provides the general characteristic about the content of protein fractions of raw (whole) milk. The structures, main and minor components, genetic variations, polypeptide chain and amino acid pool of protein compounds of milk raw materials are diverse. They constantly replenish. The information of Professor K. K. Gorbatova [12] with additions by A. Tepel [6] is given below about the amino acid pool of main fractions of proteins of raw milk. An exceeding information file of researches on the interpretation of primary structure of all fractions and genetic variations with the elements of poetic allegory and a practical component is at the back of each digit. For example, the polypeptide chain can give a sweet (aspartame) and a bitter taste in cheeses; it is possible to receive antibiotics (nisin) and, unfortunately, poison. And this all can be received from the protein structure of milk. The elementary composition of milk proteins (for your reference) according to various researchers [6, 12] is accurately traced in a hierarchy, with the contents at the level of, %: carbon (C) - 53.0, oxygen (O) - 23.0, nitrogen (N) - 15.6, hydrogen (H) - 7.0, sulfur (S) - 1.5, phosphorus (P) - 0.8. Table 1. Content of albumins in raw milk Description of raw material Content of % Solids, g/100ml Protein, g/100ml Raw milk 13.30 3.20 24.06 Cream, fat percentage 35% 41.30 2.40 5.81 Non-fat milk, fat percentage 0.05% 8.70 3.20 36.80 Butter milk 9.10 3.20 35.16 Whey 6.30 0.80 12.70 Table 2. Main groups of albumins of raw milk Name of fractions Content, g/100ml % Total content 3.27 100.0 Total of caseins 2.60 79.5 including: aS1-casein 1.00 30.6 aS2-casein 0.26 8.0 β-casein 10.01 30.8 k-casein 3.30 10.1 Total of serum proteins 0.63 19.3 including: a-lactalbumin 0.12 3.7 β-lactoglobulin 0.32 9.9 blood serum albumin 0.04 1.2 immunoglobulins 0.07 2.1 proteosopeptones 0.08 2.4 proteins of fat globules membranes 0.04 1.2 minor proteins trace amounts - Table 3. Amino acid pool of main fractions of proteins of raw milk Fractions of polypeptide chain Amino acid pool, pcs.×a-AA aS1-casein 199 aS2-casein 207 β-casein 209 k-casein 169 a-lactalbumin 123 β-lactoglobulin 162 blood serum albumin 582 The content of the main component of nitrogen-containing compounds of milk - casein is up to 80% of protein complex and is 2.6 … 3.2% (the more the better). Casein has a lot of fractions (it is considered that there are more than 20 now), genetic variations, fragments and groups (from 1 to 6 for each fraction), which is little considered in practice yet, for example, in cheese making and in the production of cottage cheese. A purposeful search with the use of all achievements of modern analytics of organic compounds is necessary. The content of whey proteins - at the level of 20% (not to confuse with whey proteins) in milk is 0.4 … 0.7%. Whey proteins of milk, just as casein, are fractional - up to 19 names with 7 genetic variations. The same goes to proteins o covers of fat globules (8 fractions). It is necessary within Lacoomics to point to a special, quite a new group of nitrogen-containing compounds, - minor (a low content, an important role) proteins - up to 2% of total mass (9 fractions and the mass of genetic variations). In general, macrocomponents (casein and whey proteins) and the minor component allow to draw a conclusion about a genetically full-weight set of clusters of nitrogen-containing compounds in milk, an irreplaceable food component during all the life cycle of mammals. From exactly this perspective, in relation to industrial processing of raw milk, this group of compounds shall be considered for all the assortment groups of products and production cycles. It is not really available in the existing study books yet! The methodology of study of albumins of raw milk and the received products is quite enough fulfilled [13, 14]. Is based on the gnoseology of coefficient 6.38 for nitrogen and the indispensability of Kyeldal's formula. At the same time, "the floating indicator of self-deception" for nonprotein nitrogen (NPN) in milk reaches 8%, and 30% in whey. Devices like MilkoScan neutralize this problem, but they are not absolutely recognized. There are breakthroughs, especially in the field of chromatography - gas, liquid and ionic chromatography [15]. For dairy business, of special interest are HPLC with reversed phases (RP-HPLC), fast protein liquid chromatography (FPLC) and size-exclusion chromatography (SEC), and also gel filtration, applicable for casein and serum proteins. RESULTS AND DISCUSSION Modern classification and nomenclature of milk proteins (is constantly replenishing and changing) [6, 12]. It is necessary to emphasize within Lactoomics and Proteomics the earlier mentioned polycomponent character of all four groups of milk proteins (caseins, whey proteins, protein of fat globules membranes, minor proteins) and to reconcile it once again with the road map of practical use of components, as a whole. Separately - with a possibility of modification for the primary components - peptides and amino acids, and also microparticulation. In such a logic of knowledge and analysis the modern postulates of Proteomics are considered. They are in the dynamics of development according to the achievements of fundamental sciences and practice of researches of creative teams of the international "dairy community". Let us consider the road map, in respect of technological monitoring, of each of the macrocomponents of milk protein complex. At the same time, we use the information file obtained from survey information [16] and an education guide [17], special researches [18, 19, 20], system publications [7-10] and the generalizing material [6]. Caseins, from the perspective of Lactoomics and Proteomics, are of special cognitive and practical interest. They draw attention of theorists and practitioners as ideal natural protein (especially in respect of polymorphism) and a source of profit for business since the time of a curious Dutch practician Mulder and the great scientist Gammersten. The depth of cognition of milk casein and the constant attention to this problematics is confirmed by the information stated above, and also by my personal observations at special IDF World Dairy Summits in the Netherlands (1973) and Austria (1975). This subject is discussed in all the events of IDF and at all Industry Summits. And the need of interest for the object of cognition can be confirmed with a simple question, which is not cleared up yet, - "why is milk white?". The trick is in casein. The confirmation follows from an observation, trivial and available to everyone, of color of milk after spontaneous or controlled (heating - cooling - souring) souring - serum is transparent with a yellowish-greenish shade, and the product is of white color. Fig. 1 provides the modern model of casein in the system HyperChem using the example of k-casein. It is considered that it reminds figuratively a jumping horse - there is nothing to do for chemists, physicists and biotechnologists but to "bridle a racer". Even more indicative is the computer model of casein clusters on the basis of fractal views. Fig. 2 provides the systematized information by Smykov [19] about the models of aggregation of clusters of casein micelles: diffusion limited aggregation (DLA) [21]; ballistic aggregation (BLA) [22-24]; rotation limited aggregation (RLA) [25-28]; diffusion limited cluster aggregation (DLCA) [29]; reaction limited cluster aggregation (RLCA) [30-32]; ballistic limited cluster-cluster aggregation (BLCA) [33-36]. The provided compositions of clusters characterize the alternative variants of process of structurization ("clotting") of casein micelles (daily performed by cheesemakers). They also cover the allegory of "the synthesis of the Universe" in case of the thermal denaturation of serum proteins (try it by holding your own observation). These provisions are interpreted by the outstanding author of the term and theory of fractals, American academician Benoit Mandelbrot [37], they underline the uniqueness of casein as an object of biocenosis in the Universe. And confirm once again the whey phenomenon. The genetic polymorphism of casein is well seen in the chromatogram (Fig. 3). Fig. 1. Variant of visualization of model of micelle of k-casein according to HyperChem - public information. (a) (b) (c) (d) Fig. 2. Beginning. Computer models of formation of casein clusters: (a) DLA, (b) BLA, (c) RLA,(d) DLCA, (e) RLCA, and (f) BLCA. (e) (f) Fig. 2. Ending. Computer models of formation of casein clusters: (a) DLA, (b) BLA, (c) RLA, (d) DLCA, (e) RLCA, and (f) BLCA. Подпись: Absorption 0 5 10 Volume of elution, ml 0.10 0.05 0 45 40 35 30 Fig. 3. Сhromatogram of milk caseins. The primary structure of casein, - a consecutive compound of amino acids, depends on a type of fraction and its genetic variation. This fine picture, worthy of "poetization", should be the cornerstone of Proteomics and demands individual consideration. The controlled hydrolysis (proteolysis) of protein compounds of casein and serum proteins turns into an independent branch which should be mastered in the dairy industry (the meat industry has earlier given complete control over it) - there were special shops by meat-processing plants. Are considered separately in respect of the controlled phenylalanine elimination. The secondary structure - peptides, is already in technology [38] and will be considered below. The tertiary structure of proteins is of great importance for the technological properties of milk. The three-dimensional tertiary structures can only be established in the previously crystallized proteins during the research by means of the X-ray diffraction analysis. It was not possible to perform casein crystallization till now - it is considered that it is hardly possible [6]. Is it the second problem for cognition, for example, in Skolkovo or within IDF?! The associates of micelles form a number of variants of quarternary structure of casein which is unstable and constantly demands the controlled regulation by processing methods (temperature, active acidity, mechanical effect). 95% of casein in its native state is in the type of casein micelles or associations of subunits (casein submicelles) which are the complexes of casein monomeric molecules. The behavior of milk during technological processing and industrial conversion is determined generally by the properties of casein micelles. The size of casein micelles is from 30 (nanolevel) to 300 (colloidal state), they have a spherical shape. It is they what determines the so-called yield of protein and fat products - cottage cheese and casein. Whey proteins are polymorphic, which is well seen in the chromatogram (Fig. 4), their structure is unique (Table 4) and their rheomorphism is incomparable (Fig. 5). Their nanosize - at the level of 10 nanometers - is accurate, which forms, along with lactose, an authentically soluble system of milk (serum and ultrafiltrates). The protein complex of whey is specific, for example, due to the availability of k-casein, and demands individual consideration with the elements of repetition with Table 2. Lactoglobulin (β-LG) - the main whey protein is non-uniform in structure. It is presented by several genetic variations A, B, C, D, E, F and G differing in amino acid structure. Their content is 50-60% of the total of serum proteins. Lactoalbumin (α-La) is the second protein in order of importance, it is presented by the genetic variations A and B. Immunoglobulins (IgG) is a non-uniform group of proteins - glycoproteids of monomers and polymers IgG1, IgG2, IgA and IgM. Serum albumin (Sa) is presented by a polypeptide chain folded in four bound disulfide threads of globular segments of non-uniform proteins. Lactoferrin, as well as transferrin, is an iron blood protein. Osteopontin (OPN) - a multifunctional protein, is found recently, plays an important role in preserving the immune status of the newborns. All fractions of whey proteins have small sizes and high hydrophily, which explains their high stability in solution. Unlike casein, whey proteins do not form micelles, do not coagulate under the effect of enzymes and do not precipitate in case of milk souring. A thermal effect (denaturation) is required for the realization of this process. Whey proteins have a rather low molecular weight - from 14 000 to 69 000. α-La β-La B β-La A Подпись: D with 214 nm 0.1 0.8 0.6 0.4 0.2 0 0 5 10 15 Fig. 4. Сhromatogram of whey proteins. Table 4. Characteristic of milk serum proteins Name of fractions Content, % Molal weight β-lactoglobulin 0.32 18 400 α-lactalbumin 0.12 14 000 Proteose-peptone fraction 0.12 from 10 000 to 200 000 Immunoglobulins 0.09 160 000 Serum albumin 0.03 70 000 Lactoferrin 0.02 93 000 L-Carnitine 0.03 - Osteopontin trace amounts - Total 0.73 - β-lactoglobulina-lactalbumin Fig. 5. Molecular structures of some whey proteins. Minor proteins of milk and protein compounds of fat globules membranes draw an increasing attention which obviously follows from the materials of seven International Whey Conferences, especially that of the fifth and sixth ones [39, 40]. For example, angiogenin was the subject of special researches [41] but the result is an original drug Milkang [42], which waits for its practical realization. The derivation (receipt) of proteins from raw milk is an indispensable component of Proteomics and is constantly studied in respect of scientific knowledge for practical application [43]. The receipt of a complex of milk proteins in the form of milk protein concentrates is developed and relates to the basic researches [6, 16]. Unfortunately, this attractive technology is not widely scaled because of an inevitable loss of native properties. The problem waits for a decision. Our researches on soft cheeses [44, 45], cottage cheese products [46], the formation of the brand “LipKA” (a lipid casein albumin concentrate) [47] and the method “TermoLakt” [48], with an attempt to implement the paradigm of complex release of proteins from raw milk, confirm this provision. A search of the optimal solution within Proteomics is forthcoming. Probably, in this case the breakthrough innovations of Professor Z.S. Zobkova and her colleagues concerning enzymatic cross-linking of milk proteins are quite perspective, for example, using transglutaminase for the receipt of new structures and original functions of the derived special purpose products [49]. Coagulation of raw milk caseins - acid, rennet, acid and rennet, chlorcalcic coagulation, coagulation with the use of electrophysical methods, polysaccharides and membrane technology is quite well studied by and is applied at the level of traditions and innovations [11]. The traditions are acid and rennet coagulations. The innovations are membrane technologies (ultrafiltration). The supertechnologies are the biomembrane technology with the use of polysaccharides (membraneless reverse osmosis) and microparticulation (nanotubes). The whey protein complex, in respect of release, is quite well studied and realized in practice [10]. The traditions are thermal denaturation; the innovations are membrane technologies. The supertechnologies are microparticulation (nanotubes). The fundamentals of development of technology of dairy products, using the example of casein coagulation - (Laktoomics base), were worked out by A.M. Osintsev [18] in a system type. The complex extraction of milk proteins using the example of soft cheeses is thoroughly studied by I.А. Smirnova [50] and O.A. Suyunchev [48]. They are widely realized (scaled) in practice. However, the search is not finished. As an example, we will briefly consider "the membraneless technology", one of the alternative variants of extraction of casein with the implementation in the line (series) of products of the brand “Bio-TON” [51]. Historically, the cycle of development of new technology of thermodynamic fractionation of milk components (sometimes called as membraneless reverse osmosis) includes the stage of accidental observations in the thirties of spontaneous milk separation when adding polysaccharide (Patent DE 555273) for two fractions - casein concentrate and a serum fraction. It was the stage which proved a unique possibility of separation of milk into a casein and non-casein fractions. The method did not find practical application and was forgotten for more than 50 years. Then, at A.N. Nesmeyanov Institute of Organoelement Compounds (INEOS) of Academy of Sciences of the USSR, within a number of large-scale basic researches of Professor V.B. Tolstoguzov and his colleagues studying the problem of search of food reserves, a possibility of controlled separation of protein solutions by polysaccharides was theoretically proved and experimentally demonstrated. Milk was one of the ideal objects. In the State Enterprise «Scientific Research Institute of the Complex Utilization of Dairy Raw Material (formerly known as All-Union Research Institute of the Complex Utilization of Dairy Raw Material and The North Caucasian branch of All-Union Research Institute of the Cheesemaking and Buttermaking Industry) by efforts of the school of my dear colleague from RAS Professor V.V. Molochnikov complex specific researches [52, 53] on the development of non-waste technology of fractionation of milk by polysaccharides using the principle of membraneless return osmosis [54], which was named as “Bio-Ton”, were performed. Fig. 6 provides the schematic diagram of separation of milk by biopolymers (polysaccharides). As a result of separation, natural casein concentrate is received, the characteristic of which (in respect of Proteomics) we will consider in more detail [55]. Natural casein concentrate (NCC) is a light-cream thixotropic uniform liquid with creamy consistence and with a pure milk taste and smell. It completely dissolves in water. The thermal effect up to 100°C on liquid natural casein concentrate does not change its solubility, which testifies to naturalness as it is known that milk casein has a unique structure in its native state, which specifies its resistance to the effect of denaturant agents and its high splittability by proteolytic enzymes. Table 5 provides the average physical and chemical values of NCC in comparison with skim milk.
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