Effect of heat treament on food protein

Thông qua việc cung cấp các axit amin, protein cần thiết cho sự phát triển của con người, nhưng chúng cũng có một loạt các đặc tính cấu trúc và chức năng có ảnh hưởng sâu sắc đến chất lượng thực phẩm. Protein đóng một vai trò cơ bản không chỉ trong việc duy trì sự sống mà còn trong các loại thực phẩm có nguồn gốc từ thực vật và động vật. Thực phẩm khác nhau về hàm lượng protein của chúng và thậm chí nhiều hơn về đặc tính của những protein đó. Ngoài việc chúng đóng góp vào các đặc tính dinh dưỡng của thực phẩm thông qua việc cung cấp các axit amin cần thiết cho sự tăng trưởng và duy trì của con người, protein còn truyền cơ sở cấu trúc cho các đặc tính chức năng khác nhau của thực phẩm. Định nghĩa về protein Từ “protein” được định nghĩa là bất kỳ nhóm hợp chất hữu cơ phức tạp nào, bao gồm cơ bản là sự kết hợp của các axit amin trong các liên kết peptit, chứa cacbon, hydro, oxy, nitơ và thông thường, lưu huỳnh. Phân bố rộng rãi trong thực vật và động vật, protein là thành phần chính của nguyên sinh chất của tất cả các tế bào và rất cần thiết cho sự sống. (“Protein” có nguồn gốc từ một từ tiếng Hy Lạp có nghĩa là “đầu tiên” hoặc “chính” vì vai trò cơ bản của protein trong việc duy trì sự sống.) Protein trong thực phẩm Axit amin, peptit và protein là những thành phần quan trọng của thực phẩm. Chúng cung cấp các chất xây dựng cần thiết cho quá trình sinh tổng hợp protein. Ngoài ra, chúng trực tiếp góp phần tạo nên hương vị của thực phẩm và là tiền chất cho các hợp chất tạo mùi thơm và màu sắc được hình thành trong quá trình phản ứng nhiệt hoặc enzym trong sản xuất , chế biến và bảo quản thực phẩm. Các thành phần khác của thực phẩm, ví dụ như carbohydrate, tham gia vào các phản ứng như vậy. Protein cũng đóng góp đáng kể vào các đặc tính vật lý của thực phẩm thông qua khả năng ổn định gel, bọt, bột nhào, nhũ tương và cấu trúc sợi. Các nguồn protein quan trọng nhất và sự đóng góp của chúng trong việc sản xuất protein trên toàn thế giới. Ngũ cốc đóng góp hơn một nửa vào sản xuất protein, tiếp theo là hạt có dầu và thịt. Bên cạnh động thực vật, tảo, nấm men và vi khuẩn có thể được sử dụng để sản xuất protein ( protein đơn bào (SCP)). Hàm lượng protein thay đổi như sau:> 20% (pho mát, thịt, các loại đậu, hạt có dầu); 10–20% (cá, trứng); 5–10% (ngũ cốc); và < 5% (sữa, rễ, củ, rau, quả, nấm).

Effect of heat treatment on food protein

Nguyen Le Huy 20190624 Through their provision of amino acids, proteins are essential to human growth, but they also have a range of structural and functional properties which have a profound impact on food quality Proteins play a fundamental role not only in sustaining life, but also in foods derived from plants and animals Foods vary in their protein content and even more so in the properties of those proteins In addition to their contribution to the nutritional properties of foods through provision of amino acids that are essential to human growth and maintenance, proteins impart the structural basis for various functional properties of foods

Definition of protein

The word “protein” is defined as any of a group of complex organic compounds, consisting essentially of combinations of amino acids in peptide linkages, that contain carbon, hydrogen, oxygen, nitrogen, and usually, sulfur Widely distributed in plants and animals, proteins are the principal constituent of the protoplasm of all cells and are essential to life (“Protein” is derived from a Greek word meaning “first” or “primary” because of the fundamental role of proteins in sustaining life.)

Protein in food

Amino acids, peptides, and proteins are important constituents of food They supply the required building blocks for protein biosynthesis In addition, they directly contribute to the flavor of food and are precursors for aroma compounds and colors formed during thermal

or enzymatic reactions in production, processing, and storage of food Other food constituents, e.g., carbohydrates, take part in such reactions Proteins also contribute significantly to the physical properties of food through their ability to stabilize gels, foams, doughs, emulsions, and fibrillar structures

The most important protein sources and their contribution to world-wide production of protein Cereals contribute to protein production by more than half, followed by oil seeds and meat Besides plants and animals, algae, yeasts and bacteria may be used for protein production (single-cell protein (SCP)) The protein content varies as follows: > 20% (cheeses, meat, legumes, oil seeds); 10–20% (fish, eggs); 5–10% (cereals); and < 5% (milk, roots, tubers, vegetables, fruits, mushrooms)

Cereals and cereal products

Cereals and cereal products are amongst the most important staple foods of mankind Proteins provided by bread consumption in industrial countries meet about one-third of the daily requirement The major cereals are wheat, maize, rice, barley, sorghum, oats, millet, and rye Wheat and rye have a special role since only they are suitable for

bread-making With the example of wheat, the cereal proteins have been separated by the basis

of their solubility, into four fractions: the water-soluble albumins, the salt-soluble globulins, the 70% aqueous ethanol-soluble prolamins, and the remaining glutelins The levels of fractions differ amongst cereals with albumins amounting to 4–44%, globulins 3–12%, prolamins 2–48%, and glutelins 24– 77% of the whole protein fraction Each of fractions consists of a larger number of proteins Albumins and globulins contain the enzymes, whereas prolamins and glutelins are storage proteins

Meat and meat products

Meat and meat products are other important staple foods, in particular in industrial countries The main meat-producing warm-blooded animals are pig, cattle, poultry, sheep, goats, and buffalo Meat proteins, i.e., the proteins of the muscle, are divided into three groups: proteins of the contractile apparatus (myofibrillar proteins), soluble proteins (sarcoplasma proteins), and insoluble proteins (connective tissue and membrane proteins) The myofibrillar proteins of a typical mammalian muscle amount to about 60% of total muscle protein, with myosin (29%) and actin (13%) as their predominating components and about 20 minor components including connectin, tropomyosins, troponins, and actinins The sarcoplasma proteins form about 30% of total protein They contain most of the enzymes, in particular those of glycolysis and the pentosephosphate cycle, but also considerable amounts of creatine kinase (2.7% of total protein), myoglobin, and some hemoglobin The insoluble proteins contain collagen as the main component, besides

elastin, insoluble enzymes, and cytochrome c In connective tissue, collagen forms a

triple-stranded helix composed of α-helices Covalent cross-links are formed between the fibers

of collagen during maturation and aging, thus improving its mechanical strength When heated, collagen fibers shrink or are converted into gelatine, depending on the temperature The structure of the gelatin obtained after cooling depends on the gelatine concentration and temperature gradient Collagen contains two unusual amino acids, 4-hydroxyproline and 5-hydroxylysine Since the occurrence of the former is confined to connective tissue, its determination provides data on the extent of connective tissue content of a meat product

Milk and dairy products

Milk and dairy products form a further important group of staple foods Milk generally means cow’s milk, but milk from buffalo, goats, and sheep is of importance in some regions Milk proteins, in particular the caseins, play an important role in processing to yield dairy products such as cheese and sour milk products The caseins make up about 80% of total milk proteins They have been separated later into different fractions: -, -, -,-, and -caseins, constituting 34, 8, 9, 25, and 4% of total protein, respectively In cheese-making, the specific cleavage of -casein by chymosin into para casein and a glycopeptide reduces the solubility of the casein complexes and casein micelles, thus leading to their aggregation followed by gel formation (curd formation) The whey proteins (about 20% of total protein) consist of -lactoglobulins, -lactalbumins (both in different genetic variants),

serum albumin, immunoglobulins, and proteose-peptone Also, more than 40 enzymes occur in the whey protein fraction, but in much lower quantities than the other components Whey proteins can be incorporated into the curd using several new processing methods of cheese-making in order to increase the yield and also to reduce waste water or whey treatment costs

Legumes

Legumes (pulses) are very important staple foods in some parts of the world, e.g., soya beans in South-east Asia and common beans in Latin America Other legumes, some of greater regional importance, include peas, peanuts, chick peas, broad beans, and lentils Legume proteins, when fractionated in a similar way to cereal proteins, yield three fractions: albumins, globulins, and glutelins The portion of the fractions varies, depending

on the legume species, but globulins always predominate The globulins are subdivided, initially according to sedimentation during ultracentrifugation, into 11S and 7S globulins (legumins and vicilins, respectively) Again, the subfractions derived from different legumes are sometimes designated by special names, e.g., glycinin and arachin for soya bean and peanut

legumin, as well as -conglycinin and phaseolin for soya bean and common bean vicilin, respectively Soya protein isolates, produced by diluted alkali extraction of defatted soya bean flakes followed by acid precipitation, are texturized and flavored for use as meat substitutes or are added to foods to raise their protein level and improve their processing qualities such as the water-binding capacity or emulsion stability The isolates contain about 95% protein and consist of 11S and 7S globulins The similarity between the caseins from bovine milk and the globulins from soya beans is reflected by the production of some typical Asian foods such as soy milk, soy curd (tofu), and soy cheese (sufu)

Eggs

Eggs are used as a food not only because of their 0excellent nutritional quality but also because of their functional properties Eggs generally means chicken eggs; those of other birds (geese, ducks, plovers, seagulls, quail) are less important Egg proteins are divided into those of egg white and those of egg yolk Egg white proteins (about 10% of total egg white) are ovalbumin, conalbumin (ovotransferrin), ovomucoid, ovomucin, lysozyme, ovoglobulin G2, ovoglobulin G3, and some minor components (54, 12, 11, 3.5, 3.4, 4, 4, and 2.5% of total egg white protein, respectively) Ovalbumin, conalbumin, ovomucin, and the ovoglobulins G contribute to foam formation and foam stability Yolk proteins (about 17% of total yolk) are phosvitin, the livetins, and the protein moieties of high-density lipoproteins (HDL) and low-density (LDL) lipoproteins (13, 31, 36, and 20% of total yolk protein, respectively) Apart from phospholipids, LDL and proteins are responsible for the

egg yolk alone Owing to the ability of all egg proteins, except ovomucoid and phosvitin, to coagulate when heated, egg products are important food binding agents

The nutritional quality of a food protein depends on the absolute content of essential amino acids, the relative proportions of essential amino acids, and their ratios to nonessential

amino acids In addition, the digestibility of the food protein, the influence by other food components such as dietary fibers, polyphenols, or proteinase inhibitors, and also the total food energy intake are of importance During pregnancy and lactation, the first 6 months, and after 6 months, the daily requirement increases by 13, 24, and 18%, respectively The biological value of a protein is generally limited by the following amino acids:

 Lysine: deficient in proteins of cereals and other plants;

 Methionine: deficient in proteins of bovine milk and meat;

 Threonine: deficient in wheat and rye;

 Tryptophan: deficient in casein, corn and rice

Protein properties

Conformation

Primary structure

The primary structure gives the sequence of amino acids in a protein chain with peptide linkage The peptide bonds have partial (40%) double-bond character with p-electrons shared between the C–O and C–N bonds Normally the bond has a trans configuration, i.e., the oxygen of the carbonyl group and the hydrogen of the NH group are in the trans position; a cis configuration only occurs in exceptional cases

Secondary structure

The secondary structure reveals the arrangement of the chain in space The peptide chains are not in an extended or unfolded form

-sheet The peptide chain is always lightly folded on the C atom, thus the R side chains extend perpendicularly to the extension axis of the chain, i.e., the side chains change their projections alternately from to Such a pleated structure is stabilized when more chains interact along the axis by hydrogen bonding, thus providing the crosslinking required for stability When adjacent chains run in the same direction, the peptide chains are parallel This provides a stabilized, planar, parallel sheet structure When the chains run in opposite directions, a planar, antiparallel sheet structure is stabilized

Helical structures The peptide chains are coiled like a threaded screw These structures are

stabilized by intrachain hydrogen bridges which extend almost parallel to the helix axis, cross-linking the CO and NH groups

Reverse turns An important conformational feature of globular proteins is the reverse

turns, -turns, or -bends They occur at “hairpin” corners, where the peptide chain changes direction abruptly Such corners involve four amino acids residues, among them frequently proline Glycine is favored in the third position of -bends on the basis of energy

considerations Different types of -turns are known, for which different amino acids are allowed

Super secondary structures Analysis of known structures has demonstrated that regular

elements can exist in combined forms Examples are the coiled-coil -helix, chain segments with antiparallel -structures (-meander structure) and combinations of -helix and -structure

Tertiary and Quaternary Structure

Proteins can be divided into two large groups on the basis of conformation: (1) fibrillar (fibrous) or scleroproteins, and (2) folded or globular proteins

Fibrous proteins The entire peptide chain is packed or arranged within a single regular

structure for a variety of fibrous proteins Examples are wool keratin helix), silk fibroin (-sheet), and collagen (a triple helix) Stabilization of these structures is achieved by intermolecular binding (electrostatic interaction and disulfide linkages, but primarily hydrogen bonds and hydrophobic interactions)

Globular proteins Regular structural elements are mixed with randomly extended chain

segments (random-coiled structures) in globular proteins The proportion of regular structural elements is highly variable: 20–30% in casein, 45% in lysozyme, and 75% in myoglobin Five structural subgroups are known in this group of proteins: (1) -helices occur only; (2) -structures occur only; (3) -helical and -structural portions occur in separate segments on the peptide chain; (4) -helices and -structures alternate along the peptide chain; and (5) -helices and -structures do not exist The process of peptide chain folding occurs spontaneously, probably arising from one center or from several centers of high stability in larger proteins Folding of the peptide chain packs it densely, by formation of a large number of intramolecular noncovalent bonds

Quaternary structures In addition to the free energy gain by folding of a single peptide

chain, association of more than one peptide chain (subunit) can provide further gains in free energy In principle, such associations correspond to the folding of a larger peptide chain around several structural domains without covalently binding the subunits

Denaturation

It is a reversible or irreversible change of native conformation (tertiary structure) without cleavage of covalent bonds (except for disulfide bridges) Denaturation is possible with any treatment that cleaves hydrogen bridges, or ionic or hydrophobic bonds This can be accomplished by changing the temperature, adjusting the pH, increasing the interfacial area, or adding organic solvents, salts, urea, guanidine hydrochloride, or detergents such as sodium dodecyl sulfate Denaturation is generally reversible when the peptide chain is stabilized in its unfolded state by the denaturing agent and the native conformation can be reestablished after removal of the agent Irreversible denaturation occurs when the unfolded peptide chain is stabilized by interaction with other chains (as occurs for instance

with egg proteins during boiling) During unfolding, reactive groups, such as thiol groups, that were buried or blocked may be exposed Their participation in the formation of disulfide bonds may also cause an irreversible denaturation Denaturation of biologically active proteins is usually associated with loss of activity The fact that denatured proteins are more readily digested by proteolytic enzymes is also of interest

Physical properties

Dissociation

Proteins, like amino acids, are amphoteric Depending on pH, they can exist as polyvalent cations, anions, or zwitterions Since -carboxyl and -amino groups are linked together by peptide bonds, the uptake or release of protons is limited to free terminal groups, and mostly to side chains In contrast to free amino acids, the pKa values fluctuate greatly for proteins since the dissociation is influenced by neighboring groups in the macromolecule

In the presence of salts, e.g., when binding of anions is stronger than that of cations, the isoelectric point is lower than the isoionic point In most cases the shift in pH is consistently positive, i.e., the protein binds more anions than cations

Solubility, Hydration, and Swelling Power

Protein solubility is variable and is influenced by the number of polar and apolar groups and their arrangement along the molecule Generally, proteins are soluble only in strongly polar solvents such as water, glycerol, formamide, dimethylformamide, or formic acid In a less polar solvent such as ethanol, few proteins have appreciable solubility The solubility in water is dependent on pH and on salt concentration Protein solubility is decreased (“salting-out” effect) at higher salt concentrations due to the ion hydration tendency of the salts

Since proteins are polar substances, they are hydrated in water The degree of hydration (grams of water of hydration per gram protein) is variable It is 0.22 for ovalbumin (in ammonium sulfate), 0.06 for edestin (in ammonium sulfate), 0.8 for -lactoglobulin, and 0.3 for hemoglobin

The swelling of insoluble proteins corresponds to the hydration of soluble proteins in that insertion of water between the peptide chains results in an increase in volume and other changes in the physical properties of the protein The amount of water taken up during swelling can exceed the dry weight of the protein by several times

Chemical Reactions

In contrast to free amino acids, except for the relatively small number of functional groups

of the terminal amino acids, only the functional groups in protein side chains are available for chemical reactions

Lysine Residue

Reactions involving the lysine residue can be divided into several groups: (1) reactions leading to a positively charged derivative; (2) reactions eliminating the positive charge; (3) derivatizations introducing a negative charge; and (4) reversible reactions The last are of

particular importance.

Arginine Residue

The arginine residue of proteins reacts with - or -dicarbonyl compounds Reaction of the arginine residue with 1,2-cyclohexanedione is highly selective and proceeds under mild conditions Regeneration of the arginine residue is again possible with hydroxylamine

Glutamic and Aspartic Acid Residues

These amino acid residues are usually esterified with methanolic hydrochloric acid There can be side reactions, such as methanolysis of amide derivatives or N,O-acyl migration in serine or threonine residues Diazoacetamide reacts with a carboxyl group and also with the cysteine residue to carboxamidomethyl derivatives

Amino acid esters or other similar nucleophilic compounds can be attached to a carboxyl group of a protein with the help of a carbodiimide Amidation is also possible by activating the carboxyl group with an isoxazolium salt to an enolester and its conversion with an amine

Cystine Residue

Reductive cleavage of cystine occurs with sodium borohydride and with thiols odification Cleavage of cystine is also possible by a nucleophilic attack

Electrophilic cleavage occurs with Ag+ and Hg+ or Hg2+ The sulfenium cation which is formed can catalyze a disulfide exchange reaction In neutral and alkaline solutions a disulfide exchange reaction is catalyzed by the thiolate anion

Cysteine Residue

A number of alkylating agents yield derivatives which are stable under the conditions for acid hydrolysis of protein The reaction with ethylenimine gives an S-(2-aminoethyl) derivative and, hence, an additional linkage position in the protein for hydrolysis by trypsin lodoacetic acid, depending on the pH, can react with cysteine, methionine, lysine, and histidine residues

Cysteine is readily converted to the corresponding disulfide, cystine, even under mild oxidative conditions, such as treatment with iodine or potassium hexacyanoferrate(III) Stronger oxidation of cysteine, and also of cystine, e.g., with performic acid, yields the corresponding sulfonic acid, cysteic acid

Methionine Residue

Methionine residues are oxidized to methionine sulfoxide with hydrogen peroxide The sulfoxide can be reduced, regenerating methionine, using an excess of thiol reagent With performic acid, methionine sulfone is formed

-Halogen carboxylic acids, -propiolactone, and alkyl halides convert methionine into sulfonium derivatives, from which methionine can be regenerated in an alkaline medium with an excess of thiol reagent Reaction with cyanogen bromide, which splits the peptide bond on the carboxyl side of the methionine molecule, is used for selective cleavage of proteins

Histidine Residue

Diethyl pyrocarbonate reacts with histidine to form N-(ethoxycarbonyl)histidine With iodoacetamide, N-1-(carboxamidomethyl)-, N-3-(carboxamidomethyl)-, or N-1,N-3-di(carboxamidomethyl) histidine are formed

Selective modification of histidine residues present on active sites of serine proteinases is possible Substrate analogs such as halogenated methyl ketones inactivate such enzymes by N-alkylation of the histidine residue

Tryptophan Residue

N-Bromosuccinimide oxidizes the tryptophan side chain and also tyrosine, histidine, and cysteine Other oxidative cleaving reagents are -iodosobenzoic acid and 3-bromo-3-methyl-(nitrophenylmercapto)-3H-indole Selective modification of histidine is possible with 2-hydroxy-5-nitrobenzyl bromide (Koshland reagent I) and 2-nitrophenylsulfenyl chloride

Tyrosine Residue

Selective acylation of tyrosine can occur with 1- acetylimidazole as a reagent Diazotized -arsanilic acid reacts with tyrosine ( substitution) and with histidine, lysine, tryptophan, and arginine Tetranitromethane introduces a nitro group into the position

Bifunctional Reagents

Bifunctional reagents enable intra-and intermolecular cross-linking of proteins Examples are bifunctional imidoester, maleimides, fluoronitrobenzene, and isocyanate derivatives

Interactions and Reactions Involved in Food Processing

Reaction with carbohydrates

Many foodstuffs contain reducing sugars and amino compounds such as proteins, peptides, amino acids, and amines Reactions between these components are usually classed under the term ‘nonenzymatic browning.‘ They occur especially at a higher temperature, low water activity and during longer storage Reactive sugars are glucose, fructose, maltose, lactose, and, to a smaller extent, reducing pentoses On the side of the amino components, primary amines are more important than secondary amines because their concentration in

foods is usually higher Exceptions are, for example, malt and corn products, which have a high proline content In the case of proteins, the e-amino groups of their lysine residues react predominantly, but guanidino groups of arginine residues can also react These reactions

result in:

 Brown pigments (known as ‘melanoidins‘): baking and roasting,

 Volatile compounds: contribute aroma of cooking, frying, roasting, baking besides the generation of off-flavors in food storage and processing

 Bitter substances: desired to coffee, but can cause off-flavor

 Reductones: highly reductive properties and contribute to the stabilization of foods against oxidative deterioration

 Losses of essential amino acids

 Mutagenic compounds

 Cross-linking of proteins

Reaction with lipid oxidation products

Products Formed from Hydroperoxides

Fatty acid hydroperoxides formed thermally or enzymatically in food are usually degraded further This degradation can also be of nonenzymatic nature In nonspecific reactions involving heavy metal ions, heme(in) compounds or proteins, hydroperoxides are transformed into oxo, epoxy, mono-, di-and trihydroxy carboxylic acids Unlike hydroperoxides, i.e., the primary products of autoxidation, some of these derivatives have a bitter taste Such compounds are detected in legumes and cereals They may play a role in other foods rich in unsaturated fatty acids and proteins, such as fish and fish products

Lipid–Protein Complexes

Studies related to the interaction of hydroperoxides with proteins have shown that, in the absence of oxygen, linoleic acid 13-hydroperoxide reacts with N-acetylcysteine, yielding an adduct that consists of several isomers However, in the presence of oxygen, covalently bound amino acid–fatty acid adduct formation is significantly suppressed; instead, oxidized fatty acids are formed

Protein Changes

Some properties of proteins change when they react with hydroperoxides or their degradation products This is reflected by changes in food texture, decreases in protein solubility (formation of cross-linked proteins), changes in color (browning), and changes in nutritive value (loss of essential amino acids)

Decomposition of Amino Acids

Studies of model systems have revealed that protein cleavage and degradation of side-chains, rather than the formation of protein networks, are the preferred reactions when

the water content of protein–lipid mixtures decreases The strong dependence of this loss

on the nature of the protein and on reaction conditions is obvious

Reaction under alkaline condition

Losses of available lysine, cystine, serine, threonine, arginine, and some other amino acids occur at high pH values Hydrolysates of alkali-treated proteins often contain some unusual compounds such as ornithine, -aminoalanine, lysinoalanine, ornithinoalanine, lanthionine, methyllanthionine and isoleucine, as well as other -amino acids

Reaction under oxidative conditions

Oxidative changes in proteins primarily involve methionine, which forms methionine sulfoxide relatively readily After in reduction to methionine, protein-bound methionine sulfoxide is apparently biologically available

Functional properties

Water absorption

Meat, sausages, bread, cakes

Elasticity

Hydrophobic bonding in gluten, disulfide

Whipped toppings, chiffon desserts, angel cakes

Heat treatment for food protein

Amino acid composition and sequence determine the native structure, functionality, and nutritional quality of a protein in a set environment During food processing, heat is often added to the protein’s environment, and this addition of energy can change any or all of the structural, functional, and nutritional characteristics of the native protein Foods are complex systems, and it is important to recognize that pH, water activity, food composition, and interactions of these with temperature also affect protein properties to varying extents