What Is Rice Protein?

Rice is one of the world's staple foods, with more than half of the world's population and more than two thirds of China's population eating rice as their staple food. Therefore, rice protein is an important source of protein in people's diets. China has a large area under rice cultivation, with an annual output of 180 billion kilograms of rice. In addition to supplying people's daily dietary needs, the processed rice is also used as a raw material for the fermentation of monosodium glutamate (MSG) and the production of starch sugar. In these processes, a large number of by-products such as rice bran and rice residue are produced. Rice bran is rich in nutrients, of which the protein content is about 12%, and the protein content in skimmed rice bran can be as high as 18%.

The protein content of rice dregs is above 40%, commonly called rice protein powder and rice protein concentrate (RPc). They are valuable protein resources, foreign countries attach great importance to the development and utilization of rice and rice bran, and produce nutritional health food and cosmetics with high added value. In the past, our country used them as animal feed, and the resources have not been reasonably utilized. In recent years, domestic attaches great importance to this, some scientific research institutions and enterprises have increased the research and development efforts. In this paper, from the perspective of development and utilization, the latest progress of research on rice and rice bran protein in recent years at home and abroad is introduced.

1 Structure, composition and properties of rice protein

Understanding the structure, composition and properties of rice proteins is fundamental to their utilization. There are many kinds of rice protein, and they are generally categorized by their solubility characteristics. Firstly, the protein fraction obtained by extracting the protein in rice or rice bran with water is called albumin; the protein fraction obtained by extracting the residue with dilute salt solution is called globular protein; the fraction extracted with 75% ethanol is called alcohol-soluble protein; and finally, the protein in the residue can only be dissolved by acid or alkali, which is called acid-soluble protein and alkali-soluble protein, and both of them are collectively called glutenin.

Gluten and alcohol-soluble proteins, also called storage proteins, are the main protein components in rice, with gluten accounting for more than 80% of the total protein and alcohol-soluble proteins accounting for about 10%; and clear and globular proteins, which are physiologically active proteins in rice, play an important physiological role in the early stage of rice grain germination.

The amino acid composition of different proteins is unique. Clear proteins have a high content of uncharged hydrophobic amino acids and a low content of acidic amino acids; globular proteins have a high content of basic amino acids, more than 15%, while alcohol-soluble proteins have only about half of the basic amino acid content of globular proteins, but their hydrophobic amino acids are much higher than those of other types of proteins [1].

The solubility of proteins is not only related to their amino acid composition, but also to their state of existence. Studies have shown that proteins in endosperm mainly exist in the form of two types of aggregates, i.e., PB-I and PB-" types. Electron microscopy showed that PB-I aggregates were lamellar in structure, with dense particles of 0.5~2 μm in diameter, while PB-" was ellipsoidal, undivided, with a diameter of 0.5~2 μm. PB-I aggregates were observed by electron microscopy, which showed that PB-I aggregates had a lamellar structure with dense particles of 0.5-2 μm in diameter, and the alcohol soluble proteins were found in PB-I; while PB-" was ellipsoidal, non-stratified, and homogeneous, with particles of about 4 μm in diameter, and the peripheral membranes of the particles were inconspicuous, and glutenin and globulin were found in PB-". The two aggregates are often found together [2 - 3].

During rice germination, the two protein aggregates disintegrated, but their digestibility differed significantly, with PB-" being more easily digested and hydrolyzed due to the absence of dense hard cores, whereas PB-I retained its lamellar structure at 9 days after germination. The SDS-PAGE technique demonstrated that new electrophoretic bands, i.e., new protein fractions, were constantly appearing in PB - ", whereas the fractions of PB - I were stable [4]. This suggests that there is a difference in the metabolism of the two protein molecules.

Rice proteins are high in cystine and contain a high number of S-S-bonds. These intra- or inter-chain S-S-bonds allow the protein polypeptide chains to aggregate into dense molecules and may be important for the formation of protein aggregates. Polyacrylamide gel electrophoresis (PAGE) analysis has shown that proteins in PB-II aggregates contain molecular weights of 64, 140, 240, 320, 380 and 500 Kda and even more than 2000 Kda [5]. Molecular biology studies have shown that the first protein molecule synthesized during gene expression of rice storage proteins is a molecular weight of 57KDa, which is then cleaved into two subunits of 22KDa and 37KDa. The protein molecules of different sizes in gluten are assembled by these two subunits through the - S - S - assembly [6].SDS can destroy the - S - S - connection, and by changing the amount of SDS, the molecular weights of 22-23KDa and 37-39KDa can be found, so these two components are actually the basic units of the macromolecular aggregates [5].

Protein fractions with molecular weights as high as 100 KDa are also present in clear proteins, but due to the low cystine content of clear proteins, it is not easy to form the - S - S - bond, and thus clear proteins are more soluble in water, which suggests that the presence of disulfide bonds is very important for stabilizing the protein aggregates.

After extracting the proteins and analyzing their amino acid composition, it was found that some proteins in rice are not simple proteins composed exclusively of amino acids, but are bound proteins containing sugars (rhamnose) or lipids [7]. These non-amino acid components not only affect the properties of proteins, but also give them special physiological functions.

In addition, a large number of studies have shown that the types of proteins in rice are not fixed. During the aging process of rice, although the total protein content remains unchanged, its structure and type change, which in turn affects the rheological properties of rice. The prominent changes are the increase in the number of disulfide bonds, the increase in the molecular weight of proteins, and the densification of protein aggregates. The network structure of proteins and starch is dense after cooking, which restricts the water-absorbing expansion and softness of starch grains, and thus the viscosity of the rice decreases while the hardness increases. The viscosity of rice decreased and the hardness increased.

At this time, the viscosity of rice was improved by adding an appropriate amount of reducing agent to break the disulfide bond [8 - 11]. Ren Shuncheng et al. also demonstrated this change in the molecular weight of proteins before and after aging by SDS-PAGE [12], and Teo et al. demonstrated that the change in proteins in rice is an important factor leading to the change in the rheological properties of rice [13]. These experiments have shown the importance of the S-S-bonds on the properties of proteins.

Rice proteins not only form larger molecules during aging, but there is also significant polymerization of protein molecules when heated.Mujoo noted that molecules with molecular weights of 24, 34, and 68 KDa could be polymerized into very large aggregates of 4 × 104 KDa when popping rice flowers, but alcohol-soluble proteins with molecular weights of 13-16 KDa did not participate in this proteasomal formation [14].

Thus, the development and utilization of rice proteins should pay particular attention to the effects of aging, heating and the oxidation and reduction of disulfide bonds on the properties of proteins.

The contents of the four types of proteins in rice bran were significantly different from those in rice. The contents of clear, globular alcohol-soluble, acid-soluble and alkaline-soluble proteins obtained by sequential extraction with square, salt number, according to alcohol, acid and alkaline solutions were 34%, 15%, 6%, 11% and 32%, respectively, of which acid-soluble and alkaline-soluble proteins were gluten proteins, that is to say, the content of water-soluble proteins in rice bran was very high.

Analysis shows that the molecular weight range of the first four proteins are 10-100KDa, 10 ~ 150KDa, 33 ~ 150KDa and 25 ~ 100KDa. The molecular weights of the main components of alkali soluble proteins are still distributed in the range of 45~150KDa even though the disulfide bonds are broken during the process of extraction, and the molecular weights of all these cereal proteins are much larger in molecular weight and harder to be soluble in water. However, if the disulfide bond is broken, more than 98% of rice bran protein can be dissolved [15]. It should be pointed out that before and after rice bran is stabilized (generally heat inactivation), the content of its various protein components changes greatly, mainly manifested in the decrease of clear protein content (due to denaturation), glutenin content increases obviously [16].

2 Nutritional value of rice protein

Rice protein is recognized as high quality food protein, mainly because the amino acid composition of rice protein is balanced and reasonable, conforming to the ideal pattern recommended by WHO/FAO, and the content of methionine is high, which is incomparable with that of other plant proteins. Rice protein and rice bran protein have high biological value, and their nutritional value is comparable to that of eggs and cow's milk.

In addition, rice protein is a low antigenic protein that does not cause allergic reactions, which is very favorable for the production of infant foods. Rice protein powder for infants and young children is available in many countries around the world. Many plant proteins contain antinutritional factors, such as trypsin inhibitors and hemagglutinins in soy protein and peanut protein, a type of clear protein in wheat, and bromelain in pineapple, which often cause an immune response that can lead to allergic or toxic reactions in the eater. There are also allergenic factors in animal foods, such as lactoglobulin in milk and ovalbumin in egg whites, to which infants and young children are most sensitive. In contrast, rice proteins are the safest, and rice is the only cereal that can be exempted from allergy testing [17]. As the technology of rice protein research is improving, rice protein-fortified foods for infants, children and the elderly are becoming more popular in the market.

In addition to its basic nutritional functions, rice protein also has other health benefits.Morita's experiments with rice protein isolate (RPI) and casein in mice showed that RPI significantly reduced serum concentrations of cholesterol, glycerol and phospholipids, and that liver weights were lower than those of the casein-fed group [18].

Dimethylbenzanthracene (DMBA) is a mutagen for breast cancer. Mice were fed 30 mg DMBA/Kg body weight, and the proteins in the basal diet were RPI, soybean protein isolate (SPI) and casein. The results showed that the tumor weight of RPI-fed mice was lower than that of casein-fed mice, and there was no significant difference in the activity of phenol hydroxylase in the serum of the mice in each group at 7 days of feeding. This indicates that RPI has the effect of resisting DMBA-induced carcinogenesis [19]. RPI extracted from rice bran also showed the same effect [20]; further analysis of the composition of RPI by chromatography and mass spectrometry found that there were triterpene alcohols, ferulic acid and other components in RPI [21], which indicated that RPI was a binding protein. These non-amino acid components may be necessary for the specific action of the protein.

Neriega's experiment is also interesting. He compared the endurance of sub-maximal physical training in people who consumed rice and bread, and found that those who consumed rice had greater endurance and lower levels of lactic acid in their blood [22].

Rice bran also has anti-diabetic properties. Streptozotocin (StrePt0z0t0cin, STZ) is inducer of diabetes. In the function test of rice bran, it is found that feeding rice bran to experimental rats for two months can significantly reduce the symptoms of diabetes induced by STZ, the content of glycerol and cholesterol in serum of experimental rats is lower than that in the control group, and the symptom of polyuria is also improved. It can be inferred that the protein in rice bran plays an important role [23].

The above studies have shown that rice protein not only has unique nutritional functions, but also has many potential health care effects. This is one of the important reasons why foreign countries attach great importance to the research and development and utilization of rice protein. Domestic research on the functionality of rice protein is relatively small.

3 Exploitation and utilization of rice protein

The main component of rice is starch, and the protein content is only about 9%, so it is obviously uneconomical to extract protein directly from rice. The protein content of the residue (i.e. rice residue) in the production of rice starch sugar and monosodium glutamate (MSG) is 40% ~ 65%, which can also be called rice protein concentrate, and it is the main raw material for the development and utilization of rice protein, which is a large amount of valuable resources. In the past, it was mainly used as protein feed for animals, but from the perspective of resource utilization, this is not economical. As the value of rice protein is recognized, more and more of it is being developed into high value-added raw materials and additives for food production. High-protein nutritious rice flour is available on the market, but it is still mainly composed of starch, with a very low protein content, and its potential for development and utilization as a protein resource has not been fully exploited.

3 . 1 Rice isolate protein (RPI)

Rice Protein Concentrate (RPC) has been found to contain more than 40% protein, but many of its functional properties are unsatisfactory. The removal of carbohydrates by chemical or biochemical methods has resulted in rice protein isolate (RPI) with a protein content greater than 90%, which can be hydrolyzed or biochemically modified to produce a variety of edible protein supplements. Since most of the RPC is water-insoluble protein, the traditional method of extraction is alkaline solubilization and acid precipitation, which can produce RPI with high purity, but has obvious shortcomings, such as the dark color of the product, the destruction of lysine in the protein, the formation of bitter-tasting and toxic substances as a result of side reactions, and the low recovery of proteins.

Based on the fact that the proteins in RPC are water insoluble and the non-protein components are mainly carbohydrates, the extracted proteins should be further purified (Purificati0n). Treatment with cellulases, pectinases and isoamylases can also be used to promote more carbohydrate solubilization. This approach in rice amylose production has resulted in higher yields of amylose as well as high purity RPI and satisfactory protein recovery [24 - 26].

Protein content in rice bran is 10% ~ 12%, as mentioned before, about 35% of it is water-soluble, but because there are a large number of fibers in rice bran, and most of rice bran has been stabilized, and the solubility of protein has been changed greatly by heating, and it is difficult to extract it effectively, and the current research on this problem is mainly the homogenization treatment of rice bran and the application of enzyme technology. The milling size of rice bran has great influence on the solubility of protein, especially the rice bran without heating treatment. It is pointed out that grinding and homogenization can dissolve 38% of protein, which is 75% higher than the original dissolved amount, and the molecular weights of dissolved components are very different [27].

Bio-enzymes are used in the extraction of rice bran protein with more obvious effect, and the enzymes that can be used include cellulase, ligninase, protease and phytase. Cellulase, ligninase can release the rice bran cellulose to protein binding, make the extract of protein content can be up to 50% above [28 - 29]. If use phytase and cellulase, ligninase and so on combined treatment of defatted rice bran, will obtain the protein content of 92% rice bran separated protein (RBPI) its yield can reach 74 . 6% [30]. 6% [30].

The application of protease can also achieve satisfactory results. Hamada et al. treated rice bran with protease to achieve a protein hydrolysis degree (DH) of 10%, and the protein extraction rate was 92%. If Na2 SO3 and SDS are applied to break the protein disulfide bonds, even if the degree of hydrolysis is only 2%, the protein recovery can reach 84% [31]. The use of two or more proteases with different hydrolysis sites in the extraction process resulted in protein hydrolysates with better physical and chemical properties than those obtained with one enzyme [32].

The above experiments were conducted to improve the extraction of protein by increasing the solubility of protein, and the foaming and emulsifying properties of the protein obtained were also improved to some extent. This is obviously different from the technical direction of enzymatic extraction of rice protein and the nature of the product.

3 . 2 Rice protein foaming powder

More than ten years ago, the emergence of rice protein foaming powder provided an option for the large-scale application of rice protein in food production. However, this kind of foaming powder is made from rice protein concentrate, and the product is hydrolyzed with NaOH, which is dark in color, high in pH value, and bitter in taste. The above disadvantages can be overcome by using protease to hydrolyze rice protein. Rice protein has a high molecular weight and contains a lot of hydrophobic amino acids, so its solubility is poor and it cannot show its physical and chemical functionality. After proper hydrolysis with protease, more - COOH and - NH2 can be released, increasing the polarity of protein molecules, which will increase the solubility of the protein and at the same time, the colloidal nature of the solution will also be enhanced, showing a certain degree of emulsification and foaming ability, which can be widely used as a raw material for food processing, and give the food a certain degree of processing performance.

It can be widely used as food processing raw materials, giving certain processing performance to food. At present, the hydrolysis of soybean protein and wheat gluten protein has been studied in China. Wang Zhangcun and others used protease to hydrolyze soybean isolate protein and achieved good results [33]. Enzymatic hydrolysis of rice protein as raw material for the production of edible foaming powder has also been reported in recent years [34]. It is believed that with the improvement of technology, enzymatic rice protein foaming powder will be widely used in food production.

3 . 3 Protein hydrolysates

It takes rice protein as raw material, and through different degrees of hydrolysis, it can obtain protein hydrolysate with different uses, most of which can be used as ready-to-drink protein nutrient fortification, and some of which contain special flavor or health care functions.

The preparation of amino acid nutrient solution is a traditional method of utilizing plant protein, and the acid hydrolysis method is more researched and utilized domestically, and the so-called chemical soy sauce is based on this method, but it should be eliminated due to environmental protection and safety problems. Protease hydrolysis is limited by enzyme specificity, and there is no single enzyme that can completely hydrolyze proteins, so it is not economical to use multiple enzymes.

In fact, it is not necessary to completely hydrolyze proteins in nutritional products aimed at replenishing amino acids, but only to hydrolyze them into small peptides. Nutritional studies now show that small peptide molecules are more easily absorbed and utilized by the small intestine than amino acids. Absorption of peptides is achieved through the active transport of peptide carriers in the striatal margin of the intestinal mucosa using a proton gradient. Small peptides have a low permeability pressure, do not cause dysentery or allergic reactions, and have a better sensory effect than amino acids, making them suitable for use as protein nutritional enhancers. NutriBiotics Rice Protein Powder, which is well known in the US, is one such product.

More interestingly, many small peptide molecules have important physiological functions, such as immunomodulation, antioxidant, anticholesterol, antithrombotic, antidiabetic and so on, which are also known as active peptides. Currently, the hydrolysis of animal proteins to produce biopeptides has become a worldwide trend, and many active peptides with potential applications have been identified [35]. However, there are relatively few studies on active peptides from rice, and one of the most widely reported active peptides from rice is the Gly-Tyr-Pro-Met-TYR-Pro-Leu-Arg peptide molecule named oryzatensin, which has been shown to have ileocecal contraction-inducing, anti-morphine, and immunomodulatory effects in guinea-pig experiments. It induces contractions mainly by activating phospholipases that hydrolyze lysophosphatidic acid to release arachidonic acid [36].

In addition, hydrolysis of rice protein can produce certain flavor peptides. Modern instrumental analysis shows that the content of glutamic acid in these flavor peptides is very high, and it combines with salt to form monosodium salt of glutamic acid, which presents fresh flavor. When rice protein is hydrolyzed, this product is mixed with dextrin and spray-dried to obtain commercially available food flavor modifiers [37]. 3 . 4 Chemical modification of rice protein

The physical and chemical functionality of natural plant proteins is generally poor. Researchers are interested in chemically improving the properties of proteins to increase their use in foods. This would meet the needs of food processing performance and increase the nutritional value of the food. Many studies have been conducted on the modification of soy proteins. The main methods are the introduction of phosphoric acid and acetyl groups, or the removal of amide groups such as glutaminylammonium and asparagine from proteins, and these measures are safe and effective. These measures are safe and effective. However, the chemical modification of rice isolate protein has not been reported.

In summary, rice protein is a valuable protein resource that needs to be vigorously developed. It is a protein polymer molecule consisting of a large number of disulfide bonds, and rice protein and its hydrolysates have not only important nutritional functions but also potential health care effects. Enzymatic hydrolysis and chemical modification of rice proteins can improve their physical and functional properties. These products have broad application prospects. Foreign countries have conducted more research on rice protein and achieved certain results. It is believed that China's research and development of rice protein will also have greater progress.

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