Several Methods of Synthesizing Peptides
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Protein Hydrolysis
The method of using certain hydrolysis methods to break the protein peptide chain of macromolecules to generate small molecular peptides with different activities is protein hydrolysis. The preparation of peptides by protein hydrolysis is currently the most important production method for peptides. Its hydrolysis is easy to control, and specific peptides can be produced in a targeted manner. The product has high safety and low cost. At present, there are mainly two methods for preparing peptides by hydrolyzing proteins, namely chemical hydrolysis and enzymatic hydrolysis.
- Chemical Hydrolysis
The chemical hydrolysis method uses acid and alkali to hydrolyze protein. It is a traditional method of protein hydrolysis. It is to mix protein with a certain concentration of acid or alkali solution, heat it to a certain temperature and keep it for a period of time to crack the peptide chain of the protein. Although this method is easy to operate and low in cost, due to severe conditions, the damage to proteins or peptides is great, and there is no rule in the degradation. At the same time, the structure of some amino acids may also change, making L-type amino acids into D-type amino acids. Lys-Ala can form toxic substances, and it is difficult to control the degree of hydrolysis, so this method is basically not used in production. - Enzymatic Hydrolysis
Peptides with specific activity can be generated by enzymolysis and digestion of protease. The enzymatic hydrolysis has mild reaction conditions, avoiding the more extreme reaction conditions in the chemical hydrolysis, and has the advantages of low salt content of the obtained enzymatic hydrolysis products, no amino acid destruction and racemization, non-toxic by-products, and no pollution to the environment. Therefore, whether in the laboratory or in industrial production, enzymatic hydrolysis is the most commonly used method for preparing peptides.
The selection of protease in the process of enzymatic hydrolysis is the key to the preparation of peptides. The main source of protease is microorganism, followed by plant and animal. Microbial protease, such as alkaline protease, flavor protease, complex protease, etc., has the characteristics of good solubility, high activity, not strong specificity, and strong effect on proteins. Plant-derived proteases include bromelain, papain, sisal protease, etc. These proteases have many action sites and are not specific, but their sources are abundant and the price is relatively cheap, so they are widely used in production. Animal-derived proteases, such as trypsin, pepsin, etc., have strong specificity, but due to limited sources, high extraction technology requirements, the price is generally relatively expensive.
Enzymatic production of peptides is the focus of current research, and it is also the most commonly used method to obtain bioactive peptides. Enzymatic hydrolysis is to select appropriate proteases to hydrolyze various proteins to obtain active peptides, which can improve the physicochemical properties and functions of proteins without reducing the nutritional value of food proteins, and expand its application in food processing. Enzymatic hydrolysis has many physical and chemical modification incomparable advantages, its reaction conditions are mild, reaction time is short, high efficiency, under certain conditions for positioning hydrolysis, the product is mainly peptide rather than amino acid, hydrolysis process is easy to control, product purity is high, easy to separate, low cost, so it can better meet the needs of peptide production. Enzymatic hydrolysis is a common method for obtaining food grade bioactive peptides. At present, a variety of bioactive peptides with different functions have been prepared from the enzymolysis of bone protein, casein protein, whey protein, soybean protein, corn protein and aquatic protein.
Synthesis Method
Synthesis is a method in which people have known the sequence of a specific active amino acid and use it to produce a specific active peptide. Synthesis of peptides can be divided into chemical synthesis, enzyme synthesis and gene recombination.
- Chemical Synthesis
Chemical synthesis of peptides is mainly through solid phase or liquid phase synthesis method to obtain the required bioactive peptides. Solid phase synthesis is often used in the laboratory for the study of proteins, peptides and other aspects, which is characterized by high product purity, high cost, low yield, short direct synthesis sequence, low synthesis efficiency, and high toxicity of synthetic reagents, so this method is generally limited to laboratory preparation. The solid phase synthesis method can only be used for the synthesis of peptides with fewer amino acids and smaller molecular weight, and it is still difficult to obtain macromolecular peptides completely by solid phase synthesis method. Compared with solid phase synthesis method, liquid phase synthesis method has the advantages of high product purity, large yield, suitable for industrial production, after continuous improvement and perfection, can be used in the production of high value of short-chain or medium-chain health-care active peptides products. However, the cost of chemical synthesis is relatively high, it is difficult to achieve large-scale production. It is inevitable that organic reagents will be used in the synthesis process, which will cause environmental pollution, many side reactions, and the synthesis of target peptides is more toxic. Many factors limit the development of chemical synthetic peptides. - Enzyme Synthesis
Enzyme synthesis uses protease to catalyze the synthesis of peptides, which is mostly used to synthesize relatively short peptides. Enzymatic synthesis reaction conditions are mild, the toxicity of the agents used is low, the product will not racemize, but the reaction by-products are more, the yield is low. - Gene Recombination
Gene recombination is often used to synthesize peptides with relatively long peptide chains, especially those consisting of several hundred amino acids. Gene recombination can recombine the gene sequence regulating the synthesis of peptides into the genetic material of microorganisms, etc., and translate the genetic material into the recipient organism. The advantage of this method is that it can reduce the production cost of expensive or low-yield peptides obtained by natural methods through gene recombination, and it is high safety, but the genetic recombination preparation of peptides is difficult in the preliminary research technology, and has not been applied to industrial production.
Microbial Fermentation
Microbial fermentation refers to the method of enzymolysis of proteins by using some proteases produced by microorganisms in the fermentation process to obtain bioactive peptides. The factors affecting the preparation of active peptides by microbial fermentation mainly include the preparation of medium, screening of strains, fermentation temperature, fermentation time and age of strains, among which the key technology is the selection of strains. The difference of bacterial species can directly lead to the difference of protease types, and the difference of protein tangent point can directly affect the biological activity of the produced peptides. Bacterial strains commonly used in microbial fermentation are Bacillus subtilis, Aspergillus Niger, etc., the source of microbial strains is relatively wide, the protease produced does not need to be separated, the production cost is lower than that of enzymatic hydrolysis, and the peptide obtained by microbial fermentation does not produce bitter taste, which greatly reduces the process. However, many of the molds produced by microbial fermentation are toxic or harmful to human body, and there are not many strains that have been proved to be non-toxic and can be used for food development. Safety issues have become the biggest obstacle to the preparation of active peptides by microbial fermentation. At the same time, due to the strict control conditions in the microbial fermentation process, the intermediate metabolites and end products in the fermentation metabolism process will inhibit the enzyme synthesis, and the protease system generated by metabolism is determined by the strain and fermentation conditions and other conditions, and can not fully meet the preparation of target active peptides, and it is difficult to isolate and purify the target active peptides.
