At the beginning of this century, clinics began to measure enzymes in body fluids to diagnose diseases. For example, Wohlgemuth determined urine amylase (AMY) as early as 1908 to diagnose acute pancreatitis. In the 1930s, alkaline phosphatase (ALP) was clinically used to diagnose skeletal diseases. Later, it was found that many liver and gall bladder diseases, especially when obstructive jaundice appears, this enzyme is often significantly increased. These enzymes became routine test items in clinical laboratories at that time. Until the 1960s, ALP was still the most frequently measured enzyme in the world. However, before the 1950s, enzyme measurement accounted for only a small part of the routine work of the laboratory.

The real development of diagnostic enzymes began with the continuous monitoring of enzyme activity concentration by spectrophotometry in the 1950s. It can measure many enzymes that cannot be measured with the old “fixed time method” and is used to diagnose diseases. The results showed that the sensitivity of lactate dehydrogenase (LD), aspartate aminotransferase (AST) and α-hydroxybutyrate dehydrogenase (HBDH) in the diagnosis of acute myocardial infarction (AMI) far exceeded that of other diagnostic methods. In the early 1960s, it was affirmed that creatine kinase (CK) increased in diagnosis of AMI earlier than the above enzymes, and the specificity was also high. At present, this enzyme has replaced ALP as the most frequently measured enzyme in the world. At the same time, it was found that alanine aminotransferase (ALT) and AST are not only highly sensitive to hepatitis diagnosis, but also significantly increased as early as the early stage of hepatitis and jaundice.

These achievements aroused the widespread interest and attention of clinical and laboratory workers at that time. They have carried out a lot of clinical and experimental work, tried and evaluated the clinical significance of hundreds of enzyme determinations, of which about ten enzymes have become important measurement items commonly used in the laboratory. Enzyme determination accounts for about 1/4 to 1/2 of the current total workload of clinical chemistry.

With extensive application and research, it has also been found that the specificity of the total enzyme activity concentration measurement for disease diagnosis is far from being as high as people initially expected. Since the 1970s, scholars have gradually focused their attention on the determination of isozymes, and found that the diagnosis of CK-MB and LD1 is more specific than the above total enzymes. CK-MB has become a recognized “gold index” for the diagnosis of AMI. The determination of these two isozymes has also become a mandatory test item in major hospitals.

Since the 1980s, it has been found that there may be changes after isozymes in tissues enter body fluids. For example, Ck-MM can be further divided into Ck-MM1, MM2, and MM3, and Ck-MB can be divided into MB1 and MB2. It is superior to CK total enzyme and isoenzyme in the diagnosis of AMI, and has become a research hotspot in clinical enzymology.

For a long time, the clinical understanding of the mechanism of serum enzyme changes has been very simple: that is, diseased cells release high concentrations of enzymes in their cells into the blood. The greater the enzyme concentration gradient between the two, the greater the increase in serum enzymes. This is far from explaining various clinical phenomena. For example, the absolute amount of AST in the liver is about 4 times that of ALT, but in acute hepatitis ALT increases much more than AST. However, in chronic liver disease, especially cirrhosis, the blood AST is higher than ALT, which is obviously difficult to clarify from the above concentration gradient theory. We must fully understand the various factors that affect the changes in serum enzymes, as well as understand the classification of serum enzymes, because the different types of enzyme change patterns are different.

Enzymes are an active substance produced by organisms and catalysts for various biochemical reactions in the body. The digestion, absorption and utilization of various nutrients must depend on the action of enzymes. Enzymes have the characteristics of specificity, high efficiency and specificity. At present, there are thousands of enzymes found, more than 300 kinds can be produced artificially, and more than 20 kinds are used in the feed industry. Bioactive methods are used to produce active enzyme products, called enzyme preparations. Enzyme preparations are a kind of feed additives widely used in feed in recent years, most of which are digestive enzymes. Enzyme preparation can effectively improve feed utilization rate, save feed material resources, and has no side effects, so it is an environmentally friendly green feed additive with a broad market prospect and application potential.

The Main Types of Enzyme Feed Additives

Amylase. Amylase mainly includes α-amylase and saccharification enzyme. Alpha-amylase can decompose starch macromolecules into easily absorbed medium and low molecular substances. The saccharification enzyme can further hydrolyze the medium and low molecular substances decomposed by α-amylase into glucose, which is absorbed and utilized by animals.

Protease. Proteases are hydrolytic enzymes that degrade protein peptide chains, mainly including pepsin, trypsin, papain, etc.

Cellulase. Cellulase can destroy the crystalline structure of cellulose, hydrolyze cellulose macromolecules into oligosaccharide fragments, and decompose oligosaccharide materials into glucose.

β-glucanase. Beta-glucan is widely present in a variety of plant raw materials, and has a high viscosity. It is an important antioxidant factor that affects the transmission and absorption of nutrients. Beta-glucanase can hydrolyze large molecules such as glucan, reduce the viscosity of substances in the digestive tract, and promote the absorption of nutrients. β-glucanase is an important and widely used enzyme in feed additives for enzyme preparations.

Pectinase. Pectin is an anti-nutritional factor in plant raw materials, which affects the utilization rate of feed. Pectinase can effectively destroy pectin quality and promote the digestion and absorption of nutrients. Pectinase is also a more commonly used feed enzyme preparation.

Phytase. The vast majority of phosphorus in cereals is in the form of phytate phosphorus. Animals do not secrete phytase, so the utilization rate of phosphorus in cereals is low. By adding phytase secreted by microorganisms in the feed, this part of phosphorus can be decomposed and released, thereby reducing the amount of inorganic phosphorus added to the feed and reducing the cost of feed. And it can reduce the excretion of phosphorus in animal feces and reduce environmental pollution. It is a green feed additive with more applications and the best prospects.

Compound enzyme. A compound enzyme is a product obtained by mixing two or more enzymes with biological activity. The compound enzyme is formulated according to the characteristics of different animals and different growth stages, has a good effect, and is currently the most commonly used feed additive.

The Role of Enzyme Feed Additives

Direct decomposition of nutrients, improve feed utilization. Active multiple enzymes can effectively decompose and digest some molecular polymers in feed into nutrients easily absorbed by animals, or into small fragments of nutrients for further digestion by other digestive enzymes. Some macromolecular substances are difficult for animals to decompose and absorb, so adding enzyme preparations can promote the decomposition and digestion of nutrients in feed, thereby improving feed utilization.

Eliminate anti-nutritional molecules and improve digestive function. There are some non-starch polysaccharides, pectin, phytic acid, and cellulose polymers in plant raw materials. These substances increase the content and viscosity of the animal’s digestive tract, affecting the animal’s digestion and absorption of effective nutrients. Various enzymes in enzyme preparations, especially β-glucanase, pectinase, phytase and cellulase can decompose these substances into small molecules. Thereby reducing the viscosity of substances in the digestive tract, and effectively eliminate the adverse effects of these anti-nutritional factors, improve the digestive performance of animals.

Due to the use of enzyme preparations, more substrates for various enzymes can be provided, thereby activating the secretion of various digestive enzymes in the animal body to increase the effective content of digestive enzymes. And accelerate the digestion and absorption of nutrients, thereby improving feed utilization and accelerating animal metabolism, and promote animal growth.

Simple Proteinases and Conjugated Proteases

Proteins are divided into simple proteins and combined proteins. Similarly, according to chemical composition, enzymes can also be divided into two major categories: simple proteinases and conjugated proteases. General hydrolytic enzymes, such as urease, protease, amylase, lipase, ribonuclease, etc., are simple proteases. The activity of these enzymes depends only on their protein structure. The enzyme is composed only of amino acids and contains no other components. And transaminase, lactate dehydrogenase (LDH), carbonic anhydrase and other oxidoreductases are all bound proteases. In addition to protein components, these enzymes also contain non-protein small molecules that are stable to heat. The former is called apoenzyme, the latter is called cofactors. When enzyme protein and cofactor exist alone, there is no catalytic activity. Only when the two are combined into a complete molecule can they have enzyme activity. This complete enzyme molecule is called holoenzyme.

holoenzyme = enzyme protein + cofactor

Some of the cofactors of enzymes are metal ions and some are small molecule organic compounds. Sometimes both of them are required for enzyme activity. These small molecule organic compounds are usually called coenzymes or prosthetic groups. The metal is in the enzyme molecule, either as a constituent of the active site of the enzyme, or to help form the conformation necessary for enzyme activity. Enzyme proteins use polar groups on their side chains to bind cofactors through covalent, coordinate, or ionic bonds through reaction. Generally speaking, the small molecule organic substances that are loosely bound to the enzyme protein and easily detached from the enzyme protein and can be removed by dialysis are called coenzymes; and the small molecule substances that are tightly bound to the enzyme protein and are not easily removed by dialysis are called prosthetic groups. There is no essential difference between coenzymes and prosthetic groups, and there is no strict boundary between the two, except that they are strongly bound to the enzyme protein.

In the catalytic reaction of holoenzymes, enzyme proteins and cofactors play different roles. The enzyme protein determines the specificity and high efficiency of the enzyme reaction, and the cofactor directly acts as a carrier for electrons, atoms or certain chemical groups, participates in the reaction and promotes the entire catalytic process.

Usually an enzyme protein can only be combined with a coenzyme to form an enzyme, which acts as a substrate to carry out a chemical reaction in one direction. A coenzyme can be combined with several enzyme proteins to form several enzymes, catalyzing the same type of chemical reaction of several substrates. For example, the enzyme protein of lactate dehydrogenase can only be combined with NAD to form lactate dehydrogenase, which makes the substrate lactic acid dehydrogenate. But there are many kinds of enzyme proteins that can bind to NAD, such as lactate dehydrogenase, malate dehydrogenase (MDH) and glycerophosphate dehydrogenase (GDH) contain NAD, which can catalyze separately Lactic acid, malic acid and glycerol phosphate are dehydrogenated. It can also be seen that the enzyme protein determines the type of reaction substrate, that is, the specificity of the enzyme, and the coenzyme (base) determines the reaction type of the substrate.

Monomer Enzyme, Oligomerase and Multi-enzyme Complex System

According to the structural characteristics of proteins, enzymes can be divided into three categories:

Monomer enzyme

Enzymes with only one polypeptide chain are called monomeric enzymes, and they cannot dissociate into smaller units. Its molecular weight is 13 000 ~ 35 000. There are few such enzymes, and most of them are enzymes that promote the hydrolysis reaction of the substrate, that is, hydrolytic enzymes, such as lysozyme, protease, and ribonuclease.

Oligomerase

Enzymes composed of several or more subunits are called oligomeric enzymes. The subunits in the oligomerase can be the same or different. The subunits are connected by non-covalent bonds, which are easy to separate for acids, bases, high-concentration salts or other denaturants. The molecular weight of oligomerase ranges from 35,000 to several million. Such as phosphorylase a, lactate dehydrogenase, etc.

Multi-enzyme compound system

The complex formed by the chimerization of several enzymes with each other is called the multienzyme system. The multi-enzyme complex is conducive to the continuous progress of a series of reactions in the cell to improve the catalytic efficiency of the enzyme, and at the same time it is convenient for the body to regulate and control the enzyme. The molecular weights of multiple enzyme complexes are all above several million. Such as pyruvate dehydrogenase system and fatty acid synthetase complex are multi-enzyme systems.

The enzymes are the general term for important protein biochemical substances. Both plants and animals have enzymes. As early as more than 100 years ago, scientists have isolated a physiological enzyme, trypsin, from the digestive tract fluids of animals. In a test tube, this substance can break down various proteins into amino acids. As a result of extensive participation in various physiological activities, enzymes have been more and more widely used in medicine. In recent years, foreign research and application have been on the rise.

In the last century, with the advent of cephalosporin β-lactam antibiotics, the serious environmental pollution caused by the production of cephalosporins is worrying. At that time, British and Dutch researchers first reformed the chemical lysis method that has been used for many years into a clean and efficient “enzymatic lysis”, that is, the enzymes used are penicillin lyase and cephalosporin lyase. Thereby eliminating the environmental pollution caused by the production of semi-synthetic penicillin or cephalosporin products. These two enzymes can be used to prepare a series of semi-synthetic penicillins such as ampicillin and amoxicillin and other key raw materials for various cephalosporins. With the implementation of enzymatic cracking process, antibiotic production has entered a new “enzymatic production era”.

The enzyme can be used not only in the cleavage process, but also in important reactions such as condensation and carboxylation. Creative Enzymes is the world’s leading research and producer of enzyme preparations. The new industrial enzymes invented by the company can be used to produce chiral amino acids, an important raw material for the synthesis of various chiral drugs.

The rise of functional oligosaccharide health products increases the amount of raw materials. In recent years, many new functional oligosaccharide products have been developed at home and abroad, which have become new raw materials for health food. Functional oligosaccharides include mannan, xylan, dextran, chitosan (chitosan), soy glycans, fructooligosaccharides, etc. It has a variety of health effects such as improving human immunity, anti-cancer, anti-aging, lowering blood fat and removing intestinal waste, etc., which has greatly promoted the demand in the international market. Developed countries such as Japan and the United States have become the world’s largest producer and consumer of functional oligosaccharides. The hot sale of functional oligosaccharides has further promoted the research and development of new functional oligosaccharide enzymes and new oligosaccharide products.

In the past 10 years, developed countries led by the United States, the United Kingdom, and Japan have successively developed and marketed a variety of medical enzyme preparations and have formed a large-scale medical enzyme market. Its annual sales have exceeded $ 1 billion. There are at least dozens of brands of digestive enzyme preparations for sale in the European and American markets. In addition, there are some new types of enzyme preparations for medical use, such as the anti-trypsin developed by the United Kingdom, which is one of the leading ones. It is used to treat emphysema, rheumatoid arthritis, ulcerative colitis and other diseases.

In recent years, some therapeutic enzyme preparations such as antibacterial enzymes, fibrinolytic enzymes, mucolytic enzymes, analgesic enzymes, antitumor enzymes, immune activator enzymes, etc. have been newly developed. In addition, the researchers also found that combining protease preparations with existing antibiotic preparations such as ampicillin, tetracycline, SMP-CO and fluoroquinolone antibacterial agents can greatly improve the antimicrobial effect of these drugs and reduce bacterial resistance .

Some hospitals have combined enzyme preparations with antibiotic preparations to treat intractable urinary tract infections and achieved good results. These new clinical uses have laid a solid foundation for the medical market for enzyme preparations. Medical enzymes such as diagnostic enzymes have become a new type of therapeutic agent at home and abroad. With the advent of more and more new enzymes, medical enzymes will become a new bright spot in the growth of the international pharmaceutical market.