Among the hundreds of famous enzyme companies in the world, NOVO of Denmark firmly holds the leading position with more than 50 % of the market share, followed by Genentech with about 25 % of the market share, and other enzyme producers in other countries share the remaining 25 % of the market share.
Enzymes used in industry are basically divided into two categories: hydrolytic enzymes, including amylase, cellulase, protease, lipase, pectinase, lactase and so on, which account for more than 75% of market sales. Currently, more than 60% of the enzymes are produced by genetically modified strains, and 80% of the strains used by NOVO are recombinant strains. The second category is non-hydrolytic enzymes, which account for about 10 % of market sales and tend to increase year by year, mainly enzymes for analytical reagents and enzymes for the pharmaceutical industry.
In the food industry, the proportion of enzymes used in starch processing is still the largest, 15 %; followed by the dairy industry, accounting for 14 %. The traditional application of enzymes in the food, textile and tanning industries is still evolving, although it is already quite extensive and technically mature. The following is a brief introduction to the production safety of enzymes and new developments in industrial applications in recent years:
1 Safety and hygiene management of enzyme production
With China's accession to the WTO just around the corner, the safety and hygiene management of enzyme production must not go unnoticed. Enzymes for food are used as food additives in foreign countries, and their safety and hygiene regulations are very strict. Although the enzyme itself is a biological product, which is safer than chemical products, the enzyme preparation is not a simple product, but often contains culture medium residues, inorganic salts, preservatives, diluents, etc. The production process may also be affected by sand, sand, sand, sand, sand, sand, sand, sand, sand, sand, sand, sand, sand, sand, sand and sand. They may be contaminated with Salmonella, Staphylococcus aureus, and E. coli during the production process. In addition, it may contain biotoxins, especially aflatoxins, which may be produced by some strains of Aspergillus niger even. Aflatoxins are either produced by the strain itself or introduced by the raw material (moldy grain material). In addition, inorganic salts are used in the culture medium, which inevitably include mercury, copper, lead, arsenic and other toxic heavy metals. In order to ensure the absolute safety of the product, the raw materials, strains, post-processing and other processes should be strictly controlled. The production site should meet the requirements of GMP (Good Manufactur2ing Practice). For the safety requirements of enzyme products, the Food and Agriculture Organization of the United Nations (FAO) and the World Health Organization (WHO) Expert Committee on Foodadditives (Joint FAO/ WHO Expert Committee on Foodadditives, J ECFA) as early as the 21st session of the General Assembly of the WHO in 1978, put forward to the safety of enzyme sources of the assessment of the standards. :
(1) Enzymes from edible parts of plants and animals and from strains of bacteria traditionally used as food ingredients, or traditionally used in food, which meet appropriate chemical and microbiological requirements, may be considered as food without the need for toxicity testing.
(2) Enzymes produced by non-pathogenic, general food-contaminating microorganisms require short-term toxicity testing.
(3) Enzymes produced by unusual microorganisms require extensive toxicity testing, including long-term rat feeding.
This standard provides a basis for safety assessment of national enzyme production. Production strains must be non-pathogenic and not produce physiologically active substances such as toxins, antibiotics and hormones, and strains must be proven harmless in a variety of safety tests before they can be used in production. For the determination of toxins, in addition to chemical analysis, biological analysis is also required. The safety of additives in the UK is assessed by the Committee on Chemical Toxicity
(abbreviated COT), which makes recommendations to the government's advisory committee of experts, FACE (Food Additives and Contamination Committee). The COT is most concerned about the toxicity of strains, recommending that microbial enzymes be subjected to a minimum of 90 days of rat-feeding and that bioassays be carried out to a high standard. COT believes that strain modification is necessary, but bioassays should be performed after each modification. There are two regulatory regimes for enzymes in the United States: GRAS (General recognized as safe) compliant substances and food additive compliant. Enzymes that are recognized as GRAS substances can be manufactured as long as they comply with GMP. Enzymes that are considered food additives must be approved and registered in the Code of Federal Regulation (CFR, TheCode of Federal Regulation) before they can be marketed. GRAS applications are subject to two major evaluations, namely, technical safety and acceptability of product safety test results. In addition to the FDA's authority to conduct GRAS accreditation, any expert qualified to evaluate the safety of a food ingredient may also conduct the evaluation independently. Animal raw materials used in the production of food enzymes in the United States must meet the requirements of meat inspection and GMP production, while plant materials or microbial cultures must not leave residues in food that are injurious to health under normal conditions of use. The equipment, diluents, auxiliaries, etc. used shall be suitable for food. Production methods and cultivation conditions must be strictly controlled so that the production bacteria do not become a source of toxins and health hazards
In addition, in recent years, the world's food market to implement the KOSHER food certification system, i.e., kosher food system. With the KOSHER certificate, you can enter the market of the world's Jewish organizations. In the United States not only Jews, even Muslims, vegetarians, people allergic to certain foods, most of them also buy KOSHER food. KOSHER foods are prohibited from containing pig, rabbit, horse, camel, shrimp, shellfish, winged insects and reptiles. Processing KOSHER food enzymes should also meet the requirements of KOSHER food. Therefore, many foreign food enzyme preparations are in line with the KOSHER food labeling. It is important to pay attention to this in order to develop our enzymes overseas. KOSHER food compliance is approved by specialized authority, which is more strict than FDA.
2 New industrial uses of enzymes
2. 1 Manufacture of functional oligosaccharides
In the last 20 years, probiotics, mainly bifidobacteria and lactobacilli, and prebiotics, mainly oligofructose, isomaltose, and oligogalactose, have become popular in countries all over the world as the new-generation health foods. The annual sales volume of various functional oligosaccharides through enzymatic conversion has exceeded 100,000 tons. Functional oligosaccharides are those oligosaccharides which are not digested or difficult to be digested and absorbed by human body, and after ingesting them, they enter into the large intestine, and are selectively utilized by human body's own beneficial bacteria (bifidobacteria, etc.) in a preferential manner. It is also known as bifidogenic factor because it makes bifidobacteria multiply hundreds of times in the body and promotes the health of the host. These oligosaccharides are not utilized by Streptococcus mutans, which is the source of dental caries, so they do not cause tooth decay. A daily intake of 3-10 g of functional oligosaccharides can improve gastrointestinal function, prevent constipation and mild diarrhea, reduce the production and absorption of toxins in the intestines, and improve the body's immune function against diseases. Functional oligosaccharides are becoming a popular source of healthy sugar in the 21st century.
(1) Isomaltooligosaccharides: are indigestible oligosaccharides that are not broken down by saliva and pancreatic juice, but can be partially broken down and absorbed in the small intestine. The calorific value is about 70%~80% of sucrose and maltose. It has less direct irritation to the intestinal tract. The LD50 of acute toxicity test in mice is more than 44g/kg, the safety is not inferior to sucrose and maltose. The maximum non-effective amount in human body is 1.5 g/kg (limited to 24 hours after ingestion without diarrhea), while the maximum non-effective amount of other indigestible oligosaccharides or sugar alcohols is only 0.1~0.4 g/kg. After ingesting 16g of isomaltose for one week, the beneficial bacteria such as bifidobacteria and lactobacilli in the intestine obviously increased, while the harmful bacteria such as bacillus and clostridium were inhibited, constipation was improved, the fecal pH decreased, organic acid increased, spoilage was increased, and the fecal pH decreased, and the fecal pH decreased. The pH of feces decreased, organic acid increased and spoilage decreased. The test in mice showed that the immune system was enhanced and the blood lipids were improved after the intake of isomaltulose. Isomaltose is stable in high temperature, slightly acidic and acidic environments, and can be added to various foods and beverages.
Isomaltooligosaccharides are syrups of branched oligosaccharides such as isomaltose, pentose, and isomaltotriose, which are produced by liquefaction of starch by alpha amylase, saccharification by beta amylase, and transglycosylation by alpha glucosidase and contain alpha 1,6-bonded isomaltose. There are two types of isomaltose available on the market, 50 % and 90 %, the latter being produced by removing glucose from 50 % isomaltose by ion exchange or yeast fermentation. Powdered sugar is produced by spray drying the syrup.
The α-glucosidase for isomaltose production is a by-product of Aspergillus niger's saccharase production, and the saccharase fermentation broth is eluted and concentrated by removing the α-glucosidase by ion-exchange adsorption. Although there have been many published reports on the study of α-glucosidase production by culturing Aspergillus niger, it has not been used in commercial production. The production of isomaltooligosaccharides by converting maltose with α-glucosidase is generally only about 50 %, and contains 20-40 % of maltose and glucose. In order to increase the yield of isomaltooligosaccharides, a number of studies have been reported, such as the use of Aspergillus oryzae α-glucosidase, in which the yield of mannose can be as high as 30% and the amount of glucose can be reduced to 20%. Takasaki found that the enzyme produced by Bacillus thermophilus lipophilus had a transglycosylation effect in the presence of high maltotriose concentration. By introducing its structural gene into Bacillus subtilis NA-1, the new Pullulanase produced by the enzyme, together with Bacillus subtilis saccharifying α-amylase (which produces maltotriose), acted on starch, and the yield of isomaltooligosaccharides was up to 60%, and the amount of glucose was reduced from 40% to 20%. In order to improve the α-glucosidase activity of Aspergillus niger, the Department of Bioengineering of the University of Tokyo introduced the α-glucosidase gene, AGLA, into Aspergillus niger GN-3 and obtained the transformant, GIZ 155-A3-4, which increased the enzyme-producing capacity by 11 times.
At present, there are as many as 50-60 enterprises producing isomaltose in China, with a production capacity of more than 50,000 tons, and the amount of α-glucosidase is 0.1 %, which requires 50 tons, and consumes a huge amount of foreign exchange (750,000 yuan per ton, which requires 37.5 million yuan). It is necessary to base on self-sufficiency.
(2) Alginate: two molecules of glucose to α, α- 1. 1 bond linking non-reducing oligosaccharide. It is widely found in plants, animals and microorganisms (e.g., bacteria, seaweed, shrimp, brewer's yeast, baker's yeast), and is the main blood sugar of insects, utilized as a source of energy when flying. Alginate protects certain plants and animals from dry and frozen environments. Alginose is a good source of sugar, non-reducing, therefore acid and heat resistance, not easy to react with proteins and amino acids. It has strong inhibiting effect on aging of starch, denaturation of protein and oxidation of fat. In addition, it can eliminate the bitter and astringent taste of certain foods and the fishy odor of meat. Alginate is not utilized by Streptococcus mutans for dental caries, so it does not cause tooth decay. The survival rate of active dry yeast depends on the content of alginate in the yeast cells. In the past, alginate was extracted from yeast (the maximum content was only 20%) at a high cost of 20,000 to 30,000 yen per kilogram. Now, it is possible to produce it by enzyme or fermentation method, and the cost has been greatly reduced. Kubota et al. discovered a group of alginate-generating enzymes (alginate synthase MTSASE and malt oligosaccharides alginate hydrolase MTHASE) from soil bacteria such as Arthrobacter, Chlorella, Flavobacterium, and Sulfolobacterium, and obtained an 85% yield of alginate by combining these enzymes with isoamylase, cyclodextrinase, α-amylase, and saccharolytic enzymes in liquefied starch.
(3) Palatinose (Palatinose) scientific name for Isomaltotulose (Isomaltotulose): sucrose as raw material, through the prion or Serratia marcescens α-glucosyltransferase (also known as sucrose transformase Sucrose multase) action, sucrose molecules of glucose and fructose by the α-1 ,2 bonding The glucose and fructose of the sucrose molecule are converted from the α-1 ,2 bond to the α-1 ,6 bond. As a result of the structural change, its sweetness is reduced to 42% of sucrose, hygroscopicity is low, stability to acid is increased, heat resistance is slightly reduced, biological and physiological properties are changed, and it cannot be utilized by most bacteria and fungi. It is not decomposed by the enzymes in the mouth and stomach after eating, and can be hydrolyzed into glucose and fructose by the enzymes until the small intestine and enter into the metabolism. Para Gold is not utilized by Streptococcus mutans for oral caries, so it is not easy for tooth decay to occur, and blood sugar will not rise rapidly after eating, so it can be used by diabetic patients.
Pallakin Sugar will be condensed into 2~4 molecules of oligomeric Pallakin Sugar under low water and low pH, the sweetness is 30% of sucrose, not digested by intestinal digestive enzymes, and can be used by bifidobacteria selectively in the large intestine, which can play the health care role of bifidobacterial factor. Para gold sugar in the high temperature and high pressure, with Rainier nickel as a catalyst for oxidation will generate Para gold sugar alcohol. This sugar alcohol sweetness of sucrose 45 ~ 60%, the calorific value of one-half of sucrose. After eating is not easy to digest and absorb, will not cause blood sugar and insulin elevation, will not cause tooth decay, suitable for diabetics, the elderly, the obese as a sweetener. Because of its physical properties similar to sucrose, it can be used to make low calorie candies, which is a new generation of sweetener popular in the world. The above three kinds of sugar in Europe and the United States, Japan and so on has been mass production, and is widely used; and in the domestic research has been successful, but in the production and application of a lot of resistance still exists.
(4) Oligofructose: Sucrose as raw material by the role of Aspergillus niger β2 fructosyltransferase, the sucrose molecules of D2 fructose to β22,1 chain connecting 123 fructose molecules from sucrose trisaccharides, sucrose tetrasaccharides, as well as sucrose fructofructose, sucrose, glucose, and fructose mixtures, sweetness of sucrose for 60%. After removing glucose and fructose with ion exchange resin, the product containing more than 95% of oligofructose can be obtained, and the sweetness is 30% of sucrose. Sucrose trisaccharides and sucrose tetrasaccharides, which are the main components of oligofructose, are not hydrolyzed by α2 glucosidase in saliva, digestive tract, liver and kidneys in the human body. They are dietary fibers, which can reach the large intestine directly after eating, and can be utilized by beneficial bacteria in the large intestine preferentially. Oligofructose does not cause the increase of blood sugar and insulin level, and its calorific value is 1.5kCal/g. Through the proliferation of bifidobacteria, the intestinal tract can be purified, the immunity of the body can be strengthened, the nutrition can be improved, and the blood lipids can be reduced. In the test with the age of 50-90 years old people, the daily diet of oligofructose 8g, 8 days after the intestinal bifidobacteria can be increased from 5% to 25%. Constipated people consume 5-6g of oligofructose per day, after 4 days, 80% of constipated people's symptoms improve, stool becomes soft, color turns yellow, odor decreases, and intestinal corruption is controlled.
Oligofructose also exists in inulin, chicory, asparagus and other plants, Western Europe are using inulin as raw material, with inulin enzyme local hydrolysis and become. The Japanese government will oligofructose approved as a specific health food; Western Europe, Finland, Singapore, Taiwan and other places will oligofructose as a functional food ingredients, widely used in a variety of foods. The annual production capacity of oligofructose in mainland China is 15,000 tons, including 10,000 tons of Quantum High-Tech in Jiangmen, Guangdong, 3,000 tons of Tianyuan in Yunnan, 1,000 tons of Liangfeng in Zhangjiagang, and 500 tons of Aoligao in Guangxi University. In addition, Wuliangye Brewing Company and Shanghai Zhongke Biomedical Technology Development Co.
(5) Oligosaccharide is characterized by strong acid and heat stability, so it can be used in fruit juice and other acidic beverages, because it is not utilized by most of the intestinal bacteria, only a few bacteria such as bifidobacteria can be utilized, so it is a strong bifidogenic factor, and it can be effective if you ingest 0.7g of it every day. This sugar is produced by extracting xylan from corn kernels and hydrolyzing it with Aspergillus xylanase. It was firstly produced by Suntory Company in Japan and successfully developed by Shandong Longli Company in China with the support of China Agricultural University. Shandong Food Fermentation Research Institute has also announced the successful development. In addition, other functional oligosaccharides such as oligogalactose, oligomannose, etc. have also been developed successfully.
2.2 Enzymes for the production of functional peptides
In recent years, it has been found that the peptides produced by protease hydrolysis of proteins are better absorbed than proteins or amino acids composed of proteins, and can be used as infusion, food for athletes, and health food, etc. Among the protein hydrolysates, the peptides produced by protease hydrolysis of proteins have been successfully developed in China. In protein hydrolysates, some peptides have physiological activity, such as casein hydrolyzed by trypsin or alkaline protease can generate casein phosphopeptide (CPP), which has the function of promoting Ca and Fe absorption. The hydrolysate obtained by enzymatic hydrolysis of fish, soybean and casein contains a heptapeptide with the sequence Ala - Val - Pro - Tyr - Pro - Gln - Arg, which is a kind of Angiotensin Converting Enzyme Inhibitor (ACEI, An2giotensin Converting Enzyme Inhibitor). It can combine with angiotensin to affect the expression of its activity, thus preventing the increase of blood pressure, and is an ideal antihypertensive food. Some of the peptides, which are hydrolyzed by different protein raw materials and different proteolytic enzymes, have the physiological functions of lowering blood lipids, promoting alcohol metabolism, anti-fatigue, and anti-allergy. Eating soybean paste, tempeh, natto, emulsions and other brewed food is good for health, and the reason is also here. Peptones are raw materials for bacterial culture media, because they are found to have physiological functions, even
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2.3 Enzymes in the oil industry
The application of enzymes in the oil industry is still in its infancy. (1) cellulase, hemicellulase used in the oil industry: oil with solvent extraction oil, residual solvents in the residue is difficult to completely remove, affecting the application of feed, so Japan developed the use of cellulase, hemicellulase and pectinase decomposition of plant tissues, to extract oils and fats. The method is to crush or heat-treat olive and rapeseed, then add hemicellulase for a few hours and centrifugally separate the oil from the residue. This process has been used in olive oil, orange oil extraction, rapeseed oil has entered the pilot stage. In the production of animal fats and oils, the use of protease treatment, so that the proteins and fats separated from the high temperature treatment can be avoided, the quality of oil and grease is also better. In order to remove the residual lecithin from the oil, phosphatase is used to remove the water-soluble lecithin from the oil.
(2) Manufacture of fatty acids
Lipase has position specificity and non-specificity to the substrate, and also has selectivity to the length and unsaturation of the fatty acid chain of the substrate. Lipase, which has no position specificity, hydrolyzes lard to produce fatty acids, which are used as raw materials for making soap. The lipase that has no effect on unsaturated fatty acid esters is used to hydrolyze fish oil, but it is difficult to hydrolyze the triglyceride of the highly unsaturated fatty acid DHA and retains the triglyceride, so it is used to manufacture ω3 fatty acids such as DHA.
(3) Ester exchange
The use of lipase ester exchange to change the fatty acid composition of fats and oils can change the properties of fats and oils, such as palm oil modified to become cocoa butter.
2.4 Transglutaminase (TGASE) for meat processingTransglutaminase catalyzes the transacylation reaction between γ2 amide group and various primary amines on glutamic acid residues in protein molecules. When ε2 amino group of lysine residue of protein acts as acyl acceptor, it can form ε2(γ2Gln) Lys *** valence bond between the molecules and cross-linking, which increases the gel strength of protein. It can improve the structural and functional properties of proteins, and by utilizing this effect, it can reorganize low-value minced meat, improve the appearance and texture of fish and meat products, and reduce wastage, thus increasing the economic value. It can also improve the nutritional value by introducing essential amino acids such as Met, Lys, etc. into proteins lacking these amino acids. This enzyme can also be used in woolen fabric processing, immobilization of enzymes, linking of different molecules, linking of antibodies to pharmaceuticals, and so on. The production of strains of Streptomyces reptoverticillummobaracens (S t reptoverticill ummobaracens) has been commercially produced in Japan, and China's Wuxi University of Light Industry has been successfully researched and transferred to the trial production stage.
2.5 New uses of enzymes in fruit and vegetable processing
(1) Protopectinase is used for pectin extraction:
Pectin in fruits is in the form of insoluble protopectin before ripening, and it is gradually transformed into soluble pectin during ripening of fruits. Proto-pectin can also be transformed into soluble pectin under the effect of acid and heat. Protopectinases produced by Bacillus subtilis, Aspergillus niger, yeasts, and tambourines have been developed for the extraction of pectin from orange peel, apple, grape peel, and carrot. Enzymatic extraction of pectin with acid-heat method compared to the process is simple, non-polluting, low cost, product quality in addition to slightly higher sugar content, there is no difference.
(2) Macerating enzymes (Macerating enzymes) used to improve the juice yield:
Macerating enzymes are a mixture of pectinase, hemicellulase (including xylanase, arabinogalactanase, mannanase), cellulase, and act in the collapse of the fruit to promote filtration and improve the juice yield than a single pectinase is better. Good. It has been the main enzyme in juice processing.
(3) Vacuum or pressurized osmosis enzyme treatment of intact fruits and vegetables:
The use of pressurized or vacuum impregnation of fruits and vegetables, so that the pectinase enzyme penetrates into the cell interstitial space or the cell wall to play a role. This method has been used for the softening of whole oranges, the peel can be easily removed. It has also been used to harden peaches by infiltrating the enzyme pectin methyl esterase and Ca2+ into the flesh of the peaches to increase the hardness of canned candied peaches by a factor of 4 (because the pectin of the demethyl ester binds to the Ca2+ and enhances the hardness). This process prevents softening and maintains the crispness of pickled vegetables. This method is also used for naringinase de-bittering of orange peels, with a de-bittering rate of 81 %.
(4) Seven enzymes are used to remove phenolic compounds
Clarified fruit juice is filtered by ultrafiltration, and after concentration, white turbidity still occurs, which is caused by phenolic compounds in the juice, so before filtration, it can be used to treat with seven enzymes, so that it oxidizes and polymerizes into an insoluble molecule, and then filtration is performed to remove it.
(5) Pectinase is used to clean the membrane of pectin contaminants.
(6) β2 Glucanase is used to remove β-glucan from grape musts infected with Cotrytis cinerea, and Vinozyme is used to precipitate insoluble matter.
2. 6 Application of enzymes in the textile industry
Cotton fabrics have been desizing with amylase for more than 100 years, and with the development of the enzyme industry, enzymes such as cellulase, pectinase, xylanase, gaseasease, protease, etc., have been successively adopted by the textile industry.
(1) Cotton finishing enzyme
With the popularity of denim, cellulase finishing cotton to improve the textile look and feel, has been widely valued by the textile industry. Cellulase enzyme action in the natural fiber non-crystalline area, so that part of the fiber degradation and modification, can make the fabric soft, smooth, feel and look comfortable. Usually after treatment with enzymes, the weight of cotton fabrics is reduced by 3-5%, but the fastness is lost by about 20%. In developed countries for the pursuit of fashion, do not care about the fastness of the cloth.
Hydrogen peroxide enzyme is often used to remove the residual H2O2 after bleaching with H2O2, and recently it is found that Arthromyces ramosus and Coprinus cinereus can produce large amount of hydrogen peroxide enzyme, and hydrogen peroxide enzyme is also used in detergents. Pectinase is used in cotton finishing, mainly to decompose the pectin on the surface of cotton and linen fabrics, so as to facilitate bleaching and dyeing. Seven enzyme is a kind of phenol oxidase, with O as the H receptor, mainly used in denim indigo dyeing decolorization treatment, NOVO company using gene technology to improve the production of Aspergillus niger. The enzyme can also act on lignin and decompose lignin. Xylanase is used in the bleaching treatment of cloth blanks to remove lignin and cotton husk adhering to the fiber.
(2) Woolen fabric protease anti-felt shrinkage finishing
Woolen fabrics without finishing after washing will be shrinkage felting can not be worn again (such as poor quality wool sweater shrinkage after washing is very small), must be anti-shrinkage and anti-felting treatment to maintain the original state after washing. Anti-felting antiseptic treatment has a history of more than 100 years, in the past with chlorine, H2O2, persulfate treatment, serious pollution, the 90's developed a non-chlorine shrinkage inhibitor. The use of protease to change the structure of wool can be used to prevent felting and shrinkage treatment in the 40's, it was studied in the 60's Japan reported that the papain treatment can prevent felting and shrinkage, and can be used for low-temperature dyeing to improve the dyeing rate, reduce the sewage and improve the feel and feel of woolen fabrics. In the 70's, we have also tried to use acid protease treatment to carry out low-temperature dyeing, and good results were achieved, with an increase in dyeing rate of 3.6%, and a reduction of 62% of sewage. The dyeing rate was increased by 3.6 % and the waste water was reduced by 62 %. The yarn breakage rate per thousand spindles was reduced to 145, and the elongation resistance, tensile strength and hand feeling were improved obviously. Since the 80's, the enzyme method of anti-felt shrinkage has attracted renewed attention both at home and abroad, and a large number of research articles have been published in Japan, the United Kingdom, the United States and other countries and some progress has been made. The researched proteases are trypsin, papain, alkaline protease, neutral protease, acid protease, etc. It is believed that these processes will soon be mature and popularized.
2. 7 Application of enzymes in the paper industry
The paper industry is an important source of environmental pollution. As people's awareness of environmental protection increases, the use of biotechnology in the paper industry has been emphasized. Enzymatic pulp production has aroused strong interest in various countries, the key is to degrade lignin. Recently, some people in China have utilized a variety of microorganisms to produce pulp, and have made promising progress, and are now preparing to expand the test. Enzyme applications in the paper industry is now mainly lipase used for log de-resin, cellulase hemicellulase and lipase used in the recycling of waste newspaper de-inking; and xylanase used in pulp bleaching.
(1) log de-resinning:
The logs used for papermaking contain resin, which pollutes the equipment and affects the production and reduces the quality of the paper when it is pulped and copied. For this reason, it needs to be stacked outdoors for a long time (more than 3 months), so that the resin decomposition. This affects the production cycle and takes up a lot of space. The Japan Paper Research Organization (JPRO) studied the composition of logs and found that 96% of the resin is oleic and linoleic acid, which can be removed by lipase treatment. Since its introduction in the 1990s, the quality of paper has improved, log stacking costs have been reduced, the amount of resin adsorbent used has been reduced, and economic benefits have been realized. At that time, the lipase used was supplied by NOVO, and it worked well at pH 6-10, 40-60 ℃. Recently, it has been found that the lipase with a heat resistance of 70 ℃ is more effective.
(2) pulp bleaching:
Pulp in order to remove the pigment source of lignin, chlorine, hypochlorite, chlorine dioxide and other chlorine treatment, pollution is serious, so in the 60's it was considered to decompose the lignin enzyme. Lignin is a polymer with phenylpropane as its backbone, and only when it is broken down will the lignin disintegrate. Lignin peroxidase (LIP), manganese-dependent peroxidase (MNP), and laccase (LAC) have been identified as enzymes that have the ability to break down lignin, but no suitable ligninase has been found so far. In recent years, Finland has proposed a combination of chemical and enzymatic treatment method, and achieved better results. First use xylanase to cut off lignin and cellulose with cellulose between the linkage between the material (xylan and hemicellulose), so that lignin free, and then alkali cooking, from the pulp free of xylan can be adsorbed again on the surface of the fiber, with xylanase to decompose, can increase the pore space, so that chlorine permeability increased, and make lignin easy to come out of the pulp interior, the process of active chlorine dosage can be reduced by 30%.
(3) Deinking in the recycling of waste newspaper
When pulp is made after recycling of waste paper, alkali, nonionic surfactant, sodium silicate and H2O2 are used for deinking. Japan in the deinking of alkaline cellulase, hemicellulase 0.1 % reaction for 2 hours, the whiteness of the paper can be increased by 4 ~ 5 %, the strength did not decrease. Due to prevent ink prints dirty hands, ink with linoleic acid, linolenic acid and oleic acid and other high-level triglycerides, so deinking and then add lipase effect is better, whiteness can be increased by 2.5 %. Waste newspaper deinking, Shandong University in China has also carried out a lot of research.
2. 8 Other
Phytase is used as feed additive to improve the utilization of organic phosphorus in feed, reduce the phosphorus in the feces pollution of the environment, save the feed plus the amount of phosphate. In recent years, phytase has also been used in brewing to improve the utilization of phosphorus in raw materials, as well as in the production of de-potassiumated soy protein food, which becomes a source of protein for renal patients. α-Glucosyltransferase is also used in the processing of stevia, which is used to remove the bitterness and astringency. Liquefaction and saccharification of starch account for almost the majority of industrial enzyme reactions, because the current enzyme liquefaction, saccharification to be carried out at different pH and temperature, in order to simplify the process, saving water and energy, it is necessary to develop acid-resistant high-temperature α2 amylase and heat-resistant saccharification enzyme, if the α2 amylase can be liquefied in the pH 4.5, and saccharification enzyme can be carried out in the temperature of more than 60 ℃, imagine how much these will bring the benefits? How much benefit will these bring? In addition, liquefaction at pH 4.5 prevents the formation of maltodextrins. Acid-resistant α2 amylase and heat-resistant saccharase have been studied for many years in foreign countries, there have been many reports. For example, Japan reported that a strain of acid-resistant α2 amylase (KOD-1) has been selected and reacted in 30% starch slurry at pH 4.5 for 10 minutes at 105 ℃, and the residual enzyme activity was 75%. The enzyme was liquefied in 30% starch slurry at pH 4.5 at 60 ℃ for 60 min to obtain DE14 liquefied liquid, which was saccharified by adding 0.1 % saccharase for 48 h. The glucose content reached 95.5 %, which was the same as that of the control Bacillus subtilis α2 amylase liquefied at pH 5.8 (glucose content of 95.7 %). In addition, the acid resistance of B. licheniformis α2 amylase was enhanced by replacing seven methionines in the molecule with other amino acids using protein engineering. Once the industrialization of this kind of enzyme is successful, it will greatly change the face of saccharification-related industries.
3 Conclusion
As the world's energy is decreasing and the population is increasing, water and food are becoming increasingly scarce. The increased awareness of environmental protection has made it more urgent for the industry to reform the traditional processes with enzymes. Therefore, it is imperative to increase enzyme production, reduce production costs and develop new varieties of enzymes for new applications. The development of genetic engineering and protein engineering has created favorable conditions for the development of enzyme industry. The development of heat-resistant, acid and alkali resistant enzymes with special effects on the substrate, as well as plant and animal production of enzymes by microbial fermentation method to produce, or will not be able to use the microbial enzymes produced by safe strains of bacteria to produce, will become a reality.