CN1052632C - Slow-calcifying compound epoxy cross-linking method for collagen tissue material - Google Patents

Abstract

The method for slowing calcification of collagen artificial valve materials is to treat natural collagen tissues such as bovine pericardium, porcine aortic valve, large animal aortic blood vessels, etc. with compound epoxy cross-linking agent, so that the strength of such collagen tissues, stress relaxation, biological Compatibility, slow calcification and resistance to enzymolysis are all better than glutaraldehyde-treated collagen tissue products. Among them, the biological valves sewn with composite epoxy collagen tissue bovine pericardium and porcine aortic valve materials are implanted in the human body. The bioprosthetic valve has the obvious function of slowing calcification, and the service life is extended from 7-8 years to more than 15 years. After the valve is replaced, the patient can restore health and prolong life.

Description

Method for Slowing Calcification of Collagen Tissue Artificial Valve Material

The invention belongs to a method for delaying calcification of collagen tissue artificial valves. After the natural collagen tissue is treated with biochemical methods, the diseased heart valves of human body can be surgically replaced, so that patients can be exempted from suffering.

There are three types of artificial heart valves: biological valves, mechanical valves and plastic valves. A biological valve has a central blood flow channel similar to a natural heart valve. It meets the physiological requirements of patients, does not need to take anticoagulant drugs for a long time, and is beneficial to patients with contraindications to anticoagulants; the failure process of heart valve function occurs gradually, which is conducive to treatment or re-valve replacement; it will not cause discomfort to patients The noise of the feeling; the source of raw materials is abundant, easy to make, low cost and other advantages. Therefore, the research and development of artificial heart valves are very fast.

In the mid-1960s, the replacement of diseased heart valves was first developed from the replacement of blood vessels. Ross.D.J., J.Thorac Cardiovasc.Surg 47, 713 (1964) selected biological valve materials including the same kind of heart valve or heterogeneous aortic valve treated with formaldehyde. After several years of research and clinical practice, the biological valve cannot be improved. The function and protective effect on collagen are eliminated.

Since the early 1970s, the second-generation collagen cross-linking agent glutaraldehyde has dominated. As early as 1968, A.Carpentier used glutaraldehyde instead of formaldehyde to treat porcine aortic valve for the first time. Glutaraldehyde can form a relatively stable cross-linking bond between collagen molecular bonds, so the bioprosthesis of the biological valve treated with glutaraldehyde Its physical and mechanical functions are far superior to formaldehyde, and it is also very convenient to use. However, in the long-term clinical investigation of more than 20 years, it was gradually found that glutaraldehyde has the following disadvantages: (1) glutaraldehyde reacts with the ε- _NH of collagen to form a pyridinium bond or only one aldehyde interacts with collagen, and the remaining The free aldehyde groups and pyridinium bonds in the bioprosthesis can cause endogenous collagen tissue calcification; (2) when the bioprosthesis material is fixed, the pentanediol polymer accumulated in the tissue gradually decomposes in the body as the bioprosthesis is implanted into the human body. Polymerization, the moldyness of the released glutaraldehyde monomers, inactivates blood cells and humoral cells that enter the leaflet material, thus causing exogenous collagen tissue calcification. Regardless of endogenous or exogenous calcification, it seriously affects the service life of biological valves. (3) Glutaraldehyde cannot improve the primary failure of biological valves under the action of force; (4) inhibits the participation of collagen calcification Alkaline phosphatase (AP) activity of alkaline phosphatase (AP) is not as strong as Al ⁺⁺⁺ ion; (5) The hydrophilicity, biocompatibility and elasticity of glutaraldehyde-treated bioprosthetic materials need to be improved, and in addition, during the storage period The yellowing of color and luster fully proves that glutaraldehyde is not an excellent collagen cross-linking agent.

The Achilles' heel that affects the development of biological valves is the calcification and tearing of the valve leaflets of biomaterials, which leads to shortened service life of the valve. There are many reasons for calcification and tearing, which involve various biochemical treatments on the material structure, mechanical behavior and other factors. Improving the strength of biomaterials depends on the type and degree of crosslinking agent used to immobilize natural biomaterials. Since the cross-linking reaction of collagen should be controlled under extremely mild conditions, most of the cross-linking agents that can be used in the field of polymers cannot be used, and only protein cross-linking agents have the possibility of selection. Since 1983, Chinese scholars Le Yilun and Wan Changxiu have carried out comprehensive basic research on the histology, biochemistry, immunology, biomechanics, physical chemistry and chemical modification of collagen of yak pericardium in Aba, Sichuan. At the same time, a series of researches have been carried out on the fixation technology, slit flap technology and preservation methods of the pericardium, and the yak pericardial biological valve has been successfully used clinically in cooperation with West China University of Medical Sciences. "YPV-1 Yak Pericardial Valve Appraisal Materials" one to twelve (1986), fully affirmed that the yak pericardial material is a promising biological valve material with great development prospects.

In order to further improve the fatigue resistance of the pericardial material and prevent the occurrence of calcification, the inventors also studied the feasibility of hydrophilic diepoxide as a cross-linking agent for the yak pericardial material. Under room temperature and neutral conditions, the self-polymer with a low degree of polymerization formed by the diepoxide can react with ε-NH ₂ between collagen molecular bonds to form collagen microfibrils and bridges, thereby improving the mechanical properties of collagen fibers. The hydrophilic epoxy crosslinking agent can also combine with other active groups on the collagen molecular chain such as , Guanidyl and methylthio and other reactions. Therefore, there are more opportunities to form bridges between collagen microfibrils than glutaraldehyde. The product of low polymerization degree epoxy crosslinking agent after polymer degradation is a non-toxic or very little toxic alcohol product. The analysis of mechanical properties such as stress-strain, elastic modulus and stress relaxation behavior of the material proves that the yak pericardium material treated with epoxy composite cross-linking has better mechanical behavior and better affinity than the traditional glutaraldehyde-fixed pericardium material. Water-based and biocompatible, and significantly better than glutaraldehyde materials in slowing down calcification.

The purpose of the present invention is to provide a kind of slow calcification method to collagen tissue artificial valve material in view of the deficiencies in the prior art, and its characteristic is that under normal temperature condition, adopts hydrophilic double epoxy composite cross-linking agent to biological tissue material, For example, biochemical treatment of yak pericardium, etc., to obtain artificial biological valves or sewn into other organs, after implantation in the human body, there is an obvious effect of slowing calcification, and the service life is extended from 7-8 years to more than 15 years. The strength and stress are relaxed, and the biological Compatibility, slow calcification and resistance to enzymolysis, etc., are superior to collagen tissue materials treated with glutaraldehyde, which is beneficial to the recovery of patients.

The method for slowing calcification of collagen tissue artificial valve materials is that the natural collagen tissue (bovine pericardium, porcine aortic valve, dura mater, fascia lata, large animal arteries, hoof tendon and sheep peritoneum, etc.) is biochemically treated. It is used clinically or sewn into organs to be implanted in the human body.

1. Rough selection: Under the condition that the environment is clean and the equipment and liquid medicine in contact with collagen tissue are sterilized and sterilized according to surgical requirements, the collected fresh collagen tissue is immediately removed at room temperature within half an hour to remove the fat and reticular tissue on its surface. connective tissue.

2. Rinse: wash the roughly selected fresh collagen tissue with sterilized physiological saline (0.9% NaCl) solution for several times until no blood seeps out. Each solid-liquid ratio is 1:10-50, that is, the ratio of weight to volume.

3. Soaking and extracting soluble protein: the HanKs solution without calcium salt is used for extraction, the solid-to-liquid ratio is the same as in (2), and washed until no soluble protein is leached out by ninhydrin reagent test.

4. Selection: Select a uniform part of the collagen tissue and cut it into a rectangular piece of a certain size.

5. Antibiotic sterilization: add 40,000 to 80,000 units of gentamicin per 500 ml of normal saline, and the solid-liquid ratio is the same as before (2).

6. Cross-linking agent fixation: 1-15% multifunctional epoxy compound is used for cross-linking agent n=1～10, with the ^pH 9-12 of the carbonate buffer solution of 0.1N) and 5-50% (volume/volume) water-soluble organic solvent (available ethanol, acetone, dioxane) as Cosolvent, prevent collagen swelling, accelerate the reaction process, ^pH 9-10, crosslink and fix collagen tissue, react at room temperature for more than 96 hours.

7. Store in preservation solution for later use: sterilize and preserve in 2% formaldehyde phosphate buffer solution and 0.5-3% water-soluble surfactant (available polyethylene glycol, polypropylene glycol and sodium lauryl sulfate). The collagen tissue material prepared by the above method can also be used as heart patch, blood vessel or other medical repair materials besides being used for sewing biological heart valves.

8. Fixation of bioprosthetic valve: ZrOCl ₂ , Al ₂ (SO ₄ ) ₃ , AlCl ₃ , FeCl ₃ , Cr ₂ (SO ₄ ) ₃ , CrCl _{3 ,} etc. The above mixtures are placed in physiological saline solution to make the sewn bioprosthesis rapidly fixed and molded into slow calcification composite epoxy cross-linked products, and then put into 2% formaldehyde phosphate buffer and surfactant for sterilization and preservation.

Table 1 below compares the effects of the yak pericardium treated with the slow calcification compound epoxy crosslinking method of the collagen tissue material of the present invention and the yak pericardium treated with the traditional glutaraldehyde method. Practice has proved that the present invention overcomes a series of disadvantages and shortcomings brought about by the traditional glutaraldehyde treatment of collagen tissue.

Performance test of yak pericardium treated by compound epoxy method and glutaraldehyde method:

Table 1 Effects of two cross-linking agents on yak pericardium project Treatment of Yak Pericardium with Compound Epoxy Crosslinking Agent Treatment of Yak Pericardium with Glutaraldehyde Color Shrinkage Temperature (°C) Breaking Strain (%) Constant Load Tensile Stress Relaxation Test (Residual Stress When Stretched for 500 Seconds) Softness and Elasticity Biocompatibility Antigenic Immunity Resistant to Enzyme Hydrolysis Hemolysis Test Cytotoxicity Acute Toxicity Subcutaneous stimulation pyrogen test Subcutaneous implantation test Antigenic immune response Determination of calcium content μg/mg dry tissue White or green 82～90°C 70±10.687.6±1.8 soft, thick and elastic animal tissue compatibility good very slightly excellent less than 5% 96 grade non-toxic negative negative negative negative excellent very small 0.57±0.19 Light yellow 8549±5.886.5±2.8 Rough feeling, thin and stiff cells are not easy to grow Very slightly good Less than 5% Grade 96 non-toxic Negative negative Negative better Minimal 23.2±4.6

1. Amino acid analysis

The samples processed by the composite epoxy method were analyzed for amino acids by the central laboratory of the Sichuan Academy of Agricultural Sciences, and the samples processed by the glutaraldehyde method were used for comparison, and were determined by Hitachi 835-50 high-speed amino acid analyzer.

Table 2 Amino acid content after two kinds of cross-linking agents treated yak bag amino acid residue Number of amino acid residues Number of responses new style composite epoxy glutaraldehyde composite epoxy Glutaraldehyde Arg (Arginine) Lys (Lysine) His (Histidine) Thr (Phenyl) Met (Methionine) Ser (Serine) Tyr (Tyrosine) Glu (Glu) Asp (Tianmen aspartic acid) 350198511622925070576382 35023511621325029576382 35267511622925270576382 -175--16-41** -131-----**

Amino acid residues per 1000 total amino acids

The ester bond formed by epoxy and acidic amino acid is hydrolytically broken under acidic hydrolysis conditions.

Amino acid analysis showed that, compared with glutaraldehyde-treated materials, the materials treated with epoxy cross-linking introduced a variety of cross-linking bonds, and the number of cross-linking points also increased.

2. Mechanical performance test

The comparison test of composite epoxy cross-linked materials and glutaraldehyde-treated materials by Shimadzu-AG-5000A biological material testing machine is shown in Figure 1 and Figure 2 by the Drug Control Institute of the Ministry of Health.

Figure 1. Stress-strain curves of yak pericardial material ^*

1. Fresh sample 2. Compound epoxy method, 3. Glutaraldehyde method

*Circumferential orientation of samples, number of samples n=9, each point on the curve is mean ± standard error.

Figure 2. Stress relaxation curve of yak pericardial material ^*

1. Fresh sample, 2. Compound epoxy method, 3. Glutaraldehyde method

* The time scale is log(t), the number of samples is n=10, the points on the curve are mean ± standard error, and the mechanical properties of the composite epoxy-treated yak pericardium material are better than those of the glutaraldehyde-treated yak under the load of 550 seconds pericardial material.

Table 3 Tensile strength and elongation at break of yak pericardium treated with two cross-linking agents sample Number of samples (n) Tensile strength KN·m ^-2 ·10 ³ Elongation at break% New compound epoxy glutaraldehyde 71410 12.5±4.412.3±3.85.0±1.1 6.1±6.570±10.649±5.8

Because epoxy compounds can react with various amino acid side groups in collagen molecules, such as ε-NH ₂ , -COOH, -OH, etc., to form cross-linking bonds with a relatively uniform distribution. In addition, epoxy crosslinkers are flexible molecular chains with ether linkages. Therefore, on the basis of maintaining the tensile strength of the material, the elongation at break of the epoxy treatment is also significantly improved, which is very beneficial for the leaflet material subjected to variable stress in the body. The stress relaxation of collagen tissue mainly comes from the slippage between fiber bundles and the viscoelasticity of fiber molecules themselves. After chemical modification, the stress relaxation rate of the materials decreased, but the decrease rate was smaller for EP pericardial material. Again, it shows that the stress relaxation effect of the modified pericardial material is consistent with the amino acid analysis results of the material, that is, the number of cross-links introduced by epoxy between collagen molecules is more than that of glutaraldehyde.

The yak pericardium material treated with compound epoxy cross-linking agent is more flexible than glutaraldehyde-treated material, its tensile strength and elongation at break are increased, and its mechanical properties are better than those treated with glutaraldehyde.

3. Calcification experiment of materials

Among the causes of bioprosthetic valve failure, non-toxic degeneration (mainly tissue degeneration) accounts for 74% of the total failure reasons, and tissue degeneration is mostly related to valve leaflet calcification. research slow

Calcification material is the key problem to solve the service life of biological valve.

According to the method of Fishbein and Levy, the samples and controls were embedded subcutaneously in the back of rats, taken out after 21 days, dried to constant weight, nitrated, and the calcium content was determined by PE3030 atomic absorption spectrophotometer.

Table 4 Calcification experiment of yak pericardium treated with two cross-linking agents sample Measurement times Ca ⁺⁺ μg/mg dry tissue Glutaraldehyde composite epoxy 99 23.2±4.60.57±0.19 Compared with the glutaraldehyde sample, the calcium content of composite epoxy treated samples decreased significantly, and the composite epoxy treated materials had obvious calcification slowing effect.

4. Enzyme hydrolysis experiment of materials:

The enzymatic hydrolysis method is expressed by the amount of hydroxyproline (Hypro) released by the collagen tissue material under the action of enzyme (type I collagenase Collagenase I, or pronase Pronase). The lower the value of hydroxyproline, the more The stronger the resistance to enzymatic hydrolysis.

Table 5 Enzyme hydrolysis experiment of yak pericardium treated with two cross-linking agents pericardial material Yak pericardium Cattle pericardium Fresh epoxy Zirconium composite epoxy Chrome Composite Epoxy glutaraldehyde Glutaraldehyde (Brazil) Glutaraldehyde (USA) Shrinkage temperature Ts℃ Hydroxyproline content Hypro value mg/g dry sample 68116.0 7952 8442.2 8350.1 8860.3 8858.8 8860.5

Experiments have shown that the yak pericardium material treated by composite epoxy crosslinking method has significantly lower hydroxyproline value than that of yak pericardium material treated by glutaraldehyde method and yellow cattle pericardium materials from Brazil and the United States under the condition of low shrinkage temperature Ts℃. Product samples.

The present invention has the following advantages:

After biochemically treated collagen tissue materials are implanted into the human body, they have good biocompatibility and little rejection, and are increasingly widely used in medical surgery. Among them, artificial biological heart valves are the most used, and only domestic and foreign valve replacements are the most important. For example, more than 1 million valve replacements have been performed in the world, about 10,000 valve replacements are performed in the United States every year, and more than 5,000 valve replacements have been performed in the Federal Republic of Germany in 1988, and the sales volume in the international market has increased by about 6.9% per year. There are about 1.4 million adults with rheumatic heart disease in my country, 80,000 of whom are in urgent need of valve replacement. According to the above growth rate, there will be an additional 5,000 people in urgent need of valve replacement every year in my country. At present, more than 100 hospitals in China have carried out valve replacement surgery, but the output is far from meeting the clinical needs. According to the survey of West China Medical University in the early 1980s, 0.8% of the 100 million people in Sichuan suffered from heart valve disease, that is, 800,000 patients, of which 10 % should be replaced, plus 10% of those who are willing or likely to be replaced, there are more than 10,000 people in this province who are urgently waiting for valve replacement. Therefore, the present invention is more beneficial to patient because:

1. Reduce or delay the calcification of collagen tissue materials in the body, thereby greatly prolonging the service life of the biological valve implanted in the human body, from 7-8 years to more than 15 years, which is conducive to the recovery of patients.

2. Compared with artificial mechanical valves, biological valves are cheaper to manufacture, and their hemodynamic functions are close to the physiological functions of the human body. They have good blood compatibility and do not need to take anticoagulant drugs. They are suitable for the national conditions of our country and developing countries. The implementation of this patent will Bring the gospel to human heart valve disease patients.

3. my country has abundant resources of biological tissue materials, especially tens of millions of pigs and tens of thousands of yaks are slaughtered every year in Sichuan Province, and there are inexhaustible and inexhaustible pig aortic valves of complete models and high-quality The advanced yak pericardial material provides a reliable guarantee for the development of the biological valve industry. Natural collagen tissue materials, before cross-linking treatment, are generally waste products from slaughtering and processing factories. After biochemical treatment, they can turn waste into treasures for the benefit of mankind. It can not only save lives, but also earn foreign exchange. For example, Guangzhou Pacific Company in China costs about 100 yuan per pericardium, and 1,500 yuan for each bioprosthesis, and it is made of traditional glutaraldehyde-treated materials. The cost of bioprosthesis imported from the U.S. is $1,000 each. If the materials produced by the method of the present invention are used to manufacture slow-calcification bioprosthetic valves, the quality is better than that of the U.S. If 5,000 pieces are produced annually and sold at the same price, the annual output value can reach 7.5 million yuan, and the export can earn 5,000 US dollars.

Example 1:

1. Rough selection: select healthy calves aged 3-5, and collect them within half an hour after slaughter. The environment is required to be clean, and the medical solution of the equipment that contacts the pericardium is sterilized according to surgery. The collected fresh yak pericardium is kept at room temperature, and the optimal temperature is 4-10°C to remove superficial fat and rough connective tissue.

2. Rinsing: Wash the rough-selected fresh yak pericardium several times with sterilized physiological saline (0.9% NaCl) solution until no blood seeps out. The best temperature is 4-10°C.

3. Soaking and extracting soluble protein: Sterilize with HanKs solution without calcium salt, the solid-liquid ratio is the same as in (2), and wash until no soluble protein is leached out in the ninhydrin test.

4. Selection: Select a uniform part of the precordial area and cut it into a rectangular piece with a size of 120×100 mm.

5. Antibiotic sterilization: Add 40,000-80,000 units of gentamycin to 500 milliliters of normal saline, the solid-to-liquid ratio is the same as in (2), and the temperature is room temperature, preferably 4-10°C.

6. Cross-linking agent fixation: the cross-linking agent can be 2-3 functional epoxy compound The biological material is cross-linked and fixed with 0.1N carbonate buffer solution and 20% ethanol as co-solvent (p ^H 9-10), the reaction temperature is 4-10° C., and the time is more than 96 hours.

7. Store in preservation solution for later use: sterilize and preserve in 2% formaldehyde phosphate buffer and 1% polyethylene glycol for about 1 month. The slow calcification collagen tissue material thus prepared can be used as a heart patch, blood vessel or other medical repair materials in addition to sewing biological valves.

8. Fixation of biological valve: use 0.2% ZrOCl ₂ in physiological saline solution to quickly fix and form the sewn biological valve. Then put it into 2% formaldehyde phosphate buffer and surfactant polyethylene glycol for sterilized preservation.

Example 2:

1. Rough selection: select fresh dog arterial blood vessels at room temperature to remove the muscle and requirements on the surface of the blood vessels, and implement (1) of 1.

2. Rinsing: Wash the roughly selected fresh dog arteries several times with sterilized physiological saline (0.9% NaCl) solution until no blood seeps out. room temperature.

4. Soak and extract soluble protein, same as (3) in Example 1.

4. Selection: Select a uniform part and cut a certain length of blood vessel.

5. Antimicrobial sterilization: the same as (5) in Implementation 1.

，n＝3与0.1N的碳酸盐缓冲液和20％丙酮作为助溶剂，p^H9-10，对血管进行交联固定，室温下反应96小时。6. Cross-linking agent fixation: 2% bifunctional epoxy compound can be used as cross-linking agent

, n=3 and 0.1N carbonate buffer solution and 20% acetone as co-solvent, ^pH 9-10, blood vessels were cross-linked and fixed, and reacted at room temperature for 96 hours.

7. Store in preservation solution for later use: sterilize and preserve in 2% formaldehyde phosphate buffer solution and 0.5% polypropylene glycol, then it can be directly used to replace diseased blood vessels.

Example 3:

1. Rough selection: select fresh porcine aortic valve to remove fat and uneven parts at room temperature, and the requirements are the same as in Example 1 (1).

2. Rinsing: 3. Soaking and extracting soluble protein, 4. Selecting and 5 antibiotic sterilization are all the same as (2)(3)(4)(5) in Example 2.

n＝10与0.1N的碳酸盐缓冲液和10％二氧环己烷作为助溶剂，p^H9-10，对该主动脉瓣进行交联固定，室温下反应96小时。6. Cross-linking fixation: the cross-linking agent can be 2% difunctional epoxy compound

n=10 and 0.1N carbonate buffer solution and 10% dioxane as co-solvent, ^pH 9-10, the aortic valve was cross-linked and fixed, and reacted at room temperature for 96 hours.

7. Store in preservation solution for later use: sterilize and preserve in 2% formaldehyde phosphate buffer and 0.5% sodium dodecyl sulfate.

Claims (6)

1. the method for the slow calcification of collagen tissue artificial valve material, is characterized in that: (1) Rough selection: Under the condition that the environment is clean and the equipment and liquid medicine in contact with collagen tissue are sterilized and sterilized according to surgical requirements, the collected fresh collagen tissue is immediately removed at room temperature within half an hour to remove the fat and fat on its surface. reticular connective tissue, (2) Rinse: Wash the roughly selected fresh collagen tissue with sterilized saline solution for several times until no blood seeps out. The solid-liquid ratio is 1:10-50 each time, that is, the ratio of weight to volume, (3) Soaking and extracting soluble protein: HanKs solution without calcium salt is used for extraction, the solid-to-liquid ratio is the same as in (2), and washed until no soluble protein is leached out in the ninhydrin reagent test, (4) Selection: select a uniform part of the collagen tissue and cut it into a rectangular sheet of a certain size. (5) Antibiotic sterilization: Add 40,000 to 80,000 units of gentamicin per 500 ml of normal saline, and the solid-liquid ratio is the same as in (2). (6) Cross-linking agent fixation: the cross-linking agent uses 1 to 15% multifunctional epoxy compound, 0.1N carbonate buffer and 5 to 50% water-soluble organic solvent as cosolvents to carry out collagen tissue The first cross-linking fixation, reaction at room temperature for more than 96 hours, (7) Store in preservation solution for later use: sterilize and preserve in 2% formaldehyde phosphate buffer and 0.5-3% water-soluble surfactant, (8) Fixation of biological valve: use 0.05-5% compound metal salt in physiological saline solution to make the sewn bovine pericardium or porcine aortic valve for the second time to form a slow calcification composite epoxy crosslinking Method products, and then put into 2% formaldehyde phosphate buffer and surfactant for sterilization and preservation. 2. according to the method for slow calcification of collagen tissue artificial valve material according to claim 1, it is characterized in that collagen tissue material is bovine pericardium, porcine aortic valve, human dura mater, fascia lata, large animal arterial vessel, tendon and sheep peritoneum. 3. according to the method for the slow calcification of collagen tissue artificial valve material according to claim 1, it is characterized in that the polyfunctionality epoxy cross-linking agent is n=1-10 homologous compounds. 4. according to the method for slowing calcification of collagen tissue artificial valve material according to claim 1, it is characterized in that the water-soluble organic co-solvent is ethanol, acetone and dioxane. 5. According to the method for slowing calcification of collagen tissue artificial valve material according to claim 1, it is characterized in that the water-soluble surfactant is polyethylene glycol, polypropylene glycol and sodium lauryl sulfate. 6. The method for slowing calcification of collagen artificial valve material according to claim 1, characterized in that the composite metal salt is ZrOCl ₂ , Al ₂ (SO ₄ ) ₃ , AlCl ₂ , FeCl ₃ , Cr ₂ (SO ₄ ) ₃ , CrCl ₃ or a mixture of more than one.

Images