Cacumen biotae extracting solution and preparation method thereof

Abstract

The invention relates to the technical field of biological medicine industry, in particular to a Chinese arborvitae twig and leaf extracting solution and a preparation method thereof, and the Chinese arborvitae twig and leaf extracting solution is prepared from the following raw materials: modified Chinese arborvitae twig and leaf extracting solution, modified panax notoginseng saponins, panax notoginseng, fibrin, chitosan, ferric oxide, glycerol and modified spinach liquid. The modified Chinese arborvitae twig and leaf extracting solution, the modified panax notoginseng saponins and the modified spinach liquid play a role through a synergistic effect, the modified Chinese arborvitae twig and leaf extracting solution contains relatively high volatile oil which can stimulate platelet aggregation and promote release of blood coagulation factors, and the modified panax notoginseng saponins can inhibit excessive activation and aggregation of platelets and prevent thrombosis; the modified cacumen biotae extracting solution and the modified panax notoginseng saponins can jointly promote platelet aggregation and blood coagulation factor release to accelerate blood coagulation, the modified spinach liquid promotes blood coagulation factor synthesis and release by providing vitamin K and accelerates blood coagulation, a dynamic balance system is formed by the modified spinach liquid, the modified cacumen biotae extracting solution and the modified panax notoginseng saponins, and blood coagulation is promoted through the synergistic effect. The hemostatic effect is achieved.

Description

Technical Field

[0001] The present invention relates to the technical field of biomedicine industry, and in particular to a Platycladus orientalis leaf extract and a preparation method thereof. Background Art

[0002] In the biomedical field, blood coagulants play a pivotal role. They are not only essential to ensure the integrity of blood samples during collection, storage and transportation, but also play an irreplaceable role in surgery, treatment of hemorrhagic diseases and research on coagulation mechanisms. The common ingredients of blood coagulants on the market are heparin salts, EDTA potassium salts and sodium citrate, most of which are organic compounds, and long-term use will cause certain side effects to patients, including gastrointestinal reactions, allergic reactions, bleeding tendencies and potential harm to specific organs or systems. In addition to the use of traditional blood coagulants, with the advancement of medical technology, some new anticoagulant drugs and monitoring devices have also emerged. These new drugs and devices have shown great potential in improving anticoagulant effects and reducing side effects, but they also face the problem of high costs. The high cost of treatment may limit the accessibility of treatment for some patients, especially in areas with limited medical resources and low-income people. Therefore, exploring safe, effective, inexpensive and less side-effect alternative anticoagulant drugs is crucial to improving the treatment and prevention of thrombotic diseases. Platycladus orientalis leaves, as a plant of the genus Platycladus in the Cupressaceae family, have attracted much attention due to their rich medicinal active ingredients. The chemical composition of Platycladus orientalis leaves is complex. Studies have shown that they contain multiple types of medicinal active ingredients, among which flavonoids and volatile oils are relatively high. Both traditional Chinese medicine and modern medical research have confirmed that Platycladus orientalis leaves have significant therapeutic effects in the treatment of various diseases. For example, they have multiple pharmacological effects such as antibacterial, anti-tumor, hemostasis, hair growth promotion, and anti-inflammatory. The potential medicinal value of Platycladus orientalis leaves and their auxiliary effects on various diseases in clinical practice are becoming increasingly apparent. Therefore, further in-depth research on the pharmacological mechanism of Platycladus orientalis leaves and the development of new anticoagulants based on Platycladus orientalis leaves will not only help solve the side effects caused by the long-term use of traditional anticoagulants, but may also provide patients with more economical and effective treatment options, which will help promote the development of the biomedical field. Summary of the invention

[0003] (1) Technical issues to be solved

[0004] The purpose of the present invention is to provide a Platycladus orientalis leaf extract and a preparation method thereof, to accurately extract components that have a significant promoting effect on blood coagulation, to optimize the main components of the blood coagulant, to combine the research content of modern medicine, and to extract and use the natural components of Chinese medicinal plants, to not only achieve a dual improvement in the safety and effectiveness of the blood coagulant, but also to reduce the production cost, to provide a more high-quality and efficient blood coagulant solution for the biomedicine field, and finally to obtain a Platycladus orientalis leaf extract.

[0005] (2) Technical solution

[0006] To achieve the above object, on the one hand, the present invention provides a Platycladus orientalis leaf extract, comprising the following raw materials in parts by weight: 20-30 parts of modified notoginseng total saponins, 5-15 parts of notoginseng, 10-30 parts of fibrin, 5-15 parts of chitosan, 5-15 parts of iron oxide, 2-6 parts of glycerol and 20-40 parts of modified spinach liquid;

[0007] The Platycladus orientalis leaf extract also includes:

[0008] Modified Platycladus orientalis leaf extract;

[0009] The mass ratio of the modified Platycladus orientalis leaf extract to the modified spinach extract is 1:(3-5);

[0010] The modified Platycladus orientalis leaves are prepared by extracting volatile oil through ultrasonic and acetone extraction method.

[0011] Furthermore, the preparation method of the modified Platycladus orientalis leaf extract comprises:

[0012] S11. Weigh 100 to 150 g of Platycladus orientalis leaf sample, wash the Platycladus orientalis leaf with distilled water, and then dry it at 30 to 50 ° C. After the dried Platycladus orientalis leaf is ground with a grinder, it is sieved through 80 to 90 mesh to obtain Platycladus orientalis leaf particles;

[0013] S12. The Platycladus orientalis leaf particles are continuously cleaned with an ultrasonic cleaner for 1 to 2 hours, and then sealed and soaked in 99 to 99.5% acetone for 2 to 3 hours to obtain a first soaking solution;

[0014] S13. The first soaking liquid is subjected to steam distillation for 3 to 5 hours to extract volatile oil to obtain a first extract, and then the first extract is extracted with 2 to 5 ml of petroleum ether. After extraction 3 to 5 times, a modified Platycladus orientalis leaf extract is obtained.

[0015] Furthermore, the preparation method of the modified notoginseng total saponins comprises:

[0016] S21. Wash and chop 200-220 g of Panax notoginseng root, then put it into 1000-1200 mL of deionized water, heat it to 90-100 ° C for 1-2 h to obtain a first mixed solution;

[0017] S22. Add 10-20 mL of sodium hydroxide to the cooled first mixed solution, adjust the pH to 9-10, and then extract in a water bath. The extraction time is 2-4 hours and the extraction temperature is 60-80°C to obtain a second mixed solution;

[0018] S23. After the extraction is completed, 10 to 20 mL of dilute nitric acid is added dropwise to the second mixed solution, the pH value is adjusted to about 3 to 5, and then placed in a freeze dryer, stirred at -5 to 0 ° C, and after stirring, filtered to remove the solid residue to obtain high-purity Panax notoginseng total saponins;

[0019] S24. Dissolve 10 to 15 g of lysine in 99.0 to 99.9% dimethylformamide in a first container, and dissolve 10 to 15 g of anhydride in 99.0 to 99.9% dimethylformamide in a second container, slowly dropwise add the liquid in the second container to the first container, and then magnetically stir at 35 to 45 ° C, at a stirring speed of 300 to 400 rpm, for 30 to 40 min to obtain an activated lysine solution;

[0020] S25. The activated lysine solution is slowly added dropwise to the high-purity total saponins of Panax notoginseng, and a coupling reaction is carried out at 10 to 15° C., followed by magnetic stirring at room temperature at a stirring speed of 500 to 700 rpm for 2 to 3 h. During the stirring process, 5 to 10 g of microbial transglutaminase is added to obtain a first modified solution;

[0021] S26. The first modified liquid is filtered, and then centrifuged with 2 to 5 ml of methanol. After centrifugation 3 to 5 times, modified Panax notoginseng total saponins are obtained.

[0022] Furthermore, the modified spinach liquid preparation method comprises:

[0023] S31. Wash the fresh radish leaves and spinach leaves separately, and then cut them into 1-3 cm small segments, and then weigh 100-150 g of radish leaves and spinach segments respectively;

[0024] S32. The weighed radish segments and spinach segments were placed in 1000-1500 mL of deionized water, boiled in a water bath at 80-90°C for 15-20 min, and then filtered to collect the filtrate, which was labeled as the first filtrate;

[0025] S33. Take 30-400 mL of a mixed solvent of acetone:n-hexane (mass ratio 1:1), add the first filtrate while stirring at room temperature, then let it stand for 30-40 minutes, then extract the first filtrate, collect the organic phase, and obtain a modified spinach liquid.

[0026] Furthermore, the modified spinach juice is rich in vitamin K, and the mass ratio of the modified spinach juice to the modified notoginseng total saponin is 2:(5-7).

[0027] Furthermore, the mass ratio of the fibrin to the modified notoginseng total saponins is (10-20): (5-10).

[0028] Furthermore, the mass ratio of the chitosan to the modified notoginseng total saponins is (1-3):(4-6).

[0029] Furthermore, the mass ratio of the iron oxide to the modified notoginseng total saponins is 3:(2-4).

[0030] On the other hand, based on the same inventive concept, the present invention also provides a method for preparing a Platycladus orientalis leaf extract, which is applied to the Platycladus orientalis leaf extract, comprising the following steps:

[0031] S41. Add deionized water to a container, add glycerol, Panax notoginseng and modified spinach liquid sequentially, stir, and heat in a water bath at a temperature of 50 to 65 ° C until fully dissolved to obtain a first mixed solution;

[0032] S42. The first mixed solution was filtered to obtain a second mixed solution, and the modified notoginseng total saponins and the modified Platycladus orientalis leaf extract were slowly added to the second mixed solution while stirring at a stirring speed of 200 to 500 rpm for 1 to 2 h to obtain a third mixed solution;

[0033] S43. The third mixed solution is subjected to steam distillation. After the distillation is completed, fibrin, chitosan and iron oxide are added in sequence, followed by magnetic stirring at a speed of 300 to 700 rpm for 5 to 6 hours. After the stirring is completed, an extract of Platycladus orientalis leaves is obtained.

[0034] The mechanism of action of the above raw material components is as follows:

[0035] Platycladus orientalis leaves, also known as cypress leaves, cypress leaves, and cypress leaves, are the dried young branches and leaves of Platycladus orientalis, a plant of the Cupressaceae family. Artificial cultivation is widespread throughout the country and is an excellent gardening tree species. According to traditional Chinese medicine theory, Platycladus orientalis leaves have a fresh scent, a bitter taste, and a slightly spicy taste. They have the effects of cooling blood and stopping bleeding, dispelling wind and dampness, resolving phlegm and relieving cough, reducing swelling and dispersing toxins, and promoting hair growth. The chemical composition of Platycladus orientalis leaves is complex, and the main chemical components are anise ketone, camphor, cypress biflavonoid apigenin, quercetin, kaempferol-7-0-glucoside, pinene, caryophyllene, palmitic acid, stearic acid, lauric acid, myristic acid, etc. It has the effects of antibacterial and anti-inflammatory, vasodilation, dandruff removal, anti-hair loss, and hair growth. Ancient books record that quercetin in Platycladus orientalis leaves has a hemostatic function. Modern medical research has found that Platycladus orientalis charcoal also has a hemostatic effect. The main component of the volatile oil in Platycladus orientalis leaves is α-ene, which has extremely strong medicinal value. It can work with other substances to promote blood coagulation, reduce bleeding, and has strong antibacterial and anti-inflammatory effects.

[0036] Modified Panax notoginseng total saponins are effective medicinal ingredients extracted from Panax notoginseng, usually in the form of brown to brown powder with a slightly bitter taste. Its main ingredients include a variety of ginsenosides and more than 20 saponin active substances, 17 trace elements, proteins, and rich vitamins. Modified Panax notoginseng total saponins can improve blood circulation, eliminate blood stasis, and also have anti-fatigue, anti-aging, and immunity-enhancing effects. It can also be used to treat symptoms such as hemoptysis, hematemesis, epistaxis, blood in the stool, metrorrhagia, and traumatic bleeding. It also has an auxiliary therapeutic effect on chest and abdominal pain, swelling and pain from falls, etc.

[0037] Modified spinach juice is a kind of food rich in vitamin K, Ga 2+ , vitamin C, iron and other nutrients. Among them, vitamin K plays an important role in the blood coagulation process. Vitamin K is a cofactor required for the synthesis of blood coagulation factors, especially coagulation factor II (heparin). Vitamin K deficiency can lead to bleeding tendency and cause coagulation factors to fail to activate normally. In addition, modified spinach liquid is rich in antioxidants that help inhibit the formation of free radicals and oxidative stress. The antioxidant effect can maintain vascular health and reduce the damage of oxidative stress to the blood system. At the same time, it can inhibit inflammatory responses and reduce vascular inflammation and fibrosis.

[0038] Fibrin is generally composed of collagen and elastin. It is a protein that is poorly soluble in water. It has a fibrous or thin rod-like appearance and a regular molecular structure. Fibrin plays an important role in maintaining blood vessel integrity, preventing blood loss, and promoting wound healing.

[0039] Chitosan is a polysaccharide naturally secreted by insects, usually produced by Asian red worms. Chitosan is mainly composed of two types of polysaccharides: aralic acid and isomers of aralic acid. Chitosan has low toxicity and can be used as a drug carrier. In addition, chitosan has good blood compatibility and biodegradability, and can promote platelet aggregation and activation of coagulation factors. It also has the ability to promote cell proliferation and differentiation, and can promote tissue repair and angiogenesis. In thrombus dressings, chitosan can promote wound healing and tissue repair, and reduce trauma and postoperative damage.

[0040] Iron oxide can affect the activity of the coagulation factor system and promote the coagulation process. It also has an antioxidant effect during the blood coagulation process. Iron oxide can bind to oxidative stress substances and inhibit the formation of free radicals, thereby protecting platelets and coagulation factor systems from oxidative damage and helping to maintain the health and normal function of the blood system.

[0041] Glycerol, also known as propylene glycol, is a colorless, odorless, sweet viscous liquid. Glycerol can be extracted from vegetable oils (such as soybean oil, palm oil, etc.) and animal oils (such as beef fat), mainly in the form of triglycerides. The antioxidant component of glycerol can be used to protect the blood system from oxidative damage and maintain the normal function of the coagulation system. Glycerol has an antibacterial effect and can prevent and treat infections, especially in trauma and postoperative applications.

[0042] (3) Beneficial effects

[0043] Compared with the prior art, the present invention has the following beneficial effects:

[0044] 1. The modified Platycladus orientalis leaf extract, modified Panax notoginseng total saponin and modified spinach liquid work through synergistic effect. The modified Platycladus orientalis leaf extract and modified Panax notoginseng total saponin jointly promote platelet aggregation and the release of coagulation factors to accelerate blood coagulation. The modified spinach liquid promotes the synthesis and release of coagulation factors, accelerates blood coagulation and achieves a hemostatic effect.

[0045] 2. The modified Platycladus orientalis leaf extract contains a large amount of volatile oil, which can stimulate platelet aggregation by binding to receptors on the surface of platelets. It can also promote the release of coagulation factors and further accelerate the coagulation process.

[0046] 3. Modified Panax notoginseng total saponins inhibit platelet activation and aggregation by regulating the increase of cyclic adenosine monophosphate content in platelets. Cyclic adenosine monophosphate, as an important second messenger molecule, plays a key regulatory role in platelets. At the same time, modified Panax notoginseng total saponins can inhibit the generation of thromboxane A2, improve vascular endothelial function, and inhibit platelet aggregation through multiple pathways;

[0047] 4. Modified spinach liquid contains various nutrients, among which vitamin K is a necessary coenzyme for the synthesis of coagulation factors 2, 7, 9, and 10. These coagulation factors can participate in the synthesis of coagulation factors during the coagulation process to form a fibrin network and achieve the purpose of hemostasis. In addition, vitamin K can regulate the release of coagulation factors, effectively promote the synthesis and release of coagulation factors, and accelerate blood coagulation. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] Figure 1 This is the SEM image of the volatile oil in the Platycladus orientalis leaves in Example 1 of the present invention. DETAILED DESCRIPTION

[0049] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0050] The experimental equipment and preparations of the embodiments described below are as follows: steam distillation apparatus, KO5200DA type numerically controlled ultrasonic cleaner, electronic balance (Sartorius, Germany), magnetic stirrer (Meiyingpu, Shanghai), high-speed centrifuge (Guangzhou Jidi), freeze dryer (Shanghai Pudong Freeze Drying), high-speed pulverizer (Shandong Tianfang Machinery), pH meter (Shanghai Yidian), oscillator (Shanghai Analyzer), rotary evaporator (Rikakikai, Japan), SB-1000 water bath (Rikakikai, Japan), scanning electron microscope (Zeiss, Germany); chemicals and reagents were purchased from Sigma-Aldrich.

[0051] Example 1: This example discloses a Platycladus orientalis leaf extract, comprising the following raw materials in parts by weight: 20 parts of modified Panax notoginseng total saponins, 5 parts of Panax notoginseng, 10 parts of fibrin, 5 parts of chitosan, 5 parts of iron oxide, 2 parts of glycerol and 20 parts of modified spinach liquid, the mass ratio of the modified Platycladus orientalis leaf extract to the modified spinach liquid is 1:3; the modified Platycladus orientalis leaf is extracted with volatile oil by ultrasonic and acetone extraction method.

[0052] In the Platycladus orientalis leaf extract, the modified Platycladus orientalis leaf extract, the modified Panax notoginseng total saponin and the modified spinach liquid work through a synergistic effect. The volatile oil component in the modified Platycladus orientalis leaf extract is the key to rapid hemostasis. It directly acts on the receptors on the surface of the platelet membrane, activates the signal transduction pathway inside the platelets, and promotes the platelets to change from a resting state to an activated state. Activated platelets release ADP and 5-HT active substances, which can further stimulate more platelet aggregation. At the same time, activated platelets also promote the release of coagulation factor V and coagulation factor VIII, thereby accelerating the coagulation cascade reaction. Then, excessive activation and activation of platelets can lead to some reactions of thrombosis. Therefore, modified Panax notoginseng total saponins can moderately inhibit the excessive activation of platelets, change the morphology of platelets by regulating the expression of platelet membrane glycoprotein receptors, and at the same time, reduce the secretion of dense granules and reduce platelet adhesion to achieve the purpose of reducing platelet concentration. In addition, modified Panax notoginseng total saponins can also reduce the calcium ion concentration in platelets, maintain platelets at a moderate activation level, and regulate platelet concentration as a whole. Fibrin forms a three-dimensional network structure, which not only provides an adhesion scaffold for platelets, but also enhances the strength of blood clots. At the same time, modified Panax notoginseng total saponins can also balance the procoagulant and anticoagulant systems by regulating the synthesis of coagulation factors, preventing the risk of thrombosis caused by excessive platelet aggregation. Chitosan uses its positive charge characteristics and strong water absorption properties to adsorb blood components and promote the activation of coagulation factors. In addition, vitamin K in the modified spinach liquid enhances the hemostatic effect by promoting the synthesis and release of coagulation factors. As an essential auxiliary factor, vitamin K participates in the γ-carboxylation modification process of various coagulation factors and promotes the synthesis of vitamin K-dependent coagulation factors such as II, VII, IX, and X. The increase of these active coagulation factors can accelerate the conversion of prothrombin to thrombin and promote the formation of fibrin, thereby consolidating the hemostatic effect and improving the activity of coagulation factors. Iron oxide forms a mechanical barrier by contracting blood vessels and promoting local tissue coagulation, and provides metal ions to accelerate the coagulation cascade reaction. Glycerol optimizes the overall performance by adjusting the viscosity of the material, maintaining water balance, and enhancing the dispersibility of other components. The three components of modified Platycladus orientalis leaf extract, modified Panax notoginseng total saponins, and modified spinach liquid form a network of mutual restraint and mutual promotion, and finally form a dynamically balanced hemostatic system, which can not only quickly and effectively promote hemostasis, but also prevent thrombosis, and ensure the continuity of the hemostatic effect.

[0053] The preparation method of the modified Platycladus orientalis leaf extract comprises:

[0054] S11. Weigh 150 g of Platycladus orientalis leaf sample, wash the Platycladus orientalis leaf with distilled water, and then dry it at 30 to 50 ° C. After drying, grind the dried Platycladus orientalis leaf with a grinder, and pass it through a 80 to 90 mesh sieve to obtain Platycladus orientalis leaf particles;

[0055] S12. The Platycladus orientalis leaf particles are continuously cleaned with an ultrasonic cleaner for 1 to 2 hours, and then sealed and soaked in 99 to 99.5% acetone for 2 to 3 hours to obtain a first soaking solution;

[0056] S13. The first soaking liquid is subjected to steam distillation for 3 to 5 hours to extract volatile oil to obtain a first extract, and then the first extract is extracted with 2 to 5 ml of petroleum ether. After extraction 3 to 5 times, a modified Platycladus orientalis leaf extract is obtained.

[0057] The preparation method of the modified Panax notoginseng total saponins comprises:

[0058] S21. Wash and chop 200 g of Panax notoginseng root, then put it into 1000-1200 mL of deionized water, heat it to 90-100 ° C for 1-2 h to obtain a first mixed solution;

[0059] S22. Add 10-20 mL of sodium hydroxide to the cooled first mixed solution, adjust the pH to 9-10, and then extract in a water bath. The extraction time is 2-4 hours and the extraction temperature is 60-80°C to obtain a second mixed solution;

[0060] S23. After the extraction is completed, 10 to 20 mL of dilute nitric acid is added dropwise to the second mixed solution, the pH value is adjusted to about 3 to 5, and then placed in a freeze dryer, stirred at -5 to 0 ° C, and after stirring, filtered to remove the solid residue to obtain high-purity Panax notoginseng total saponins;

[0061] S24. Dissolve 10 to 15 g of lysine in 99.0 to 99.9% dimethylformamide in a first container, and dissolve 10 to 15 g of anhydride in 99.0 to 99.9% dimethylformamide in a second container, slowly dropwise add the liquid in the second container to the first container, and then magnetically stir at 35 to 45 ° C, at a stirring speed of 300 to 400 rpm, for 30 to 40 min to obtain an activated lysine solution;

[0062] S25. The activated lysine solution is slowly added dropwise to the high-purity total saponins of Panax notoginseng, and a coupling reaction is carried out at 10 to 15° C., followed by magnetic stirring at room temperature at a stirring speed of 500 to 700 rpm for 2 to 3 h. During the stirring process, 5 to 10 g of microbial transglutaminase is added to obtain a first modified solution;

[0063] S26. The first modified liquid is filtered, and then centrifuged with 2 to 5 ml of methanol. After centrifugation 3 to 5 times, modified Panax notoginseng total saponins are obtained.

[0064] The preparation method of the modified spinach liquid comprises:

[0065] S31. Wash the fresh radish leaves and spinach leaves separately, and then chop them into 1-3 cm small segments, and then weigh 100 g of radish leaves and spinach segments respectively;

[0066] S32. The weighed radish segments and spinach segments were placed in 1000-1500 mL of deionized water, boiled in a water bath at 80-90°C for 15-20 min, and then filtered to collect the filtrate, which was labeled as the first filtrate;

[0067] S33. Take 30-400 mL of a mixed solvent of acetone:n-hexane (mass ratio 1:1), add the first filtrate while stirring at room temperature, then let it stand for 30-40 minutes, then extract the first filtrate, collect the organic phase, and obtain a modified spinach liquid.

[0068] The mass ratio of the modified spinach liquid to the modified notoginseng total saponin is 2:5.

[0069] The mass ratio of the fibrin to the modified total saponins of Panax notoginseng is 10:5.

[0070] The mass ratio of the chitosan to the modified notoginseng total saponins is 1:4.

[0071] The mass ratio of the iron oxide to the modified notoginseng total saponins is 3:2.

[0072] A method for preparing a Platycladus orientalis leaf extract, applied to the Platycladus orientalis leaf extract, comprises the following steps:

[0073] S41. Add deionized water to a container, add glycerol, Panax notoginseng and modified spinach liquid sequentially, stir, and heat in a water bath at a temperature of 50 to 65 ° C until fully dissolved to obtain a first mixed solution;

[0074] S42. The first mixed solution was filtered to obtain a second mixed solution, and the modified notoginseng total saponins and the modified Platycladus orientalis leaf extract were slowly added to the second mixed solution while stirring at a stirring speed of 200 to 500 rpm for 1 to 2 h to obtain a third mixed solution;

[0075] S43. The third mixed solution is subjected to steam distillation. After the distillation is completed, fibrin, chitosan and iron oxide are added in sequence, followed by magnetic stirring at a speed of 300 to 700 rpm for 5 to 6 hours. After the stirring is completed, an extract of Platycladus orientalis leaves is obtained.

[0076] Example 2: This example discloses a Platycladus orientalis leaf extract, comprising the following raw materials in parts by weight: 25 parts of modified Panax notoginseng total saponins, 10 parts of Panax notoginseng, 15 parts of fibrin, 10 parts of chitosan, 10 parts of iron oxide, 4 parts of glycerol and 30 parts of modified spinach liquid, the mass ratio of the modified Platycladus orientalis leaf extract to the modified spinach liquid is 1:4; the modified Platycladus orientalis leaf is extracted with volatile oil by ultrasonic and acetone extraction method.

[0077] The preparation method of the modified Platycladus orientalis leaf extract comprises:

[0078] S11. Weigh 100 g of Platycladus orientalis leaf sample, wash the Platycladus orientalis leaf with distilled water, and then dry it at 30 to 50 ° C. After drying, grind the dried Platycladus orientalis leaf with a grinder, and pass it through a 80 to 90 mesh sieve to obtain Platycladus orientalis leaf particles;

[0079] S12. The Platycladus orientalis leaf particles are continuously cleaned with an ultrasonic cleaner for 1 to 2 hours, and then sealed and soaked in 99 to 99.5% acetone for 2 to 3 hours to obtain a first soaking solution;

[0080] S13. The first soaking liquid is subjected to steam distillation for 3 to 5 hours to extract volatile oil to obtain a first extract, and then the first extract is extracted with 2 to 5 ml of petroleum ether. After extraction 3 to 5 times, a modified Platycladus orientalis leaf extract is obtained.

[0081] The preparation method of the modified Panax notoginseng total saponins comprises:

[0082] S21. Wash and chop 210 g of Panax notoginseng root, then put it into 1000-1200 mL of deionized water, heat it to 90-100 ° C for 1-2 h to obtain a first mixed solution;

[0083] S22. Add 10-20 mL of sodium hydroxide to the cooled first mixed solution, adjust the pH to 9-10, and then extract in a water bath. The extraction time is 2-4 hours and the extraction temperature is 60-80°C to obtain a second mixed solution;

[0084] S23. After the extraction is completed, 10 to 20 mL of dilute nitric acid is added dropwise to the second mixed solution, the pH value is adjusted to about 3 to 5, and then placed in a freeze dryer, stirred at -5 to 0 ° C, and after stirring, filtered to remove the solid residue to obtain high-purity Panax notoginseng total saponins;

[0085] S24. Dissolve 10 to 15 g of lysine in 99.0 to 99.9% dimethylformamide in a first container, and dissolve 10 to 15 g of anhydride in 99.0 to 99.9% dimethylformamide in a second container, slowly dropwise add the liquid in the second container to the first container, and then magnetically stir at 35 to 45 ° C, at a stirring speed of 300 to 400 rpm, for 30 to 40 min to obtain an activated lysine solution;

[0086] S25. The activated lysine solution is slowly added dropwise to the high-purity total saponins of Panax notoginseng, and a coupling reaction is carried out at 10 to 15° C., followed by magnetic stirring at room temperature at a stirring speed of 500 to 700 rpm for 2 to 3 h. During the stirring process, 5 to 10 g of microbial transglutaminase is added to obtain a first modified solution;

[0087] S26. The first modified liquid is filtered, and then centrifuged with 2 to 5 ml of methanol. After centrifugation 3 to 5 times, modified Panax notoginseng total saponins are obtained.

[0088] The preparation method of the modified spinach liquid comprises:

[0089] S31. Wash the fresh radish leaves and spinach leaves separately, and then chop them into 1-3 cm small segments, and then weigh 120 g of radish leaves and spinach segments respectively;

[0090] S32. The weighed radish segments and spinach segments were placed in 1000-1500 mL of deionized water, boiled in a water bath at 80-90°C for 15-20 min, and then filtered to collect the filtrate, which was labeled as the first filtrate;

[0091] S33. Take 30-400 mL of a mixed solvent of acetone:n-hexane (mass ratio 1:1), add the first filtrate while stirring at room temperature, then let it stand for 30-40 minutes, then extract the first filtrate, collect the organic phase, and obtain a modified spinach liquid.

[0092] The mass ratio of the modified spinach liquid to the modified notoginseng total saponin is 2:6.

[0093] The mass ratio of the fibrin to the modified total saponins of Panax notoginseng is 15:10.

[0094] The mass ratio of the chitosan to the modified notoginseng total saponins is 1:5.

[0095] The mass ratio of the iron oxide to the modified total saponins of Panax notoginseng is 1:1. The preparation method of a Platycladus orientalis leaf extract in this embodiment is consistent with that in the first embodiment.

[0096] Example 3: This example discloses a Platycladus orientalis leaf extract, comprising the following raw materials in parts by weight: 25 parts of modified Panax notoginseng total saponins, 10 parts of Panax notoginseng, 15 parts of fibrin, 10 parts of chitosan, 10 parts of iron oxide, 4 parts of glycerol and 30 parts of modified spinach liquid, the mass ratio of the modified Platycladus orientalis leaf extract to the modified spinach liquid is 1:4; the modified Platycladus orientalis leaf is extracted with volatile oil by ultrasonic and acetone extraction method.

[0097] The preparation method of the modified Platycladus orientalis leaf extract comprises:

[0098] S11. Weigh 120 g of Platycladus orientalis leaf sample, wash the Platycladus orientalis leaf with distilled water, and then dry it at 30 to 50 ° C. After drying, grind the dried Platycladus orientalis leaf with a grinder, and pass it through a 80 to 90 mesh sieve to obtain Platycladus orientalis leaf particles;

[0099] S12. The Platycladus orientalis leaf particles are continuously cleaned with an ultrasonic cleaner for 1 to 2 hours, and then sealed and soaked in 99 to 99.5% acetone for 2 to 3 hours to obtain a first soaking solution;

[0100] S13. The first soaking liquid is subjected to steam distillation for 3 to 5 hours to extract volatile oil to obtain a first extract, and then the first extract is extracted with 2 to 5 ml of petroleum ether. After extraction 3 to 5 times, a modified Platycladus orientalis leaf extract is obtained.

[0101] The preparation method of the modified Panax notoginseng total saponins comprises:

[0102] S21. Wash and chop 220 g of Panax notoginseng root, then put it into 1000-1200 mL of deionized water, heat it to 90-100 ° C for 1-2 h to obtain a first mixed solution;

[0103] S22. Add 10-20 mL of sodium hydroxide to the cooled first mixed solution, adjust the pH to 9-10, and then extract in a water bath. The extraction time is 2-4 hours and the extraction temperature is 60-80°C to obtain a second mixed solution;

[0104] S23. After the extraction is completed, 10 to 20 mL of dilute nitric acid is added dropwise to the second mixed solution, the pH value is adjusted to about 3 to 5, and then placed in a freeze dryer, stirred at -5 to 0 ° C, and after stirring, filtered to remove the solid residue to obtain a modified high-purity Panax notoginseng total saponin;

[0105] S24. Dissolve 10 to 15 g of lysine in 99.0 to 99.9% dimethylformamide in a first container, and dissolve 10 to 15 g of anhydride in 99.0 to 99.9% dimethylformamide in a second container, slowly dropwise add the liquid in the second container to the first container, and then magnetically stir at 35 to 45 ° C, at a stirring speed of 300 to 400 rpm, for 30 to 40 min to obtain an activated lysine solution;

[0106] S25. The activated lysine solution is slowly added dropwise to the high-purity total saponins of Panax notoginseng, and a coupling reaction is carried out at 10 to 15° C., followed by magnetic stirring at room temperature at a stirring speed of 500 to 700 rpm for 2 to 3 h. During the stirring process, 5 to 10 g of microbial transglutaminase is added to obtain a first modified solution;

[0107] S26. The first modified liquid is filtered, and then centrifuged with 2 to 5 ml of methanol. After centrifugation 3 to 5 times, modified Panax notoginseng total saponins are obtained.

[0108] The preparation method of the modified spinach liquid comprises:

[0109] S31. Wash the fresh radish leaves and spinach leaves separately, and then cut them into 1-3 cm small segments, and then weigh 150 g of radish leaves and spinach segments respectively;

[0110] S32. The weighed radish segments and spinach segments were placed in 1000-1500 mL of deionized water, boiled in a water bath at 80-90°C for 15-20 min, and then filtered to collect the filtrate, which was labeled as the first filtrate;

[0111] S33. Take 30-400 mL of a mixed solvent of acetone:n-hexane (mass ratio 1:1), add the first filtrate while stirring at room temperature, then let it stand for 30-40 minutes, then extract the first filtrate, collect the organic phase, and obtain a modified spinach liquid.

[0112] The mass ratio of the modified spinach liquid to the modified notoginseng total saponin is 2:7.

[0113] The mass ratio of the fibrin and the modified total saponins of Panax notoginseng is 20:10.

[0114] The mass ratio of the chitosan to the modified notoginseng total saponins is 1:4.

[0115] The mass ratio of the iron oxide to the modified total saponins of Panax notoginseng is 3:4. The preparation method of a Platycladus orientalis leaf extract in this embodiment is consistent with that in the first embodiment.

[0116] Example 4: This example discloses a Platycladus orientalis leaf extract, comprising the following raw materials in parts by weight: 30 parts of modified Panax notoginseng total saponins, 10 parts of Panax notoginseng, 15 parts of fibrin, 10 parts of chitosan, 10 parts of iron oxide, 4 parts of glycerol and 30 parts of modified spinach liquid, the mass ratio of the modified Platycladus orientalis leaf extract to the modified spinach liquid is 1:3; the modified Platycladus orientalis leaf is extracted with volatile oil by ultrasonic and acetone extraction method.

[0117] The mass ratio of the modified spinach liquid to the modified notoginseng total saponin is 2:5.

[0118] The mass ratio of the fibrin to the modified total saponins of Panax notoginseng is 10:5.

[0119] The mass ratio of the chitosan to the modified notoginseng total saponins is 1:4.

[0120] The mass ratio of the iron oxide to the modified total saponins of Panax notoginseng is 3:2. The preparation method of a Platycladus orientalis leaf extract in this embodiment is consistent with that in the first embodiment.

[0121] Example 5: This example discloses a Platycladus orientalis leaf extract, comprising the following raw materials in parts by weight: 20 parts of modified Panax notoginseng total saponins, 10 parts of Panax notoginseng, 15 parts of fibrin, 10 parts of chitosan, 10 parts of iron oxide, 4 parts of glycerol and 20 parts of modified spinach liquid, the mass ratio of the modified Platycladus orientalis leaf extract to the modified spinach liquid is 1:3; the modified Platycladus orientalis leaf is extracted with volatile oil by ultrasonic and acetone extraction method.

[0122] The mass ratio of the modified spinach liquid to the modified notoginseng total saponin is 2:5.

[0123] The mass ratio of the fibrin to the modified total saponins of Panax notoginseng is 10:5.

[0124] The mass ratio of the chitosan to the modified notoginseng total saponins is 1:4.

[0125] The mass ratio of the iron oxide to the modified total saponins of Panax notoginseng is 3:2. The preparation method of a Platycladus orientalis leaf extract in this embodiment is consistent with that in the first embodiment.

[0126] Control group 1: This example differs from Example 1 in that the modified Platycladus orientalis leaf extract is not contained. The preparation method of the Platycladus orientalis leaf extract in this example is the same as that in Example 1.

[0127] Control group 2: This example differs from Example 1 in that it does not contain modified notoginseng total saponins. The preparation method of a Platycladus orientalis leaf extract in this example is the same as that in Example 1.

[0128] Control group 3: This example differs from Example 1 in that the modified spinach liquid is not contained. The preparation method of a Platycladus orientalis leaf extract in this example is the same as that in Example 1.

[0129] Control group 4: This example differs from Example 1 in that it does not contain modified spinach liquid and modified Platycladus orientalis leaf extract. The preparation method of Platycladus orientalis leaf extract in this example is the same as that in Example 1.

[0130] Control group 5: This example differs from Example 1 in that it does not contain modified notoginseng total saponins and modified Platycladus orientalis leaf extract. The preparation method of Platycladus orientalis leaf extract in this example is the same as that in Example 1.

[0131] Control group 6: This example differs from Example 1 in that: Modified notoginseng total saponins and modified spinach liquid are not contained. The preparation method of Platycladus orientalis leaf extract in this example is the same as that in Example 1.

[0132] Control group 7: This example differs from Example 1 in that it does not contain modified notoginseng total saponins, modified Platycladus orientalis leaf extract and modified spinach liquid. The preparation method of Platycladus orientalis leaf extract in this example is the same as that in Example 1.

[0133] Effect evaluation: Analysis of blood coagulation performance of Platycladus orientalis leaf extract: The Platycladus orientalis leaf extract prepared in each experimental group was randomly selected and divided into 5 parts on average. Blood from the common carotid artery of rabbits was taken and the effect of Platycladus orientalis leaf extract on blood coagulation in vitro was observed by using the coagulation plate method, with coagulation time (CT), activated partial thromboplastin time (APTT) and platelet aggregation rate as indicators. Table 1 is the statistical results of the factors affecting blood coagulation of Platycladus orientalis leaf extract. It can be seen from Table 1 that there are certain differences in the proportions of Platycladus orientalis leaf extracts in each experimental group. By comparing the coagulation time of Platycladus orientalis leaf extracts prepared in Examples 1 to 5 with that of Control Groups 1 to 7, it can be found that, on the whole, the Platycladus orientalis leaf extracts prepared in Examples 1 to 5 have better blood coagulation, and the average coagulation time is lower than that of the control group, and the Platycladus orientalis leaf extract prepared in Example 1 has the best effect on blood coagulation and the shortest coagulation time. This shows that the synergistic effect of modified Platycladus orientalis leaf extract, modified Panax notoginseng total saponins and modified spinach liquid plays a great role. Modified Platycladus orientalis leaf extract and modified Panax notoginseng total saponins jointly promote platelet aggregation and the release of coagulation factors to accelerate blood coagulation. Modified spinach liquid promotes the synthesis and release of coagulation factors, accelerates blood coagulation and achieves hemostatic effect. By comparing the Platycladus orientalis leaf extract prepared in Example 1 with control groups 1 to 7, it can be found that the coagulation time, activated partial thromboplastin time and platelet aggregation rate are all in an optimal range, among which the coagulation time is the shortest among the entire experimental group and the control group, and blood coagulation is completed in only 30 seconds. The activated partial thromboplastin time is also the shortest among all experimental groups, taking 25 seconds. Among them, the platelet aggregation rate is 10.6%, which is the highest in the entire experimental group. The normal value of the platelet aggregation rate is generally 9% to 13%, indicating that when the modified Platycladus orientalis leaf extract, modified Panax notoginseng total saponins and modified spinach liquid are added simultaneously during the preparation of the Platycladus orientalis leaf extract, the biomedical value of the Platycladus orientalis leaf extract can be significantly improved.

[0134] Table 1 Statistical results of factors affecting blood coagulation in Platycladus orientalis leaf extracts in each experimental group

[0135] Experimental Group Coagulation time (s) Activated partial thromboplastin time (s) Platelet aggregation rate (%) Example 1 30 25 10.6 Example 2 33 26 10.3 Example 3 36 26 10.1 Example 4 34 26 10.2 Example 5 35 27 10.4 Control group 1 50 30 9.9 Control group 2 40 30 10.0 Control group 3 49 32 9.2 Control group 4 61 36 9.6 Control group 5 66 33 9.5 Control group 6 63 35 9.0 Control group 7 69 40 9.0

[0136] Through the above limited experiments, the Platycladus orientalis leaf extract material of Example 1 of the present invention has significant effect. By optimizing the preparation method of Platycladus orientalis leaf extracted volatile oil, modified Platycladus orientalis leaf extract, modified Panax notoginseng total saponins and modified spinach liquid and other plant ingredients are added in the process of preparing the Platycladus orientalis leaf extract, the medicinal value of Platycladus orientalis leaves is significantly improved, and the effect of Platycladus orientalis leaf extract on blood coagulation is improved, and finally a high-quality Platycladus orientalis leaf extract is obtained. The obtained Platycladus orientalis leaf extract has great potential for biomedical applications. The Platycladus orientalis leaf extract raw material can be applied to cosmetics, medicine and other fields to achieve large-scale production, thereby promoting the healthy and orderly development of the industry.

[0137] Finally, it should be noted that: Although the present invention has been described in detail with reference to the aforementioned embodiments, it is still possible for those skilled in the art to modify the technical solutions described in the aforementioned embodiments, or to make equivalent substitutions for some of the technical features therein. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A Platycladus orientalis leaf extract, characterized in that The invention comprises the following raw materials in parts by weight: 20-30 parts of modified notoginseng total saponins, 5-15 parts of notoginseng, 10-30 parts of fibrin, 5-15 parts of chitosan, 5-15 parts of iron oxide, 2-6 parts of glycerol and 20-40 parts of modified spinach liquid. The Platycladus orientalis leaf extract also includes: Modified Platycladus orientalis leaf extract; The mass ratio of the modified Platycladus orientalis leaf extract to the modified spinach extract is 1:(3-5); The modified Platycladus orientalis leaves are prepared by extracting volatile oil through ultrasonic and acetone extraction method.

2. The Platycladus orientalis leaf extract according to claim 1, characterized in that The preparation method of the modified Platycladus orientalis leaf extract comprises: S11. Weigh 100 to 150 g of Platycladus orientalis leaf sample, wash the Platycladus orientalis leaf with distilled water, and then dry it at 30 to 50 ° C. After the dried Platycladus orientalis leaf is ground with a grinder, it is sieved through 80 to 90 mesh to obtain Platycladus orientalis leaf particles; S12. The Platycladus orientalis leaf particles are continuously cleaned with an ultrasonic cleaner for 1 to 2 hours, and then sealed and soaked in 99 to 99.5% acetone for 2 to 3 hours to obtain a first soaking solution; S13. The first soaking liquid is subjected to steam distillation for 3 to 5 hours to extract volatile oil to obtain a first extract, and then the first extract is extracted with 2 to 5 ml of petroleum ether. After extraction 3 to 5 times, a modified Platycladus orientalis leaf extract is obtained.

3. The Platycladus orientalis leaf extract according to claim 1, characterized in that The preparation method of the modified Panax notoginseng total saponins comprises: S21. Wash and chop 200-220 g of Panax notoginseng root, then soak it in 99.0-99.9% dimethylformamide at room temperature for 1-2 h, take out the chopped Panax notoginseng root after soaking, put it into 1000-1200 mL of deionized water, heat it to 90-100 ° C, and heat it for 1-2 h to obtain a first mixed solution; S22. Add 10-20 mL of sodium hydroxide to the cooled first mixed solution, adjust the pH to 9-10, and then extract in a water bath. The extraction time is 2-4 hours and the extraction temperature is 60-80°C to obtain a second mixed solution; S23. After the extraction is completed, 10 to 20 mL of dilute nitric acid is added dropwise to the second mixed solution, the pH value is adjusted to about 3 to 5, and then placed in a freeze dryer, stirred at -5 to 0 ° C, and after stirring, filtered to remove the solid residue to obtain high-purity Panax notoginseng total saponins; S24. Dissolve 10 to 15 g of lysine in 99.0 to 99.9% dimethylformamide in a first container, and dissolve 10 to 15 g of anhydride in 99.0 to 99.9% dimethylformamide in a second container, slowly dropwise add the liquid in the second container to the first container, and then magnetically stir at 35 to 45 ° C, at a stirring speed of 300 to 400 rpm, for 30 to 40 min to obtain an activated lysine solution; S25. The activated lysine solution is slowly added dropwise to the high-purity total saponins of Panax notoginseng, and a coupling reaction is carried out at 10 to 15° C., followed by magnetic stirring at room temperature at a stirring speed of 500 to 700 rpm for 2 to 3 h. During the stirring process, 5 to 10 g of microbial transglutaminase is added to obtain a first modified solution; S26. The first modified liquid is filtered, and then centrifuged with 2 to 5 ml of methanol. After centrifugation 3 to 5 times, modified Panax notoginseng total saponins are obtained.

4. The Platycladus orientalis leaf extract according to claim 1, characterized in that: The preparation method of the modified spinach liquid comprises: S31. Wash the fresh radish leaves and spinach leaves separately, and then cut them into 1-3 cm small segments, and then weigh 100-150 g of radish leaves and spinach segments respectively; S32. The weighed radish segments and spinach segments were placed together in 1000-1500 mL of deionized water, boiled in a water bath at 80-90°C for 15-20 min, and then filtered to collect the filtrate, which was labeled as the first filtrate; S33. Take 30-400 mL of a mixed solvent of acetone:n-hexane (mass ratio 1:1), add the first filtrate while stirring at room temperature, then let it stand for 30-40 minutes, then extract the first filtrate, collect the organic phase, and obtain a modified spinach liquid. The modified spinach liquid is rich in vitamin K and Ga 2+ Content, the mass ratio of modified spinach liquid and modified Panax notoginseng total saponins is 2: (5-7).

5. The Platycladus orientalis leaf extract according to claim 1, characterized in that The mass ratio of the fibrin to the modified total saponin of Panax notoginseng is (10-20): (5-10).

6. The Platycladus orientalis leaf extract according to claim 1, characterized in that The mass ratio of the chitosan to the modified notoginseng total saponin is (1-3): (4-6).

7. The Platycladus orientalis leaf extract according to claim 1, characterized in that The mass ratio of the iron oxide to the modified notoginseng total saponins is 3:(2-4).

8. A method for preparing a Platycladus orientalis leaf extract, which is used to prepare a Platycladus orientalis leaf extract as claimed in any one of claims 1 to 7, characterized in that: The method comprises the following steps: S41. Add deionized water to a container, add glycerol, Panax notoginseng and modified spinach liquid sequentially, stir, and heat in a water bath at a temperature of 50 to 65 ° C until fully dissolved to obtain a first mixed solution; S42. The first mixed solution was filtered to obtain a second mixed solution, and the modified notoginseng total saponins and the modified Platycladus orientalis leaf extract were slowly added to the second mixed solution while stirring at a stirring speed of 200 to 500 rpm for 1 to 2 h to obtain a third mixed solution; S43. The third mixed solution is subjected to steam distillation. After the distillation is completed, fibrin, chitosan and iron oxide are added in sequence, followed by magnetic stirring at a speed of 300 to 700 rpm for 5 to 6 hours. After the stirring is completed, an extract of Platycladus orientalis leaves is obtained.

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