A crocetin preparation with high bioavailability and its application

Abstract

The present invention belongs to the technical field of pharmaceutical preparations, and provides a crocetin preparation with high bioavailability and its application. The present invention uses sodium crocinate as the main active ingredient, and for the first time proposes to prepare it into an oral enteric-coated preparation with high bioavailability and fast absorption. By optimizing the formulation process of the oral preparation, using lipid materials, solubilizers and pH regulators as the key components of the sodium crocinate enteric-coated preparation, the stability, solubility and absorption efficiency of the drug components are significantly increased, the bioavailability of the poorly soluble crocetin preparation by oral administration is improved, and the pharmacokinetic parameters and efficacy of the drug are improved. The oral enteric-coated preparation of the present invention can effectively prevent or treat cardiovascular and cerebrovascular diseases and metabolic diseases, greatly expanding the clinical value and drug-likeness of crocetin, and having a simple preparation process and low cost.

Description

Technical Field

[0001] The present invention belongs to the technical field of pharmaceutical preparations, and particularly relates to a crocetin preparation with high bioavailability and its application. Background Art

[0002] Crocus is the dried stigma of the plant Crocus sativus L. of the family Iridaceae. It is mild in nature and sweet in taste, and belongs to the heart and liver meridians, with the effects of promoting blood circulation to remove blood stasis, cooling blood and detoxifying, and relieving depression and calming the nerves. Crocetin is one of the active ingredients of crocus, also known as safranal, which exists in saffron and gardenia plants. It is a natural carotenoid dicarboxylic acid with the molecular formula C 20 H 24 O4 and a molecular weight of 328.40 g / mol. Crocetin is a hydrophobic compound, insoluble in water and most organic solvents. Its anionic form is soluble in water and easily dissolves in dilute sodium hydroxide aqueous solution or other alkaline aqueous solutions. Since crocetin is a polyene compound with conjugated double bonds having isoprene as residues, its structural stability is poor, its bioavailability is low, and it is easily degraded, which affects the safety and effectiveness of drugs. With the continuous in-depth research on crocetin, crocetin has various pharmacological effects such as protecting the heart, protecting the nerves, protecting the liver, antidepressant, improving Alzheimer's disease and Parkinson's disease, etc., and has great development potential.

[0003] In modern pharmacology, crocetin has various curative effects such as increasing the oxygen diffusion rate, preventing atherosclerosis, reducing blood lipid, reducing blood pressure, and increasing blood flow to the eyes, because crocetin itself has a significant antioxidant effect. Research shows that crocetin can effectively scavenge free radicals in the body, activate the mitogen-activated protein kinase (MAPK) and PI3K / ATK pathways, reduce the production of reactive oxygen species (ROS) and cardiomyocyte apoptosis, and thus play a role in protecting against heart failure. Crocetin can also inhibit mitochondrial function by scavenging free radicals in clinical experiments, activate the AMPK pathway, reduce intracellular fat oxidation and the expression of cell immune antigens at the lesion site, promote glucose uptake, accelerate fat consumption, and reverse myocardial damage. Lautenschlager et al. demonstrated that crocetin can penetrate the blood-brain barrier, which may be the mechanism of crocetin exerting its pharmacological effects in the central nervous system, making crocetin have the potential to prevent and treat cardiovascular and cerebrovascular diseases.

[0004] However, due to factors such as the high price of saffron, poor solubility and stability, and extremely low bioavailability, crocetin-related preparations are currently mostly injected intravenously in clinical trials to improve the bioavailability of the drug, but the half-life of the drug is very short. Crocetin reaches its maximum plasma concentration approximately 0.5 hours after administration, indicating that the rapid absorption of crocetin may be due to transport into the bloodstream via the portal vein. In clinical trials on healthy humans, after a single oral dose of 16 mg crocetin, the crocetin concentration range at different sampling intervals was approximately 0.09-0.35 μg / mL. Clinical experimental observations of the pharmacokinetics of crocetin in 10 healthy humans found that crocetin can be detected in human plasma 1 hour after a single dose of 7.5, 15, and 22.5 mg, and its t max Range 4.0-.8h, t 1 / 2 6.1-7.5h, C max and AUC 0-24h The average values ​​were 100.9-279.7 ng / mL and 556.5-1720.8 ng / mL, respectively, and were dose-dependent. In contrast, the crocetin metabolite - crocetin monoglucuronic acid was still very low. When administered to rats by gavage at a dose of 25 mg / kg, the oral bioavailability of crocetin was only 11.25%. Due to the rapid absorption and metabolism of crocetin, repeated oral administration of crocetin will not accumulate in plasma and increase blood drug concentration, and the bioavailability of crocetin is still very low. Therefore, the problem of improving the bioavailability of crocetin needs to be solved urgently.

[0005] Currently, in order to improve the bioavailability and stability of crocetin, there are already various novel drug delivery systems. In the study by Zhou et al., gum arabic was used as the wall material to prepare crocetin microcapsules, which improved the half-life and stability of crocetin. However, there are drawbacks such as the inability to continuously produce the microcapsules, the non - continuity of the production process of the microcapsule drugs, and relatively low efficiency. In addition, there may be problems with the unstable drug release rate of the microcapsule drugs. Langroodi et al. prepared crocetin PLGA nanoparticles using the W / O / W double emulsion method, which improved the sustained - release effect and pharmacological activity of crocetin. However, this preparation faces problems such as high production costs and preparation complexity. El - Kharrag et al. used FeCl2·4H2O, FeCl3, and an NaOH solution containing dextran as raw materials, prepared magnetic nanoparticles by the coprecipitation method, and then carried out in - situ inclusion to prepare magnetic crocetin nanoparticles. The research showed that the dispersibility and stability of crocetin in water were improved. However, the disadvantage is that the effect of the magnetic nanoparticle drugs is not only affected by the magnetic field strength and direction, and its production and use costs are relatively high, which may limit its wide clinical application. Esposito et al. prepared nanostructured lipid dispersions of crocetin using glyceryl monooleate, sodium cholate, and sodium caseinate as raw materials, which improved the stability and pharmacological activity of crocetin. However, they also face problems such as high production costs, and the storage conditions of the nanostructured lipid dispersions are more demanding.

[0006] Chinese Patent CN109091458A discloses a preparation method of a crocetin microemulsion and its lyophilized powder. Based on the principle of microemulsification, the surfactant added during the preparation process increased the solubility of crocetin in water by 800 times. However, while increasing the solubility, it also affected its stability. In addition, Chinese Patent CN112999162A discloses a crocetin solid dispersion and its preparation method. After dissolving the hydrophilic polymer carrier material and meglumine and mixing them, they are cross - linked with each other to form a carrier polymer, and then mixed and dissolved with crocetin. The solid dispersion is prepared by the solvent evaporation method or the spray - drying method. This method uses the carrier to affect the dissolution and absorption of crocetin. However, organic solvents such as petroleum ether, acetone, dichloromethane, and chloroform used in the preparation process pose certain safety risks. Foreign Patent WO2022 / 025997A1 discloses a trans - crocetin composition and a treatment plan. The trans - crocetin drug and the dosing plan provided therein significantly increase the solubility of crocetin and improve the stability of crocetin. However, the preparation process of the trans - liposome preparation is complex and the production cost is high.

[0007] The present invention aims to study a formulation that can be produced by industrial processes and has a low cost. By preparing crocetin into its sodium salt and then optimizing the formulation and process of the oral formulation of sodium crocetin, the problems that crocetin is insoluble in water and the dissolution of sodium crocetin is extremely unstable are solved. Finally, a crocetin formulation with high bioavailability and its application are provided, effectively prolonging the drug half-life, improving the solubility and solution stability of crocetin, enhancing the bioavailability of the crocetin formulation, and further improving the clinical efficacy in treating cardiovascular and cerebrovascular diseases, nervous system diseases, etc. The preparation process of the present invention has strong operability and low cost, is suitable for industrial scale-up production, and has good clinical application prospects. Summary of the Invention

[0008] Aiming at the problems existing in the prior art, the present invention provides a crocetin formulation with high bioavailability and its application. Specifically, an enteric-coated formulation is prepared with sodium crocetin as the main active ingredient, significantly improving the stability and solubility of the drug, effectively improving the metabolic situation of sodium crocetin in vivo, thereby enhancing the drug bioavailability and absorption rate, effectively preventing or treating cardiovascular and cerebrovascular diseases, and greatly expanding the druggability and clinical value of crocetin.

[0009] To achieve the above object, the technical solution adopted by the present invention is as follows:

[0010] On the one hand, the present invention provides a crocetin formulation with high bioavailability, and the formulation is an oral enteric-coated formulation prepared from crocetin active ingredients; the crocetin active ingredients are selected from at least one of crocetin and sodium crocetin.

[0011] Preferably, in addition to the crocetin active ingredients, the formulation further includes at least one of a pH regulator, a solubilizer, and a lipid material.

[0012] More preferably, the formulation includes crocetin active ingredients, a pH regulator, a solubilizer, and a lipid material.

[0013] Even more preferably, the pH regulator is selected from at least one of sodium bicarbonate, sodium alginate, meglumine, sodium hydroxide, and disodium hydrogen phosphate.

[0014] Even more preferably, the solubilizer is selected from at least one of hydroxypropyl methylcellulose, hydroxypropyl-β-cyclodextrin, polyvinylpyrrolidone, copolyvinylpyrrolidone, hydroxypropyl cellulose, polyethylene glycol, polyethylene glycol-15-hydroxystearate, and vitamin E polyethylene glycol succinate.

[0015] Even more preferably, the lipid material is selected from at least one of glyceryl monostearate, glyceryl behenate, and hydrogenated castor oil.

[0016] Even more preferably, the pH regulator is sodium bicarbonate.

[0017] More preferably, the solubilizer is selected from at least one of hydroxypropyl methylcellulose and vitamin E polyethylene glycol succinate.

[0018] More preferably, the lipid material is glyceryl behenate.

[0019] More preferably, the crocetin preparation comprises, by weight, 1 part of crocetin active ingredient, 1 - 10 parts of pH regulator, 1 - 20 parts of solubilizer and 1 - 20 parts of lipid material.

[0020] More preferably, the crocetin preparation comprises, by weight, 1 part of crocetin active ingredient, 1 - 8 parts of pH regulator, 1 - 15 parts of solubilizer and 1 - 15 parts of lipid material.

[0021] More preferably, the crocetin preparation comprises, by weight, 1 part of crocetin active ingredient, 1 - 5 parts of pH regulator, 1 - 10 parts of solubilizer and 1 - 10 parts of lipid material.

[0022] Preferably, the crocetin preparation further comprises a filler, a glidant, a lubricant and an enteric auxiliary material.

[0023] More preferably, the filler is selected from at least one of lactose, soluble starch, microcrystalline cellulose, dextrin, calcium hydrogen phosphate, mannitol and pregelatinized starch.

[0024] More preferably, the glidant is selected from at least one of colloidal silica and talcum powder.

[0025] More preferably, the lubricant is selected from at least one of magnesium stearate, calcium stearate, colloidal silicon dioxide, hydrogenated vegetable oil, polyethylene glycols and sodium lauryl sulfate.

[0026] More preferably, the weight gain after coating with the enteric auxiliary material is 8% - 20%.

[0027] More preferably, the enteric auxiliary material can be a commercially available product, which can be routinely selected by those skilled in the art to ensure that the preparation does not dissolve in gastric acid but dissolves in the small intestine.

[0028] Preferably, the dosage form of the preparation is selected from any one of enteric - coated tablets, enteric - coated capsules, enteric - coated granules and enteric - coated pellets.

[0029] On the other hand, the present invention also provides a preparation method of the above - mentioned crocetin preparation, comprising the following steps:

[0030] S1. Pretreatment: Mix the crocetin active ingredient and the filler to form mixture 1, and screen mixture 1.

[0031] S2. Granulation: Mix mixture 1, sustained-release agent, and solubilizer to form mixture 2. Dissolve the stabilizer and another solubilizer in water to form a solution. Mix mixture 2 and the solution for wet granulation to form wet granules, and then screen the wet granules.

[0032] S3. Drying and sizing: Dry and screen the wet granules to obtain dry granules.

[0033] S4. Total mixing: Mix the glidant, lubricant, and dry granules to form a total mixture.

[0034] S5. Tabletting: Tablet the total mixture to form a core tablet.

[0035] S6. Aging and coating: Age the core tablet and then perform isolation coating and enteric coating in sequence to obtain the crocetin preparation.

[0036] Preferably, in step S1, the mesh number of the screening is 80 meshes.

[0037] Preferably, in step S2, the mesh number of the screening is 40 meshes.

[0038] Preferably, in step S3, the moisture content of the dried wet granules is less than 2.5%.

[0039] Preferably, in step S3, the mesh number of the screening is 30 meshes.

[0040] Preferably, in step S5, a rotary tablet press is used for tabletting.

[0041] Preferably, in step S5, the hardness of the core tablet is 70 ± 20 N.

[0042] Preferably, in step S6, the specific conditions for aging are: temperature is 60 - 80 °C, and time is 1 - 5 h.

[0043] On the other hand, the present invention also provides the application of the above crocetin preparation in the preparation of drugs for cardiovascular and cerebrovascular diseases and metabolic diseases.

[0044] Preferably, the cardiovascular and cerebrovascular diseases include coronary heart disease, angina pectoris, heart failure, myocardial infarction, hyperlipidemia, and metabolic syndrome.

[0045] Compared with the prior art, the present invention solves the technical problem of poor pharmaceutical properties of crocetin, achieves unexpected technical effects, and significantly improves the bioavailability and in vivo pharmacological activity of oral administration. Specifically, it has the following obvious beneficial effects:

[0046] (1) The present invention first proposes to prepare an enteric-coated preparation with crocetin as the main active ingredient. By optimizing the prescription process, lipid materials, solubilizers, and pH regulators are used as the key components of the crocetin enteric-coated preparation. Both lipid materials and solubilizers can improve the dissolution rate of crocetin. Under the synergistic action of the three with the pH regulator, not only can the dissolution rate of crocetin be significantly improved, but also the solution stability can be enhanced. This prescription process solves the key pharmaceutical property technical problems such as the insolubility of crocetin in water and poor solution stability.

[0047] (2) The present invention first discovers that by preparing sodium crocetin into a sample with high water solubility and administering it through the duodenum, the bioavailability of crocetin can be extremely significantly improved, which is 9.56 times that of the oral gavage group. Further, through oral administration of the enteric-coated preparation of the present invention, the bioavailability can be increased to more than 30 times that of oral administration of crocetin, and the absorption is fast and the half-life is long.

[0048] (3) The pH regulator, lipid materials, and solubilizers in the enteric-coated preparation prepared by the present invention can not only synergistically promote the dissolution of crocetin, but also, through accelerated experiments, no degradation phenomenon is observed under the condition of accelerating for six months, significantly increasing the stability of crocetin. A comparative study on the pharmacodynamic effects of the myocardial infarction-induced heart failure model in rats was carried out using a conventional enteric-coated preparation and the full-prescription enteric-coated preparation of the present invention. It was found that the pharmacokinetics and pharmacodynamics of the full-prescription enteric-coated preparation of the present invention are not only significantly superior to crocetin, but also significantly superior to the prescription samples of the conventional enteric-coated preparation. The full-prescription enteric-coated preparation of the present invention can significantly improve the disease progression of heart failure diseases and improve various indicators of the heart, and can effectively treat or prevent cardiovascular and cerebrovascular diseases, greatly expanding the clinical value and pharmaceutical properties of crocetin.

[0049] (4) Through a large number of literature research and analysis, combined with the inventor's many years of research experience and continuous experimental exploration, the present invention finally successfully solves the pharmaceutical property problems such as the insolubility of crocetin in water and poor solution stability, breaks through the technical difficulties of existing complex preparations such as nano-formulations and liposomes, breaks through the limitation that crocetin preparations can only be administered by injection, and reduces the cost. The present invention discloses a brand-new crocetin enteric-coated preparation suitable for industrial production and with low cost, significantly improving the bioavailability, therapeutic effect, and clinical application value of crocetin. Description of the Drawings

[0050] Figure 1 Results of the cumulative dissolution rate experiment for simulating intestinal fluid.

[0051] Figure 2 Effect of each administration group on NT-proBNP in rats in the pharmacodynamic comparison experiment of the crocetin enteric-coated preparation.

[0052] Figure 3Effect of each administration group on the cardiac index of rats in the pharmacodynamic comparison experiment of crocetin enteric preparations. Detailed implementation mode

[0053] The following non-limiting examples can enable those of ordinary skill in the art to understand the present invention more comprehensively, but do not limit the present invention in any way. The following content is only an exemplary illustration of the scope claimed by the present invention. Those skilled in the art can make various changes and modifications to the invention of the present invention based on the disclosed content, and it should also fall within the scope claimed in this application.

[0054] The present invention will be further described below by way of specific examples. All chemical reagents used in the embodiments of the present invention are obtained through conventional commercial channels unless otherwise specified. Unless otherwise specified, the contents described below are all mass contents. Unless otherwise specified, it is understood that the operations are carried out at room temperature.

[0055] Crocetin (CRA) and sodium crocetin (CRAT) were prepared and provided by Xinglin Traditional Chinese Medicine Technology (Guangzhou) Co., Ltd.

[0056] Example 1 (Pharmacokinetic comparison experiment of different administration routes of sodium crocetin)

[0057] 1. Preparation of test substances

[0058] Preparation of the intravenous administration test solution of CRAT: Prepare a dosing solution with a CRAT concentration of 1.5 mg / mL using 5% glucose, and filter and sterilize it before administration;

[0059] Preparation of the gavage administration and intestinal intubation administration test solutions of CRAT: Dilute and prepare a CRAT suspension with a concentration of 1.5 mg / mL using 0.5% sodium carboxymethylcellulose (CMC-Na);

[0060] Preparation of the gavage administration test solution of CRA: Dilute and prepare a CRA suspension with a concentration of 1.5 mg / mL using 0.5% CMC-Na.

[0061] 2. Experimental animals

[0062] Twenty-four SD rats, SPF grade, weighing 223.3 - 244.9 g, 6 - 7 weeks old, with half males and half females. Provided by Hunan Slake Jingda Experimental Animal Co., Ltd., the quality certificate number of the experimental animals: No430727201101012371, the production license number of the experimental animals: SCXK (Xiang) 2019 - 0004. Breeding conditions: temperature 20 - 26 °C, humidity 40 - 70%, the alternating time of day and night is 12 h / 12 h, and the rats are allowed to eat and drink freely. After one week of adaptive feeding of the rats, the experiment was carried out.

[0063] 3. Grouping and administration

[0064] Twenty-four SFP-grade SD rats that passed quarantine were randomly divided into 4 groups according to gender and body weight, with 3 males and 3 females in each group. A comparison study was conducted with a sodium crocinate intravenous administration group (IV), a crocetin intragastric administration group (CRA), a sodium crocinate intragastric administration group (CRAT), and a sodium crocinate intestinal intubation administration group (DI) (administered from the duodenum). The experimental animals were fasted for 12 h before administration and allowed free access to water. The next day, the corresponding test substances were administered according to the dosing scheme in Table 1. After administration, water was withheld for 2 h and food was withheld for 4 h.

[0065] Table 1. Grouping of test substances and dosing scheme

[0066] Dose group Group Dosing dose / mg / kg Test substance Dosing method Number of animals Intravenous administration group IV 5 CRAT Intravenous injection 3 males and 3 females each Gavage administration group CRA 15 CRA Gavage 3 males and 3 females each Gavage administration group CRAT 15 CRAT Gavage 3 males and 3 females each Intestinal intubation administration group DI 15 CRAT Duodenal administration 3 males and 3 females each

[0067] Note: The above doses are the doses of crocetin.

[0068] 4. Sample collection

[0069] Approximately 0.25 mL of whole blood was collected from the orbital venous plexus (isoflurane inhalation anesthesia) or the jugular vein at the corresponding time points into pre-labeled heparinized blood collection tubes. Among them, blood samples were collected from the animals in each group before administration and at 5 min, 10 min, 20 min, 45 min, 1.5 h, 3 h, 6 h, 12 h, and 24 h after administration. The blood samples were temporarily stored in an ice box and then centrifuged at 4500 rpm for 10 min to separate the plasma. The separated plasma was placed in a labeled centrifuge tube and stored in an ultra-low temperature freezer for later measurement.

[0070] 5. The blood drug concentration of each group was detected using a high-performance liquid chromatograph

[0071] The pharmacokinetic parameters of each group of rats were statistically analyzed. After administration in each dosing group, the main pharmacokinetic parameters of CRA in the plasma are shown in Table 2.

[0072] Table 2. Summary of the main pharmacokinetic parameters of each dosing group (n = 6)

[0073]

[0074] As can be seen from the results in Table 2, the AUC of the CRAT intestinal intubation administration group (0-t) was 77305.3 ± 23733.1 μg / L*h, the AUC of the CRAT intragastric administration group (0-t) was 8086.2 ± 1872.5 μg / L*h. The bioavailability of the intestinal intubation administration group was 9.56 times that of the intragastric administration group and 32.86 times that of the CRA intragastric administration group at the same dose; the t of intravenous injection 1 / 2 was 0.67 ± 0.17 h, the t of intragastric administration 1 / 2 was 9.64 ± 13.09 h, and the t of duodenal administration1 / 2 was 4.65 ± 2.62 h, indicating rapid metabolism after CRA entered the blood, short drug half-life and fast peak time, resulting in C in the intravenous administration group max being higher than that in the intestinal intubation administration group.

[0075] In addition, the water solubility of crocetin after salification increased, and the bioavailability increased by about 4 times compared with crocetin. However, the absolute bioavailability of the CRA and CRAT intragastric administration groups were 1.62% and 5.58% respectively, both of which were extremely low. The absolute bioavailability of the CRAT duodenal administration group was 53.39%, significantly higher than that of the intragastric administration group. It can be seen that although the water solubility of CRAT has increased, there are still problems with insufficient druggability in ordinary gastric-soluble preparations. Through comprehensive analysis, sodium crocetin is more suitable for making enteric-coated preparations clinically. Therefore, the present invention further studies enteric-coated preparations.

[0076] Example 2 (Investigation of pH regulators for enteric-coated crocetin preparations)

[0077] Through long-term formulation research and dissolution rate investigation, it was found that samples prepared from sodium crocetin or crocetin using conventional prescription processes did not meet the qualified formulation dissolution rate requirements of the pharmacopoeia, and the dissolution rate in conventional neutral or slightly acidic media was extremely low. The solubility of crocetin in neutral and acidic environments was extremely low and it could not dissolve. In a neutral to slightly acidic environment, the surface of sodium crocetin particles was easily hydrolyzed into crocetin, resulting in the inability of both sodium crocetin and crocetin samples to dissolve smoothly in conventional media. In view of this, in addition to minimizing the particle size of the raw material drug and increasing the dispersion of the raw material drug, alkaline excipients were introduced into the prescription to increase the pH value of the microenvironment of the raw material drug particles, thereby increasing the dissolution of the raw material drug. First, the influence of pH regulators on enteric-coated preparations was investigated, and the prescription is shown in Table 3.

[0078] Table 3. Prescription of enteric-coated crocetin preparations

[0079]

[0080] Note: " / " indicates not added, the same below.

[0081] The preparation processes of Formulations 1-5 are as follows:

[0082] S1. Pretreatment: Crocetin and lactose were co-crushed with a high-shear pulverizer for 30 s, and the crushed mixture was passed through an 80-mesh sieve;

[0083] S2. Granulation: Place the above mixture, microcrystalline cellulose, and cross-linked povidone into a 5 L wet granulation pot, with a stirring speed of 300 rpm and a cutting speed of 2000 rpm, and mix for 5 min. Dissolve the pH regulator in purified water, pour the above solution into the wet granulation pot for wet granulation, with a stirring speed of 300 rpm and a cutting speed of 2000 rpm, granulate for 3 min, and pass the obtained soft material through a 40-mesh sieve;

[0084] S3. Drying and screening: Place the wet granules in a forced-air drying oven at 60 °C and dry for 2 h, control the moisture content of the granules within 2.5%, and screen the obtained dry granules through a 30-mesh sieve;

[0085] S4. Total mixing: Add colloidal silicon dioxide, magnesium stearate, and the above dry granules into a 5 L barrel of a conical mixer and mix for 5 min to obtain a total mixture.

[0086] S5. Tabletting: Tablet the above total mixture using a rotary tablet press, control the hardness at 100 ± 20 N to obtain the plain tablets.

[0087] Determine the in vitro dissolution rate of Formulations 1 - 5 (by high-performance liquid chromatography), and the test results are as follows;

[0088] In vitro dissolution method: Use pH 6.0 phosphate buffer as the dissolution medium, with a volume of 900 mL, a rotation speed of 50 revolutions per minute, conduct the dissolution experiment using the paddle method, supplement 10 mL of sample each time, determine the dissolution results by high-performance liquid chromatography, and calculate the cumulative dissolution rate. The results are shown in Table 4.

[0089] Table 4. Cumulative dissolution rate

[0090] Time / min 30 60 120 240 Prescription 1 2.13％ 2.21％ 2.30％ 2.41％ Prescription 2 5.44％ 5.49％ 5.55％ 5.62％ Prescription 3 7.14％ 7.40％ 7.54％ 7.49％ Prescription 4 2.31％ 2.28％ 2.30％ 2.21％ Prescription 5 2.82％ 2.81％ 2.84％ 2.83％

[0091] Comparing Formulations 1 - 5, it can be seen that crocetin has basically no dissolution in a medium with pH 6.0. The surface of the API particles is easily acid-hydrolyzed into crocetin, resulting in the basic inability of the API to dissolve, and the final dissolution rate is only about 2%. After adding common alkaline pH regulator excipients (meglumine, sodium bicarbonate, sodium alginate, sodium hydroxide) to the formulation, the solubility is increased, but the degree of increase varies greatly. The results show that when using sodium bicarbonate as the alkaline excipient, the solubilization effect is the most obvious, the effect of meglumine is slightly worse, while sodium hydroxide and sodium alginate have almost no effect.

[0092] Example 3 (Investigation of solubilizers for enteric-coated crocetin preparations)

[0093] By simply adding a pH regulator, the change in dissolution rate under the intestinal pH environment is limited. To further improve the dissolution rate of crocetin sodium, considering that crocetin has long hydrophobic conjugated double bonds and two hydrophilic terminal carboxyl groups, and has surfactant-like properties, the present invention considers introducing a solubilizer into the formulation, screening different solubilizers, analyzing the characteristics of this component and the results of preliminary experiments, comparing available solubilizers, and formulating 10 prescriptions for research, as shown in Table 5 specifically.

[0094] Table 5. Prescriptions of Enteric-coated Crocetin Preparations

[0095]

[0096] The preparation processes of Prescriptions 6-11 are as follows:

[0097] S1. Pretreatment: Co-crush crocetin sodium and lactose with a high-shear pulverizer for 30 s, and pass the crushed mixture through an 80-mesh sieve;

[0098] S2. Granulation: Place the above mixture, microcrystalline cellulose, crospovidone, and solubilizers (hydroxypropyl methylcellulose, hydroxypropyl-β-cyclodextrin, povidone VA064, copovidone, hydroxypropyl cellulose, polyethylene glycol 6000) into a 5-L wet granulation pot, with a stirring speed of 300 rpm, a cutting speed of 2000 rpm, and mix for 5 min. Pour purified water into the wet granulation pot for wet granulation, with a stirring speed of 300 rpm and a cutting speed of 2000 rpm, granulate for 3 min, and pass the obtained soft material through a 40-mesh sieve;

[0099] S3. Drying and sieving: Place the wet granules in a forced-air drying oven at 60 °C for drying for 2 h, control the moisture content of the granules within 2.5%, and sieve the obtained dry granules with a 30-mesh sieve;

[0100] S4. Total mixing: Add colloidal silicon dioxide, magnesium stearate, and the above dry granules into a 5-L conical mixer barrel, mix for 5 min to obtain a total mixture.

[0101] S5. Tabletting: Press the above total mixture with a rotary tabletting machine, control the hardness at 100 ± 20 N to obtain the plain tablets.

[0102] Except for the granulation process, the other processes of Prescriptions 12-15 are the same as those of Prescriptions 6-11. The granulation process is as follows:

[0103] The granulation parameters used are the same as those of Prescriptions 6-11, but the solubilizers (sodium dodecyl sulfate, Tween 80, polyethylene glycol-15-hydroxystearate, vitamin E polyethylene glycol succinate) are first dissolved in purified water respectively and then wet granulation is carried out.

[0104] The in vitro dissolution rates of Formulations 6 - 15 were determined, and the test results are as follows;

[0105] In vitro dissolution method: Using pH 6.0 phosphate buffer as the dissolution medium, the volume of the dissolution medium was 900 mL, the rotation speed was 50 revolutions per minute, the paddle method was used for the dissolution test, 10 mL was sampled and replenished each time, and the dissolution results were determined by high - performance liquid chromatography, and the cumulative dissolution rate was calculated. The results are shown in Table 6.

[0106] Table 6. Cumulative dissolution rate

[0107] Time / min 30 60 120 240 Prescription 6 11.88％ 11.07％ 10.21％ 8.75％ Prescription 7 32.12％ 11.74％ 8.47％ 3.45％ Prescription 8 9.76％ 8.84％ 7.49％ 7.27％ Prescription 9 8.44％ 6.42％ 5.55％ 4.62％ Prescription 10 7.28％ 7.02％ 6.98％ 6.75％ Prescription 11 6.24％ 6.04％ 5.89％ 5.92％ Prescription 12 4.42％ 1.45％ 1.87％ 1.54％ Prescription 13 4.25％ 1.57％ 1.38％ 1.70％ Prescription 14 18.45％ 18.14％ 16.74％ 15.58％ Prescription 15 24.41％ 23.89％ 23.14％ 22.12％

[0108] Comparing Formulations 6 - 15, it can be seen that hydroxypropyl methylcellulose, hydroxypropyl - β - cyclodextrin, polyethylene glycol - 15 - hydroxystearate, and vitamin E polyethylene glycol succinate have a relatively significant solubilizing effect on sodium crocinate as solubilizers. Other excipients have a slight solubilizing effect. Sodium dodecyl sulfate and Tween 80 not only cannot solubilize but will accelerate the precipitation of the active pharmaceutical ingredient. Among them, hydroxypropyl - β - cyclodextrin can significantly improve the dissolution rate of the active pharmaceutical ingredient in a short time, but too high a concentration promotes the violent hydrolysis of the active pharmaceutical ingredient, resulting in a low concentration of the final dissolution solution. Secondly, during the dissolution process of the samples prepared with vitamin E polyethylene glycol succinate and polyethylene glycol - 15 - hydroxystearate, the concentration of the active pharmaceutical ingredient did not show a significant decrease. The two can form dark red emulsion droplets with the dissolved crocic acid, presumably forming crocic acid micelles. For macromolecular compounds, since most molecules have multiple hydrophilic hydroxyl groups, they can reduce the activity of the active pharmaceutical ingredient molecules, so they all show a certain solubilizing effect, among which hydroxypropyl methylcellulose is the most obvious.

[0109] Example 4 (Investigation of lipid materials for enteric - coated crocic acid preparations)

[0110] By adding a pH regulator or a solubilizer to the formulation, the dissolution rate of sodium crocinate in a slightly acidic medium can be increased, but it is still low, and the cumulative dissolution rate fails to exceed 30%. Through the adjustment of the preparation process and formulation, the samples still fail to achieve a cumulative dissolution rate of more than 50%. Comparing the dissolution results of all samples in Examples 2 and 3, it is found that the samples all reach the dissolution end - point within 30 min and partial hydrolysis occurs, which is not conducive to the absorption of the drug in the body. Therefore, through exploratory research in the present invention, it is found that lipid materials are insoluble in the dissolution medium and can provide a micro - space for the dissolution of sodium crocinate. Considering whether this characteristic can be used to improve the dissolution rate and solution stability of sodium crocinate. Therefore, to further improve the dissolution rate and solution stability of crocic acid, the characteristics of each lipid material were analyzed, and through preliminary tests, lipid materials that may achieve the purpose were finally selected for further research on the formulation process. The specific design is shown in Table 7.

[0111] Table 7. Prescription of Crocin Enteric-coated Preparation

[0112]

[0113]

[0114] The preparation process of Prescription 16 - 23 is as follows:

[0115] S1. Pretreatment: Co - grind sodium crocinate and lactose with a high - shear pulverizer for 30 s, and pass the pulverized mixture through an 80 - mesh sieve;

[0116] S2. Granulation: Put the above - mentioned mixture, lipid materials (glyceryl mono - and distearate, glyceryl behenate, hydrogenated castor oil) and hydroxypropyl methylcellulose into a 5 - L wet granulation pot, with a stirring speed of 300 rpm and a cutting speed of 2000 rpm, and mix for 5 min. Dissolve sodium bicarbonate and vitamin E polyethylene glycol succinate in purified water, pour the above solution into the wet granulation pot for wet granulation, with a stirring speed of 300 rpm and a cutting speed of 2000 rpm, granulate for 3 min, and pass the obtained soft material through a 40 - mesh sieve;

[0117] S3. Drying and sieving: Place the wet granules in a forced - air drying oven at 60 °C for 2 h, control the moisture content of the granules within 2.5%, and sieve the obtained dry granules with a 30 - mesh sieve;

[0118] S4. Total mixing: Add colloidal silicon dioxide, magnesium stearate and the above - mentioned dry granules into a 5 - L conical - shaped mixer barrel, mix for 5 min to obtain the total mixture.

[0119] S5. Tabletting: Tablet the above - mentioned total mixture with a rotary tablet press, control the hardness within 70 ± 20 N to obtain the plain tablets.

[0120] Determine the in vitro dissolution rate and content (by high - performance liquid chromatography) of Prescription 16 - 23, and the test results are as follows;

[0121] In vitro dissolution method: Use pH 6.0 phosphate buffer as the dissolution medium, the volume of the dissolution medium is 900 mL, the rotation speed is 50 revolutions per minute, conduct the dissolution experiment by the paddle method, take a 10 - mL sample and supplement the sample each time, determine the dissolution result by high - performance liquid chromatography, and calculate the cumulative dissolution rate. The results are shown in Table 8.

[0122] Table 8. Cumulative Dissolution Rate

[0123]

[0124]

[0125] As can be seen from the dissolution test results in Table 8, a comparison between Formulation 18, 20, 22 and Formulation 16 shows that the lipid materials can improve the dissolution of crocetin, but simply adding lipid materials cannot solve the problem of low dissolution. As can be seen from the previous examples, adding only pH regulators and solubilizers also cannot meet the dissolution requirements. The dissolution results of Formulation 19, 21 and 23 meet the pharmacopoeia requirements and are significantly higher than those of all other formulation process samples, indicating that the lipid materials, pH regulators and solubilizers in the formulation have a synergistic effect when used together, which can significantly improve the solubility and solution stability of sodium crocetin. Whether the bioavailability and efficacy can also be significantly improved remains to be further studied.

[0126] In addition, considering that glyceryl behenate, as a lipophilic material, can provide a stable microenvironment for the dissolution of the active pharmaceutical ingredient, and the lipophilic materials are unevenly distributed in the freshly pressed tablets, resulting in an uneven or unstable microenvironment structure. At the same time, considering that the melting point of glyceryl behenate is 65 - 70 °C, the plain tablets were placed at 70 °C for 2 h for aging to make their distribution more uniform, and the formed microenvironment is smaller, more uniform and more stable. That is, the in vitro dissolution results (dissolution method same as above) of the samples prepared from Formulation 21 after aging are shown in Table 9.

[0127] Table 9. Cumulative dissolution rate of the samples prepared from Formulation 21 after aging

[0128] Time / min 30 60 120 240 Finished product content After ripening of Prescription 21 21.41％ 48.79％ 84.49％ 99.73％ 100.31％

[0129] Example 5

[0130] Prepare a conventional enteric-coated tablet according to Formulation 1 of Example 2. This enteric-coated tablet formulation does not contain pH regulators, solubilizers, or lipid materials.

[0131] First, use Opadry 88A180040-CN to perform a barrier coating on the plain tablets of Formulation 1 in Example 2, with a weight gain of about 3%, and then use Opadry 93O65142-CN to perform an enteric coating, with a weight gain of about 10%, to obtain the enteric-coated tablet.

[0132] Example 6

[0133] Prepare an enteric-coated tablet (containing pH regulator) according to Formulation 3 of Example 2. This enteric-coated tablet formulation does not contain solubilizers or lipid materials.

[0134] First, use Opadry 88A180040-CN to perform a barrier coating on the plain tablets of Formulation 3 in Example 2, with a weight gain of about 3%, and then use Opadry 93O65142-CN to perform an enteric coating, with a weight gain of about 10%, to obtain the enteric-coated tablet.

[0135] Example 7

[0136] Prepare enteric-coated tablets (containing pH regulators and solubilizers) according to Prescription 15 of Example 4. The prescription of this enteric-coated tablet does not contain lipid materials.

[0137] First, use Opadry 88A180040-CN to perform a barrier coating on the core tablets of Prescription 15 of Example 4, with a weight gain of about 3%. Then, use Opadry 93O65142-CN to perform an enteric coating, with a weight gain of about 10%, to obtain the enteric-coated tablets.

[0138] Example 8

[0139] Prepare enteric-coated tablets (full formula) according to Prescription 21 of Example 4. The prescription of this enteric-coated tablet contains pH regulators, solubilizers, and lipid materials.

[0140] Place the core tablets of Prescription 21 of Example 4 at 70 °C for 2 h of aging. After aging, first use Opadry 88A180040-CN to perform a barrier coating on the core tablets, with a weight gain of about 3%. Then, use Opadry 93O65142-CN to perform an enteric coating, with a weight gain of about 10%, to obtain the enteric-coated tablets.

[0141] Example 9

[0142] According to the prescription (full formula) of Prescription 21 of Example 4, use crocetin as the raw material to replace sodium crocetin, and prepare crocetin enteric-coated tablets containing pH regulators, solubilizers, and lipid materials.

[0143] S1. Pretreatment: Co-crush sodium crocetin and lactose with a high-shear pulverizer for 30 s, and pass the crushed mixture through an 80-mesh sieve.

[0144] S2. Granulation: Place the above mixture, glyceryl behenate, and hydroxypropyl methylcellulose in a 5 L wet granulation pot, with a stirring speed of 300 rpm and a cutting speed of 2000 rpm, and mix for 5 min. Dissolve vitamin E polyethylene glycol succinate in purified water, pour the above solution into the wet granulation pot for wet granulation, with a stirring speed of 300 rpm and a cutting speed of 2000 rpm, granulate for 3 min, and pass the obtained soft material through a 40-mesh sieve.

[0145] S3. Drying and sizing: Place the wet granules in a forced-air drying oven at 60 °C for 2 h, control the moisture content of the granules within 2.5%, and size the obtained dry granules with a 30-mesh sieve.

[0146] S4. Total mixing: Add colloidal silicon dioxide, magnesium stearate, and the above dry granules to the 5 L barrel of a conical mixer and mix for 5 min to obtain the total mixture.

[0147] S5. Tableting: Press the above total mixture with a rotary tableting machine, and control the hardness within 70 ± 20 N to obtain the core tablets.

[0148] S6. Baking the tablets: Place the above-mentioned plain tablets in an oven at 70 °C for 2 h to obtain matured plain tablets.

[0149] S7. First, use Opadry 88A180040-CN to perform isolation coating on the above-mentioned matured plain tablets, with a weight gain of approximately 3%, and then use Opadry 93O65142-CN to perform enteric coating, with a weight gain of approximately 10%.

[0150] Example 10

[0151] Prepare enteric-coated pellets according to Prescription 21 (full formula) of Example 4.

[0152] S1. Pretreatment: Co-crush sodium crocinate and lactose with a high-shear pulverizer for 30 s, and pass the crushed mixture through an 80-mesh sieve.

[0153] S2. Granulation: Place the above mixture, glyceryl behenate, and hydroxypropyl methylcellulose in a 5-L wet granulation pot, with a stirring speed of 300 rpm and a cutting speed of 2000 rpm, and mix for 5 min. Pour sodium bicarbonate and vitamin E polyethylene glycol succinate solution into the wet granulation pot for wet granulation, with a stirring speed of 300 rpm and a cutting speed of 2000 rpm, and granulate for 3 min to obtain soft materials.

[0154] S3. Pelletization: Pour the above-prepared soft materials into an extrusion spheronizer, install an extrusion screen with a diameter of 0.8 mm, and turn on the extrusion spheronizer to prepare pellets.

[0155] S4. Drying and sieving: Transfer the above pellets to a fluidized bed, set the inlet air temperature to 60 °C for drying, and after the material is dried, place it in a 10-mesh sieve for sorting, and use a 40-mesh sieve to remove fine powder to obtain dried pellets.

[0156] S5. Coating: Transfer the above pellets to a fluidized bed, spray Opadry 88A180040-CN for isolation coating, with a weight gain of approximately 3%, and then use Opadry 93O65142-CN for enteric coating, with a weight gain of approximately 10%.

[0157] Example 11

[0158] Prepare enteric-coated granules according to Prescription 21 (full formula) of Example 4.

[0159] S1. Pretreatment: Co-crush sodium crocinate and lactose with a high-shear pulverizer for 30 s, and pass the crushed mixture through an 80-mesh sieve.

[0160] S2. Place the above mixture, glyceryl behenate, and hydroxypropyl methylcellulose in a 5 L wet granulation pot, with a stirring speed of 300 rpm, a cutting speed of 2000 rpm, and mix for 5 min. Pour the sodium bicarbonate and vitamin E polyethylene glycol succinate solution into the wet granulation pot for wet granulation, with a stirring speed of 300 rpm and a cutting speed of 2000 rpm, and granulate for 3 min to obtain soft material;

[0161] S3. Granulation: Pour the above-prepared soft material into a rotary granulator, and granulate with a 0.8 mm aperture sieve and a rotation speed of 20 rpm;

[0162] S4. Drying and sizing: Transfer the above granules to a fluidized bed, set the inlet air temperature to 60 °C for drying, and after the material is dried, place it in a 10-mesh sieve for sizing, and use a 40-mesh sieve to remove fine powder to obtain dried granules;

[0163] S5. Coating: Transfer the above granules to a fluidized bed, spray Opadry 88A180040-CN for barrier coating, with a weight gain of about 3%, and then use Opadry 93O65142-CN for enteric coating, with a weight gain of about 10%.

[0164] Results and Discussion

[0165] 1. Simulated intestinal fluid cumulative dissolution rate experiment

[0166] Determine the in vitro dissolution according to the enteric preparation method 1 of the dissolution and release determination method in the 2020 Edition of the Pharmacopoeia of the People's Republic of China 0931. Use 750 mL of 0.1 mol / L hydrochloric acid solution as the dissolution medium, with a rotation speed of 50 revolutions per minute. After 120 minutes, immediately add 250 mL of 0.2 mol / L sodium phosphate solution preheated to 37 °C ± 0.5 °C, with a rotation speed of 50 revolutions per minute. Take a 10 mL sample and replenish the sample each time. Determine the dissolution result by high performance liquid chromatography. Start formal sampling before adding the sodium phosphate solution and calculate the cumulative dissolution rate. The results are shown in Table 10.

[0167] Table 10. Cumulative dissolution rate

[0168] Time / min 0 30 60 120 240 480 Prescription 1 0.23％ 1.54％ 1.56％ 1.62％ 1.66％ 1.67％ Prescription 21 0.33％ 4.34％ 4.25％ 4.75％ 4.74％ 4.77％ Example 5 0.00％ 3.12％ 3.15％ 3.11％ 3.08％ 3.10％ Example 6 0.00％ 7.98％ 8.52％ 8.48％ 8.45％ 8.58％ Example 7 0.00％ 43.47％ 44.51％ 42.12％ 41.08％ 40.72％ Example 8 0.00％ 30.66％ 59.57％ 92.24％ 99.54％ 99.78％ Example 9 0.00％ 23.76％ 42.82％ 65.62％ 74.24％ 75.21％ Example 10 0.00％ 38.43％ 65.48％ 99.48％ 100.20％ 100.42％ Example 11 0.00％ 44.25％ 78.41％ 98.45％ 98.47％ 98.50％

[0169] The above experimental results show that the key ingredient combinations of the present invention were not added simultaneously in the prescription processes of the samples prepared by Prescription 1, Prescription 21, and Examples 5 - 7, and the dissolution degrees were all too low. After the acid tolerance and dissolution degree tests, the crocetin enteric-coated tablets, enteric-coated pellets, and enteric-coated granules of Examples 8 - 11 all met the requirements of the enteric preparation general rules of the Chinese Pharmacopoeia for a relatively high dissolution degree, and the prepared enteric preparations achieved the expected drug release effect, that is, in the specified acidic medium (gastric acid), they maintained an intact form for 2 hours, and the drug was not released or hardly released, while the dissolution degree in intestinal fluid could almost reach 100%, thus achieving the slow-release purpose that the drug would not be released in the stomach but would dissolve rapidly after reaching the intestine, significantly improving the bioavailability of oral administration of crocetin and the concentration of crocetin in the intestine.

[0170] 2. Accelerated stability test

[0171] Respectively take the full-formula enteric-coated tablets prepared in Example 8 and Example 9, place them in a stability chamber at a temperature of 40°C ± 2°C and a relative humidity of 75% ± 5% for 6 months, sample at the 0th, 1st, 3rd, and 6th months respectively, and detect the content and 60-min dissolution degree of three batches of samples. The test results are shown in Table 11.

[0172] Table 11. Results of accelerated stability test

[0173]

[0174] The results of the accelerated stability experiment show that the crocetin sodium enteric preparations and crocetin enteric preparations prepared by the present invention have good stability under accelerated conditions. Through the detection of content determination, related substances, and 60-min dissolution degree, no degradation phenomenon was observed under the condition of accelerating for six months, and there were no obvious changes in the dissolution degree and content, and all the detection indexes met the regulations.

[0175] 3. Pharmacokinetic comparison experiment of crocetin enteric preparations

[0176] To observe the dissolution and absorption behaviors of different prescriptions in vivo, during the prescription screening process, it was verified by combining in-vivo pharmacokinetic experiments on animals.

[0177] 3.1. Test samples

[0178] Ordinary crocetin sodium tablets prepared by Prescription 1 of Example 2, crocetin enteric-coated tablets prepared in Examples 5, 7, 8, and 9, and crocetin sodium enteric-coated pellets prepared in Example 10.

[0179] 3.2. Experimental animals

[0180] Ordinary-grade Beagle dogs, weighing 11.84 - 13.56 kg, 16 - 24 months old, half male and half female. The experimental animal quality certification numbers are: No.110334210100040471, No.370825220100019471. The breeding environment temperature is 16 - 26 °C, and the humidity is 40% - 70%. The day-night light-dark alternation time is 12 h / 12 h. This batch of animals has been quarantined and observed in the designated breeding area of this center for at least 14 days, once a day. After no obvious abnormalities were found, the experiment began.

[0181] 3.3. Grouping and dose design

[0182] According to Table 12, a total of 6 groups were divided, with 4 - 6 animals in each group, half male and half female. The dosing dose was 100 mg / dog, and the dosing method was oral feeding.

[0183] Table 12. Grouping, dosing, and dose design

[0184]

[0185] 3.4. Animal dosing and sample collection

[0186] Before dosing in each group, first moisten the dog's oral cavity and esophagus with 15 mL of purified water, then place the drug at the root of the dog's tongue. After the dog swallows, administer 25 mL of purified water to wash it down. Before and after dosing, collect about 2 mL of whole blood from the saphenous vein of the dog's four limbs at the corresponding time points into pre-labeled heparinized blood collection tubes. Blood samples were collected once from each group of animals before dosing and at 0.5 h, 1 h, 2 h, 3 h, 4 h, 5 h, 6 h, 8 h, 10 h, 12 h, 16 h, and 20 h after dosing.

[0187] 3.5. Test results

[0188] The pharmacokinetic parameters of each group were calculated. After dosing in each dosing group, the specific pharmacokinetic parameters of CRA in plasma are shown in Table 13 and Table 14.

[0189] Table 13. Comparative study results of pharmacokinetic parameters of animals in sample A group of Example 5

[0190]

[0191]

[0192] Table 14. Comparative study results of pharmacokinetic parameters of animals in sample B group of prescription 1 of Example 2

[0193] Parameter <![CDATA[AUC (0-t) > <![CDATA[AUC (0-∞) > <![CDATA[MRT (0-t) > <![CDATA[MRT (0-∞) > <![CDATA[t 1 / 2 > <![CDATA[T max > V CL <![CDATA[C max > Unit μg / L*h μg / L*h h h h h L L / h μg / L B1 5341.9 5343.4 11.655 11.659 1.727 4 18.652 7.486 598.6 B2 3692.7 3692.7 4.123 4.123 0.955 4 14.924 10.832 936.2 B3 2830.2 2832.7 6.834 6.838 1.618 3 7.668 5.854 766.7 B4 1348.6 1348.6 5.458 5.458 0.912 2 9.844 4.479 498.8 mean 3303.4 3304.4 7.02 7.02 1.30 3.25 12.77 7.16 700.1 SD 1668.6 1668.9 3.28 3.29 0.43 0.96 4.96 2.74 192.4

[0194] As can be seen from the experimental results in Table 13 and Table 14, the bioavailability of the enteric-coated tablets of Example 5 with the same formulation and process is three times that of the uncoated plain tablets of Prescription 1 in Example 2. Therefore, to further verify the creativity and clinical value of the whole formula of the present invention, combined with the in vitro dissolution experiment, a pharmacokinetic comparative study on Beagle dogs was carried out according to the samples of Example 7, Example 8, Example 9, Example 10 and Example 5 in Table 12.

[0195] Table 15. Summary of comparative study results of pharmacokinetic parameters of Beagle dogs in each example (n = 6)

[0196]

[0197]

[0198] According to the above results, the AUC (0-t) , t 1 / 2 and C max of the conventional prescription CRAT ordinary tablets of Prescription 1 in Example 2 are lower than those of the sodium crocinate enteric-coated preparations of Examples 7 - 10, indicating that the enteric-coated preparations of the present invention significantly improve the bioavailability of crocetin. In addition, the sodium crocinate enteric-coated tablets of Example 5 without pH regulator, solubilizer and lipid material and the sodium crocinate enteric-coated tablets of Example 7 without lipid material can both improve the bioavailability and half-life of the drug, which is consistent with the conclusion of Example 1. The sodium crocinate enteric-coated tablets of Example 8, the crocetin enteric-coated tablets of Example 9 and the sodium crocinate enteric-coated pellets of Example 10 containing the whole formula all significantly increase the AUC (0-t) and t max . The experimental results show that the bioavailability of the sodium crocinate enteric-coated preparations prepared by the present invention is about 3 - 4 times that of Prescription 1 in Example 2. At the same administration dose, compared with the gastric-soluble preparations, the enteric-coated preparations prepared by the present invention can significantly accelerate the absorption rate, effectively extend the drug half-life and improve the in vivo bioavailability; compared with the conventional enteric-coated preparations, the present invention can significantly improve the oral bioavailability of the poorly soluble crocetin preparations.

[0199] 4. Pharmacodynamic comparison experiment of crocetin enteric-coated preparations

[0200] 4.1. Experimental samples

[0201] Group A: Sodium crocinate raw material, provided by Xinglin Traditional Chinese Medicine Technology (Guangzhou) Co., Ltd.;

[0202] Group B: The sample prepared in Example 5;

[0203] Group C: The whole formula enteric-coated pellets prepared in Example 10.

[0204] 4.2. Experimental animals and methods

[0205] SPF-grade male SD rats, weighing 170.6 - 258.1 g. Except for the 6 rats in the sham operation group, the remaining rats were subjected to coronary artery ligation to establish a chronic heart failure (HFrEF) model with reduced ejection fraction caused by coronary artery ligation. After the operation, they were conventionally raised for 8 weeks. At the 8th week, echocardiography was performed, and animals with an ejection fraction (EF) < 50% were randomly divided into 4 groups: model control group, group A (sodium crocetinate raw material drug), group B (ordinary enteric-coated sodium crocetinate tablets), and group C (enteric-coated sodium crocetinate pellets of the whole formula), with 6 rats in each group; each group was intragastrically administered once a day for 5 consecutive weeks.

[0206] 4.3. Grouping and Dose Design

[0207] According to Table 16, a total of 5 groups were divided, with 6 rats in each group. The dosing doses are shown in Table 16, and all were administered by gavage at a dose of 10 mL / kg body weight.

[0208] Table 16. Grouping, Administration, and Dose Design

[0209] Group Test substance Dose Dosing method Number of animals Sham operation group Blank solvent - Oral administration 6 animals Model control group Blank solvent - Oral administration 6 animals Group A Sodium crocinate raw material drug 11.3 Oral administration 6 animals Group B Ordinary sodium crocinate enteric-coated tablets 11.3 Oral administration 6 animals Group C Whole formula sodium crocinate enteric-coated pellets 11.3 Oral administration 6 animals

[0210] 4.4. Collection and Detection of Animal Samples

[0211] At the 0th week and the 6th week of administration, the ejection fraction (EF) in echocardiography was detected; after the 5th week, 0.5 mL of blood was collected from the sublingual vein of anesthetized HFrEF rats, left to stand at room temperature for about 1 h, centrifuged at 4000 rpm for 10 min to separate the serum, and N-terminal pro-brain natriuretic peptide (NT-proBNP) was detected;

[0212] The animal hearts were taken, rinsed with pre-cooled physiological saline, blotted dry with filter paper, quickly weighed, and then the cardiac index (CI) was calculated.

[0213] 4.5. The detection results are as follows

[0214] NT-proBNP is a marker of myocardial injury. The content of NT-proBNP is directly proportional to the degree of myocardial injury. EF, also known as ejection fraction, can evaluate the systolic and diastolic functions of the human heart and the effective stroke volume per minute. The detection results of NT-proBNP, cardiac index, and EF in each dosing group are shown in Table 17 and Figure 2 , Figure 3 as shown.

[0215] Table 17. Cardiac Function Detection Results of Each Dosing Group

[0216]

[0217] Note: vs sham operation group: ## , p < 0.01; vs model control group: **, p < 0.01;

[0218] As can be seen from Table 17, in the 5th week after drug administration, compared with the sham operation group, the serum NT-proBNP content and cardiac index in the model control group were both increased (p < 0.01), and the serum NT-proBNP and cardiac index in groups A, B, and C showed a downward trend (p < 0.01), indicating that crocetin can improve heart failure to a certain extent, protect cardiomyocytes, reduce myocardial oxidative stress injury, and improve the degree of myocardial fibrosis. In addition, compared with the model control group, the EF% in groups A, B, and C after 5 weeks of drug administration was increased (p < 0.01), indicating that crocetin can improve the cardiac ejection function of heart failure rats, and the enteric-coated pellets in group C have a significant effect on improving the ejection function. From the above results, it can be known that crocetin can be used to treat heart failure diseases, and the enteric-coated crocetin pellets of the whole formula containing pH regulator, solubilizer, and lipid material in group C have a significantly stronger effect than the ordinary sodium crocinate enteric tablets in group B and the raw material drug in group A, and it has the best therapeutic effect on HFrEF. The preparation of the enteric-coated crocetin preparation of the present invention can effectively prevent and treat cardiovascular and cerebrovascular diseases, greatly expanding the clinical value and drugability of crocetin.

[0219] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. An oral enteric-coated preparation of sodium crocetin, characterized in that: The preparation comprises, by weight, 25 parts of sodium crocetin, 52 parts of lactose, 2 parts of sodium bicarbonate, 5 parts of hydroxypropyl methylcellulose, 3 parts of vitamin E polyethylene glycol succinate, 10 parts of glyceryl behenate, 2 parts of colloidal silicon dioxide and 1 part of magnesium stearate; The preparation method of the preparation comprises placing the preparation at 70° C. for aging for 2 hours and isolating and coating the preparation.

2. The oral enteric-coated preparation of sodium crocetin according to claim 1, characterized in that: The dosage form of the preparation is selected from any one of enteric-coated tablets, enteric-coated capsules, enteric-coated granules and enteric-coated pellets.

3. Use of the oral enteric-coated sodium crocetin preparation according to any one of claims 1 to 2 in the preparation of drugs for treating cardiovascular and cerebrovascular diseases and metabolic diseases.

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