WO2009079902A1 - Élaboration et utilisation de composé de curcumine et zinc et sa dispersion solide - Google Patents

Élaboration et utilisation de composé de curcumine et zinc et sa dispersion solide Download PDF

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WO2009079902A1
WO2009079902A1 PCT/CN2008/001086 CN2008001086W WO2009079902A1 WO 2009079902 A1 WO2009079902 A1 WO 2009079902A1 CN 2008001086 W CN2008001086 W CN 2008001086W WO 2009079902 A1 WO2009079902 A1 WO 2009079902A1
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curcumin
group
solid dispersion
zinc
zinc compound
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PCT/CN2008/001086
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French (fr)
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Xueting Mei
Donghui Xu
Shibo Xu
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Sun Yat-Sen University
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Priority to US12/747,800 priority Critical patent/US8759562B2/en
Publication of WO2009079902A1 publication Critical patent/WO2009079902A1/zh

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    • C07—ORGANIC CHEMISTRY
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    • C07F3/00—Compounds containing elements of Groups 2 or 12 of the Periodic Table
    • C07F3/003—Compounds containing elements of Groups 2 or 12 of the Periodic Table without C-Metal linkages
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    • A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
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    • A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
    • A61K47/54—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound
    • A61K47/55—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic compound the modifying agent being also a pharmacologically or therapeutically active agent, i.e. the entire conjugate being a codrug, i.e. a dimer, oligomer or polymer of pharmacologically or therapeutically active compounds
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    • A61K9/16—Agglomerates; Granulates; Microbeadlets ; Microspheres; Pellets; Solid products obtained by spray drying, spray freeze drying, spray congealing,(multiple) emulsion solvent evaporation or extraction
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    • A61K9/1629—Organic macromolecular compounds
    • A61K9/1635—Organic macromolecular compounds obtained by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyvinyl pyrrolidone, poly(meth)acrylates
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    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/04—Drugs for disorders of the alimentary tract or the digestive system for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
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    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/06—Antihyperlipidemics
    • A—HUMAN NECESSITIES
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    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P9/00—Drugs for disorders of the cardiovascular system
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    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
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    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
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    • A61K9/0095—Drinks; Beverages; Syrups; Compositions for reconstitution thereof, e.g. powders or tablets to be dispersed in a glass of water; Veterinary drenches

Definitions

  • the present invention relates to the field of pharmaceutical compositions, and in particular to a curcumin-zinc compound structure, a curcumin-zinc compound and a method for preparing the same, and a pharmaceutical and health care use.
  • Total curcumin [including curcumin, demethoxycurcumin, bisdemethoxycurcumiii], the main active ingredient is curcumiii, with good free radical scavenging Anti-oxidation, anti-mutation, anti-aging, anti-radiation, anti-virus, anti-cancer, anti-cancer, hypolipidemic and cholesterol-lowering, anti-atherosclerosis, treatment of depression and other health and pharmacological effects.
  • curcumin Although curcumin has a wide range of pharmacological effects, it is extremely difficult to dissolve in water, and its bioavailability is extremely low. It is unstable under alkaline conditions or illumination conditions, which greatly limits its application to drug development. Oral administration of 1 g'kg- 1 curcumin to rats, about 75% excreted from feces, and only trace amounts of curcumin in the urine.
  • curcumin content in the blood and bile excretion showed poor intestinal absorption; animal pharmacological experiments were given
  • the curcumin method was intraperitoneally injected, requiring 0.5% dimethyl sulfoxide to dissolve, and the solution was unstable, indicating that curcumin has poor water solubility, poor oral absorption, and low bioavailability, which limits its rational application in human patients.
  • Curcumin is easily degraded in alkaline solution. In phosphate buffer pH 7.4, curcumin degrades rapidly. Its absorption at 426 nm decreases to about 50% after 5 min. After 10 min, only 10% remains. The final solution is Colorless. Atanu Barik has reported the use of curcumin-copper compounds to scavenge free radical reactions.
  • curcumin-copper compounds Due to the poor water solubility of curcumin-copper compounds and the extremely low bioavailability, curcumin-copper compounds are limited in their overall level. At home and abroad, it has been reported that there is no medical research report on the application of curcumin-copper compound, and there is no report on the preparation of solid dispersion of curcumin-zinc compound by water-soluble polyvinylpyrrolidone (PVP) and its medical use.
  • PVP polyvinylpyrrolidone
  • Zinc is an important component of various enzymes in the body. It has the functions of promoting growth and development, improving taste, accelerating wound healing, improving immune function, and anti-oxidation. Growth retardation caused by zinc deficiency, malnutrition, anorexia, and ecstasy And oral ulcers. '
  • Alzheimer's disease, memory loss, hyperlipidemia, cerebral ischemia, arteriosclerosis, thrombosis, blood Small plate aggregation, diabetes, viral infections, etc. are common senile diseases, and in many cases multiple diseases occur simultaneously in the same patient.
  • the existing drugs for treating these diseases generally have problems of single application and unsatisfactory efficacy. Therefore, whether it is possible to research and develop a drug with a good comprehensive therapeutic effect on the above various senile diseases is of great significance for facilitating the prevention and treatment of senile diseases and ensuring the health of the elderly.
  • the object of the present invention is to provide a curative and versatile curcumin-zinc compound and a solid dispersion preparation thereof, which are common in the treatment of existing senile diseases.
  • Another object of the present invention is to provide a process for the preparation of the above curcumin-zinc compound and solid dispersion thereof.
  • a further object of the present invention is to provide a solid dispersion of the above curcumin-zinc compound for use in the treatment of immunocompromised, gastric ulcer, senile dementia, renal failure, hyperlipidemia, arteriosclerosis, diabetes, cerebral ischemia, memory Applications in declining, myocardial ischemia, inflammation, viral infections, tumors, free radical damage, liver damage, depression medications or health supplements.
  • the present invention utilizes two adjacent carbonyl groups in the curcumin molecule to form a complex with zinc acetate, and utilizes polyvinylpyrrolidone (PVP-k30) as a carrier to utilize active hydrogen in the curcumin molecule and a carbonyl group in the PVP molecule.
  • PVP-k30 polyvinylpyrrolidone
  • 0 is hydrogen bonded to form a solid dispersion of curcumin-zinc compound, which greatly improves the water solubility of the curcumin-zinc compound, and exerts a synergistic action between curcumin and Zn+ ions, and has a stronger pharmacological action.
  • the mass ratio of curcumin-zinc to PVP-k30 in the solid dispersion of curcumin-zinc compound is 1: : ⁇ 1 : 28, the preferred ratio is 1:3 ⁇ 1: 18, and the optimum ratio is 1: 5 ⁇ 1 : 16.
  • the curcumin and zinc acetate are dissolved in an organic solvent (such as ethanol, propanol, etc.) in a molar ratio of 1.1 to 1:5, and the curcumin is contained in a nitrogen-protected or oxygen-free environment at 45 to 70 ° C.
  • the organic solution is added dropwise to the organic solution containing zinc acetate, stirred and refluxed for 3 to 5 hours, filtered by cooling, and the precipitate is collected, and the precipitate is washed 4 to 6 times with 5 to 15 ° C absolute ethanol, respectively, and the precipitate is in a vacuum. Dry to obtain curcumin-zinc compound.
  • curcumin-zinc compound and polyvinylpyrrolidone are mixed and dissolved in an ice-free ethanol solution, protected by nitrogen gas, homogenized by a high-pressure homogenizer, filtered, and spray-dried to obtain a solid dispersion of curcumin-zinc compound.
  • Curcumin-zinc compound solid dispersion can improve the water solubility of curcumin-zinc compound, and its curative effect is higher than that of curcumin. It is used in the treatment of immunocompromised, gastric ulcer, senile dementia, renal failure, hyperlipidemia. Important, high cholesterol, arteriosclerosis, diabetes, cerebral ischemia, memory loss, myocardial ischemia, inflammation, viral infections, tumors, free radical damage, liver damage, drugs or health supplements in depression The role.
  • the curcumin-zinc compound solid dispersion can be prepared by a conventional method in combination with a conventional conventional pharmaceutical dressing (such as sodium dodecyl sulfate (SDS), microcrystalline cellulose, stearic acid, etc.);
  • a suitable dosage form such as a tablet, a powder, a granule, a capsule, an ointment, a syrup, an injection, an infusion, or a suppository can be prepared as needed.
  • a conventional conventional pharmaceutical dressing such as sodium dodecyl sulfate (SDS), microcrystalline cellulose, stearic acid, etc.
  • a suitable dosage form such as a tablet, a powder, a granule, a capsule, an ointment, a syrup, an injection, an infusion, or a suppository can be prepared as needed.
  • the curcumin-zinc compound is generally used at a dose of about 1 to 1000 mg per day, and the usual amount for adults is 20 to 900 mg
  • the invention has the following beneficial effects:
  • Curcumin-zinc compound is a new compound, which enhances the activity of zinc-containing antioxidant enzymes and exerts the synergistic effect of curcumin and zinc ions, greatly improving the pharmacological action of the original curcumin.
  • the curcumin-zinc compound solid dispersion improves the water solubility of the curcumin-zinc compound, solves the disadvantages of low bioavailability and poor absorption of curcumin, and the curcumin-zinc compound solid dispersion of the present invention is widely used in treatment.
  • the curative effect is obvious in the senile diseases; it overcomes the problem that the drug application is single and the curative effect is not ideal in the treatment of the existing senile diseases.
  • the preparation method of the invention is simple and has broad application prospects.
  • Zinc acetate dihydrate (abbreviation: zinc acetate), analytically pure, supplied by the Guangzhou Chemical Reagents Wholesale Department.
  • PVP Polyvinylpyrrolidone
  • curcumin-zinc solid dispersion 10.0 g was dissolved in 400 ml of an absolute ethanol solution containing 80.0 g of FVP, protected with nitrogen, homogenized by a high pressure homogenizer, and spray dried to obtain 86.7 g of a curcumin-zinc solid dispersion.
  • mice Healthy NIH mice were selected, weighing 20 ⁇ 2g, 20 males and females. After fasting for 12 hours, the mice were given a curcumin-zinc solid dispersion (200mg/0.5). Ml) solution 0.8ml, continuous observation for 7d, the mouse activity is normal, agile, did not cause death or abnormal reaction, limited to the volume of the test sample can not be increased, so the maximum tolerance test.
  • mice Another 20 healthy NIH mice were selected, weighing 20 ⁇ 2g, half male and half female. After the mice were fasted for 12 hours, the mice were given a curcumin-zinc solid dispersion (200mg/0). 5ml) solution 0. 5ml, once every 3h, 4 times, after continuous observation for 7d, during this period, the mice were allowed to eat and drink freely. During the observation period, the mice were normal and did not cause death or abnormal reaction.
  • curcumin-zinc solid dispersion 200mg/0
  • 5ml solution 0. 5ml, once every 3h, 4 times, after continuous observation for 7d, during this period, the mice were allowed to eat and drink freely. During the observation period, the mice were normal and did not cause death or abnormal reaction.
  • mice were limited to the fact that the volume of the test sample could no longer be increased and LD 5 could not be measured. . Therefore, it was changed to the maximum tolerated dose test.
  • the results were as follows: the maximum tolerated dose of curcumin-zinc solid dispersion in mice was not less than g.kg- 1 body weight, and the maximum tolerance to curcumin-zinc was not less than 5 7g.kg - 1 body weight, during the observation of the test sample, the mice were agile, the fur was smooth, and no death was observed.
  • Curcumin-zinc solid dispersion (curcumin-zinc: PVP, mass ratio 1:6), curcumin solid dispersion (curcumin: PVP, mass ratio 1:6), the preparation method is the same as in Example 2, Provided by the Chinese Medicine and Marine Drug Laboratory of the School of Life Sciences, Sun Yat-sen University. Polyvinylpyrrolidone PVP k30 (referred to as PVP), imported and distributed, supplied by Guangzhou Chemical Reagent Wholesale Department. When used, the sample is dissolved in water and administered by intragastric administration.
  • MDA malondialdehyde
  • LDH lactate dehydrogenase
  • CK creatine kinase
  • the extent of ST segment changes can basically reflect the severity of myocardial ischemia, while the ST segment can be used as a quantitative indicator of ischemic duration.
  • the ST segment After subcutaneous injection of ISO-induced myocardial ischemia in rats, the ST segment was significantly elevated, indicating severe myocardial ischemia injury in the rat myocardium.
  • the curcumin-zinc solid dispersion is superior to the curcumin solid dispersion in reducing the abnormal elevation of the ST segment caused by subcutaneous injection of ISO ( ⁇ ST average displacement amplitude).
  • Curcumin-zinc solid dispersion (curcumin-zinc: PVP, mass ratio 1:10), curcumin solid dispersion (curcumin: PVP, mass ratio 1:10), the preparation method is the same as in Example 2, Provided by the Chinese Medicine and Marine Drug Laboratory of the School of Life Sciences, Sun Yat-sen University. Polyvinylpyrrolidone PVP k30 (referred to as PVP), imported and distributed, supplied by Guangzhou Chemical Reagent Wholesale Department. When used, the sample is dissolved in water and administered by intragastric administration. NIH mice, provided by the Guangdong Medical Laboratory Animal Center.
  • mice 18-22 g were anesthetized with sodium pentobarbital 60 mg.kg for surgery.
  • the mice were fixed in the supine position, the skin was cut in the middle of the neck about 7 mm, the common carotid arteries were separated, and the silk thread of "0" was placed.
  • the bilateral common carotid arteries were clamped with arterial clips for 2 min during ischemia, resulting in brain deficiency. Blood status, end of ischemia, release of arterial clip, recovery of blood reperfusion, short interval 5 min, repeated twice. After blocking the blood flow to the common carotid artery, the incision was sutured. In addition to bleeding at the end, the amount of blood is less than 10% of the total blood.
  • mice X ⁇ S The rats in the blank control group were intragastrically administered with the same amount of PVP, and the other groups of mice were given curcumin, curcumin-zinc content of 90 mg/kg body weight, and administered according to the dose of Table 3, continuous administration 15 day. A memory test was performed 15 days later. The effect of cerebral ischemia-reperfusion injury on brain memory avoidance test in mice X ⁇ S, The incubation period (seconds) of the group dose reaching the black box
  • Curcumin-zinc solid dispersion (curcumin-zinc: PVP, mass ratio 1:8), curcumin solid dispersion (curcumin: PVP, mass ratio 1:8), the preparation method is the same as in Example 2, Provided by the Chinese Medicine and Marine Drug Laboratory of the School of Life Sciences, Sun Yat-sen University. Polyvinylpyrrolidone PVP k30 (referred to as PVP), imported and distributed, supplied by Guangzhou Chemical Reagent Wholesale Department. When used, the sample is dissolved in water and administered by intragastric administration.
  • NIH is a mouse, S.D. is a white rat, provided by the Guangdong Medical Laboratory Animal Center; chicken, purchased from the Zhongshan University Vegetable Market.
  • mice Forty healthy NIH mice were selected, 18 ⁇ 22g, male and female, 10 randomly selected as normal control group, male and female, and the other 30 were intraperitoneally injected with cyclophosphamide 60mg/Kg body weight and randomly divided into 3 groups.
  • Each group consisted of 10 males and a half, namely a blank control group, a curcumin solid dispersion group, and a curcumin-zinc solid dispersion group.
  • the test samples were administered by gavage, and the same amount of PVP was administered to the blank control group and the normal control group.
  • the curcumin group and the curcumin-zinc solid dispersion group were given correspondingly containing curcumin or curcumin-zinc 50 mg/g.
  • the test sample of the corresponding body weight was continuously administered to the test sample for 30 days, and the mice of each group on the 30th day were fasted for one day, but drinking water was provided. On the 31st day, mice in each group were sacrificed. The body weight, thymus and spleen weight of each group were weighed. The thymus index and spleen index were calculated, and the differences between the groups were compared. Third, the experimental results
  • the test substance can significantly increase the thymus index and spleen index of the mouse, and has an immune function. Curcumin zinc solid dispersion is superior to curcumin solid dispersion in increasing thymus index and mouse spleen index in mice.
  • mice Forty mice were randomly divided into 4 groups: normal control group, pathological model group, curcumin group and curcumin-zinc solid dispersion group, with 10 rats in each group.
  • the mice in the normal control group and the pathological model group were given the same amount of PVP solution by intragastric administration.
  • the curcumin solid dispersion group and the curcumin-zinc solid dispersion group were given corresponding weights of curcumin or curcumin-zinc 50 mg Kg.
  • the test sample was prepared, and then the endotoxin was immediately injected with 0.06 g kg (the normal control group was injected with an equal amount of the same amount of PVP solution).
  • mice were harvested from the eyeballs and serum urea nitrogen (BUN) and muscle liver (Cr) concentrations were determined.
  • BUN serum urea nitrogen
  • Cr muscle liver
  • GSH-Px glutathione peroxidase
  • curcumin-zinc solid dispersion can effectively inhibit the increase of serum BUN.Cr value in mice induced by endotoxin, increase the activity of GSH-Px enzyme, and protect the acute renal failure caused by endotoxin. Used, superior to curcumin solid dispersion.
  • the experimental sample curcumin-zinc solid dispersion (curcumin-zinc: PVP, mass ratio of 1: 8), curcumin solid dispersion (curcumin: PVP, mass ratio of 1: 8), the preparation method is the same Example 2, provided by the Chinese Medicine and Marine Drug Laboratory of the School of Life Sciences, Sun Yat-Sen University. Polyvinylpyrrolidone PVP k30 (referred to as PVP), imported and packaged, supplied by Guangzhou Chemical Reagent Wholesale Department. When used, the sample is dissolved in water and administered by gavage.
  • Nffl mice 40 male and female, were randomly divided into 4 groups, 10 in each group, which were normal control group, blank model group, curcumin solid dispersion group and curcumin-zinc solid dispersion group. Group 10 only. The mice in the normal control group and the blank model group were intragastrically administered with the same amount of PVP solution. The curcumin solid dispersion group and the curcumin-zinc solid dispersion group were given 50 mg/kg of curcumin or curcumin-zinc respectively. The corresponding preparation test sample. Each group of NIH mice (except the normal group), each mouse was intraperitoneally injected with 0.45% pentobarbital sodium 0.1 ml / 10 g body weight.
  • the skull is cut longitudinally and the skull is exposed. The skull is exposed. After the bregma point is 2.0 mm, a small hole is drilled with a syringe at a distance of 2.5 mm, and a 2 ⁇ l ⁇ solution is slowly injected into the 5 ⁇ l syringe. The concentration was 7 ⁇ / ⁇ 1, the injection was completed within 30 minutes, the needle was left for 5 minutes, the syringe was removed, and the skin was sutured. The administration was started on the next day, and it was administered once a day for 30 days.
  • Platform test The principle of the platform test is: The bottom of the reaction box is covered with a 36V electric copper grid, and the animals are subjected to electric shock. The normal response is to jump into the insulated platform of the box to avoid noxious stimulation. Most animals may jump to the copper grid again or again, receive an electric shock and quickly jump back to the platform, train for 5 minutes, and record the number of electric shocks per mouse, that is, the number of errors and errors, as a result of academic achievement. The test was repeated 24 hours later, which is the memory retention test. Record the number of animals that were shocked, the latency of the first jump off the platform and the number of errors in 3 minutes. The specific method is: start training the next day after the last administration.
  • the animals were placed in a reaction chamber to acclimate to the environment for 3 min, and then immediately passed through a 36 V AC. Animals are subjected to electric shocks, and their normal response is to jump back to the platform to avoid noxious stimulation.
  • the simultaneous contact of the two feet of the mouse with the copper grid is an electric shock and is considered an error reaction.
  • Most animals may jump to the copper grid again or again, receive an electric shock and quickly jump back to the platform, so trained for 5 minutes (all mice in the same experiment have the same training time), record the number of errors 5 minutes after each mouse, as a learning Results.
  • the test was repeated 24 hours later, that is, the memory retention test. The incubation period of the animal's first jump off the platform and the total number of errors within 3 minutes were recorded, and the frequency of errors in the animals was calculated.
  • senile dementia is the death of brain nerve cells, Nerve fiber tangles, characterized by senile plaques in brain nerve cells and cerebral blood vessels. Injecting ⁇ protein into the hippocampus of mice causes senile plaques in brain nerve cells and cerebral blood vessels, thus forming a dementia model. Curcumin zinc solid dispersion showed significant enhancement of brain memory acquisition and consolidation in Alzheimer's model mice, suggesting an anti-dementia effect, superior to curcumin solid dispersion.
  • Example 9 Pharmacological action on arteriosclerosis
  • Dissection of the left common carotid artery, right external jugular vein take a piece of polyethylene soft plastic tubing (long tube 7.3cm, inner diameter 0.15cm, tube thickness 0.03cm), put a 8cm long medical suture
  • the heparin sodium aqueous solution 50 U/ml was filled with a polyethylene tube at both ends of the plastic tube with a small tube.
  • One end of the polyethylene tube was inserted into the right external jugular vein, and the other end was inserted into the left common carotid artery and ligated and fixed.
  • Healthy SD rats weighing 275 ⁇ 20 g, male, 30 total, were randomly divided into blank model group, curcumin solid dispersion group and curcumin-zinc solid dispersion group, with 10 rats in each group. Rats in the blank model group were given the same amount of PV solution by intragastric administration. The rats in the curcumin solid dispersion group and the curcumin-zinc solid dispersion group were given the corresponding preparations containing curcumin, curcumin-zinc 80 mg/Kg body weight. Product. According to Table 9, the test sample was continuously given for 15 days, and the 16th day was fasted for 12 hours.
  • the carotid artery was dissected and isolated by intraperitoneal injection of pentobarbital sodium solution (30 mg/kg body weight).
  • pentobarbital sodium solution (30 mg/kg body weight).
  • the polyethylene tube was taken for blood, and 3.8% sodium citrate was used as an anticoagulant (anticoagulant: blood volume: 1:9), centrifuged at 800 rpm for 5 min, and the upper layer of milky platelet-rich plasma (PRP) was aspirated, and the remaining portion was again 3000 rpm. After centrifugation for 10 min, the supernatant was taken as platelet-poor plasma (PPP), and PRP was adjusted with PPP to make the platelet count 60-900,000/mm 3 .
  • anticoagulant blood volume: 1:9
  • PRP milky platelet-rich plasma
  • PRP platelet-poor plasma
  • turbidimetry method in order to eliminate the influence of different platelet numbers, PRP and PPP of the same blood sample are used as the zero point and apex of the vertical scale (platelet aggregation rate), PPP is 100%, PRP is 0%, and such relative measurement is The effect of the number of platelets was ruled out.
  • Curcumin-zinc PVP group 80 49. 8 ⁇ 7. 6***### 46. 1 ⁇ 7. 2***## Curcumin group 80 60. 5 ⁇ 6. 7*** 62. 8+ 8. 3*** Compared with the blank control group: ***p ⁇ 0.001; compared with the curcumin group #P ⁇ 0.01, # *P ⁇ 0.001 From the results of rat thrombosis and platelet aggregation experiments, curcumin- Zinc solid dispersion has obvious antithrombotic effect and platelet aggregation, suggesting that it has anti-atherosclerotic effect and is superior to curcumin solid dispersion.
  • Example 10 Pharmacological action of a therapeutic drug delivery model for rat lipid metabolism disorder
  • the healthy SD rats were selected, male, 200 ⁇ 20g, and fed with common words in the experimental environment, observed for 6 days, no abnormalities were found. Then cut the tail to take blood, separate the serum, determine the blood lipid index, eliminate the abnormal blood lipid index of the animal, select the normal blood lipid index, a total of 50, supply food and drinking water, the rats are regularly fed high-fat food every morning. (80% lard + 4% sodium cholate + 4% cholesterol + 12% egg yolk powder) 20ml/kg body weight, continuous administration for 30d. On the 31st day, fasting for 12h, blood was taken from the tail, serum was separated, and the TC content of the triamcinolone was determined.
  • 30 rats above 600 mg / dl were selected and randomly divided into 3 groups, namely high fat pair.
  • the rats in the high-fat control group were given the same amount of PVP solution by intragastric administration.
  • the rats in the curcumin group and the curcumin-zinc solid dispersion group were given corresponding test samples containing curcumin or curcumin-zinc 50 mg/Kg body weight. .
  • the test sample was continuously given for 25 days, and the 26th day was fasted for 12 hours.
  • the rats were taken for blood to measure various blood lipid indexes.
  • curcumin-zinc solid dispersion can be hyperlipidemia, hypercholesterolemia It has obvious therapeutic effects and is superior to curcumin solid dispersion.
  • the experimental sample curcumin-zinc solid dispersion (the erythecin-zinc: PVP, mass ratio of 1: 8), curcumin solid dispersion (curcumin: PVP, mass ratio of 1: 8), the preparation method is the same Example 2, provided by the Chinese Medicine and Marine Drug Laboratory of the School of Life Sciences, Sun Yat-Sen University. Polyvinylpyrrolidone PVP k30 (referred to as PVP), imported and packaged, supplied by Guangzhou Chemical Reagent Wholesale Department. When used, the sample is dissolved in water and administered by gavage.
  • Experimental reagent Alloxan, produced by Sigma Chemical Co.
  • mice provided by the Guangdong Medical Laboratory Animal Center
  • mice Healthy IH mice were selected, males, 18 ⁇ 22g, 50, 10 mice were randomly selected as normal control group, and another 40 were fasted for 12h, and intraperitoneal injection of alloxan 250 mg / g After 6 hours, 2.5kg of glucose solution was administered to make the mice safely pass through the hypoglycemic phase. The mice then provided food and drinking water. After 36 hours, they were fasted for 12 hours. Blood was taken from the orbital vein of the mouse and the intraperitoneal injection of alloxan was measured.
  • the blood glucose level of the mice 30 mice with blood glucose levels greater than 200 mg/dl were selected and randomly divided into 3 groups, a hyperglycemic model group, a curcumin group, and a curcumin-zinc solid dispersion group, 10 in each group.
  • the normal control group and the hyperglycemic model group were given the same amount of PVP solution by intragastric administration.
  • the curcumin solid dispersion group and the curcumin-zinc solid dispersion group were given correspondingly containing curcumin or curcumin-zinc 40 mg/kg. Weight corresponding to the preparation Test sample. The test sample was continuously administered for 12 d.
  • HBeAg enzyme diagnostic kit produced by Shanghai Industrial Kehua Bioengineering Co., Ltd.
  • HbsAg strong positive serum provided by the Department of Clinical Laboratory of the First affiliated Hospital of Guangzhou University of Traditional Chinese Medicine, is a mixture of 20 high-value HBsAg strong positive serum (HBsAg, HBeAg, anti-HBc, HBVDNA are positive), set at -20 °C The refrigerator is kept for use.
  • Preparation of drug-containing serum 24 New Zealand rabbits weighing 2.0 ⁇ 0.2 kg were randomly divided into blank model group, curcumin solid dispersion group and curcumin-zinc solid dispersion group, 8 rats in each group, half male and half female.
  • the blank model group New Zealand rabbits were intragastrically administered with the same amount of PVP solution.
  • the curcumin solid dispersion group and the curcumin-zinc solid dispersion group New Zealand rabbits were respectively given the corresponding preparations containing curcumin or curcumin-zinc 200 mg/kg body weight. sample. Once a day for 8 consecutive days. At 1 hour after the last dose, the heart was blood-sucking. Serum was aseptically separated, and some serum was inactivated by 56 ⁇ 30 min, partially inactivated, and sterilized by 0.45 ⁇ microporous membrane filtration, and stored in a refrigerator at -20 ° C for use.
  • HBsAg-positive serum was doubled with normal saline containing 20% calf serum 1:2, 1:4, 1:8, 1:16...1:16384 Specific dilution, followed by ELISA to detect the PN value of each sample and 1 g L, 2 g / L, 5 g / L HBsAg standard, repeated 3 times, each time do double parallel experiments. Finally, the HBsAg concentration in HBsAg strong positive serum was determined according to the HBsAg concentration-P/N value standard curve, and stored in a refrigerator at -20 °C for use.
  • curcumin-zinc solid dispersion (curcumin-zinc: PVP, mass ratio 1:8: ), curcumin solid dispersion (curcumin: PVP, mass ratio 1:8), preparation method is the same as the example 2, provided by the Chinese Medicine and Marine Drug Laboratory of the School of Life Sciences, Sun Yat-Sen University.
  • Polyvinylpyrrolidone PVP k30 (referred to as PVP), imported and packaged, supplied by Guangzhou Chemical Reagent Wholesale Department. When used, the sample is dissolved in water and administered by gavage. '
  • Rats in the blank control group were given PVP 500mg/K g body weight (excluding the effect of dressing on the gastric mucosa), and the curcumin solid dispersion group and the curcumin-zinc solid dispersion group were given correspondingly containing curcumin or curcumin.
  • the rats were sacrificed by cervical dislocation, the abdominal cavity was ligated to the pylorus and the stomach was taken, and 5 mL of 4% formaldehyde solution was injected into the stomach. After fixation for 30 minutes, the stomach was cut open, and the stomach was turned over to wash away the food residue.
  • the ulcers are rounded or ellipsoidal, measuring the longest and shortest diameters of the ulcer, with their mean as the ulcer index. Statistical comparisons were made between groups, and the ulcer inhibition rate was calculated according to Equation 1-1.
  • the curcumin-zinc solid dispersion has a high serum NO 7j effect, which reduces the blood ET K leveling effect and is superior to the curcumin solid dispersion.
  • curcumin-zinc solid dispersion (curcumin-zinc: PVP, mass ratio 1:8), curcumin solid dispersion (curcumin: PVP, mass ratio 1:8), preparation method same as the example 2, provided by the Chinese Medicine and Marine Drug Laboratory of the School of Life Sciences, Sun Yat-Sen University.
  • Polyvinylpyrrolidone PVP k30 (referred to as PVP), imported and packaged, supplied by Guangzhou Chemical Reagent Wholesale Department. When used, the sample is dissolved in water and administered by gavage.
  • curcumin-zinc solid dispersion (curcumin-zinc: PVP, mass ratio 1:8), curcumin solid dispersion (curcumin: PVP, mass ratio 1:8), preparation method same as the example 2, provided by the Chinese Medicine and Marine Drug Laboratory of the School of Life Sciences, Sun Yat-Sen University.
  • Polyvinylpyrrolidone PVP k30 (referred to as PVP), imported and packaged, supplied by Guangzhou Chemical Reagent Wholesale Department. When used, the sample is dissolved in water and administered by gavage.
  • mice NIH pure mice, provided by the Guangdong Medical Laboratory Animal Center, Ehrlich ascites tumors, transplanted sarcoma (S180) and transplanted liver cancer (HepA) cells, provided by the Cancer Research Center of Sun Yat-sen University, Guangzhou. Effect on transplanted sarcoma S180 in mice
  • mice Thirty mice were inoculated with 0.2 ml of sarcoma S180 cell suspension (1.0 ⁇ 10 7 cells/L) under the right axillary sac of the mice.
  • the inoculation was randomly divided into 3 groups, which were blank control group and curcumin group.
  • the curcumin-zinc solid dispersion group 10 mice in each group, the blank control group was intragastrically administered with the same amount of capsules, and the curcumin group and the curcumin-zinc solid dispersion group were respectively given with curcumin or curcumin- Zinc 60 mg Kg body weight corresponding test sample.
  • the tumor inhibition rate (%) (1 - mean tumor weight of the administration group / mean tumor weight of the control group) x l00%.
  • mice Thirty mice were injected into the mice intraperitoneally with 0.2 ml of HepA cell suspension (L0X10 7 cells/ml). The cells were randomly divided into 3 groups on the next day, which were blank control group, curcumin solid dispersion group and curcumin.
  • - Zinc solid dispersion group 10 mice per group, blank control mice were intragastrically administered with equal amounts of PVP solution, curcumin solid dispersion group, curcumin-zinc solid dispersion group were respectively given with curcumin or turmeric Sucrose-zinc 60 mg/Kg body weight corresponding test sample.
  • Life extension rate (average survival days in the experimental group / average survival days in the control group - 1) ⁇ 00%.
  • mice Thirty mice were enrolled in the mice. The mice were intraperitoneally inoculated with 0.2 ml of Ehrlich ascites tumor (1.0 ⁇ 10 7 cells/L). The next day, the rats were randomly divided into 3 groups: blank control group, curcumin solid dispersion group and curcumin.
  • Zinc solid dispersion group 10 mice in each group, blank control group mice were intragastrically administered with equal amount of PVP solution, curcumin solid dispersion group, curcumin-zinc solid dispersion group correspondingly given curcumin or turmeric Sucrose-zinc 60 mg g body weight corresponding preparation test sample.
  • Life extension rate (average survival days in the experimental group / average survival days in the control group - 1) ⁇ 100%.
  • Inhibition of mouse transplanted sarcoma S180 ( ⁇ s, n 10)
  • Curcumin-zinc group 60 19.4+5.2*## 36.6 Curcumin group 60 17.9+5.2 26.0 Compared with the blank control group, *P ⁇ 0.01; compared with the curcumin group, ⁇ ⁇ . ⁇ compared with the blank control group, The curcumin-zinc solid dispersion has a significant inhibitory effect on s 180 sarcoma, and the survival time of Ehrlich ascites mice and HepA mice is superior to that of curcumin solid dispersion.
  • Example 16 Protective effect on liver injury induced by carbon tetrachloride in rats
  • Curcumin-zinc solid dispersion Curcumin-zinc: PVP, mass ratio 1:6)
  • curcumin solid dispersion curcumin: PVP, mass ratio 1:6
  • preparation method same as the example 2 provided by the Chinese Medicine and Marine Drug Laboratory of the School of Life Sciences, Sun Yat-Sen University.
  • Polyvinylpyrrolidone PVP k30 (referred to as PVP), imported and packaged, supplied by Guangzhou Chemical Reagent Wholesale Department. When used, the sample is dissolved in water and administered by gavage.
  • Experimental animals SD pure white rats, provided by Guangdong Medical Laboratory Animal Center;
  • the rats in the normal control group were intraperitoneally injected with peanut oil 5ml/k g , and the other five groups were intraperitoneally injected with 25% CCl4-arachi oil 5nJ/kg for 10 days, and fasted on the 11th day.
  • the water could not be forbidden for 24 hours.
  • the blood was centrifuged at 300Qrpm ⁇ 10min to determine the contents of ALT, AST and Alb in the serum.
  • the curcumin-zinc solid dispersion has anti-liver damage effect and inhibits serum
  • ALT and AST are superior to the solid dispersion of curcumin.
  • curcumin-zinc solid dispersion (curcumin-zinc: PVP, mass ratio 1:8), curcumin solid dispersion (curcumin: PVP, mass ratio 1:8), preparation method same as the example 2, provided by the Chinese Medicine and Marine Drug Laboratory of the School of Life Sciences, Sun Yat-Sen University.
  • Polyvinylpyrrolidone PVP k30 (referred to as
  • mice Thirty male Kunming mice, weighing 18 ⁇ 25g, were randomly divided into 3 groups, 10 in each group, which were normal control group, blank control group, curcumin solid dispersion group and curcumin-zinc solid dispersion group.
  • the blank control group was given an equal amount of PVP solution, and administered once a day on the morning, once a day for 8 days.
  • the mice were placed in a glass dish (30 cm high, 18 cm in diameter) with a water depth of 10 cm. After the mice were swimming 2 ⁇ , the observation was started immediately, and the observation was continued for 4 min. The mice were accumulated in the water for 4 minutes.
  • mice Male Kunming mice, weighing 18 ⁇ 25g, total 30, were randomly divided into 3 groups, 10 in each group, which were blank control group, curcumin solid dispersion group and curcumin-zinc solid dispersion group.
  • the blank control group was given the same amount of PVP solution, and was administered by intragastric administration once a day, once a day for 8 days.
  • the mice were fixed at 2 cm at the end of the mouse on a horizontal surface, and the head was placed upside down. The head was 30 cm away from the table. The surrounding eyes were separated by a plate. The mice were first adapted to 2 min, and the mice were recorded. No moving time within 6 minutes.
  • the immobile state of the mouse in the forced swimming model also reflects the desperate behavior of the animal.
  • the curcumin solid dispersion group and the curcumin-zinc solid dispersion group can significantly shorten the forced swimming time in mice, which is significantly different from the blank control group, suggesting that the curcumin solid dispersion, curcumin-zinc solid dispersion It has an antidepressant effect, while the curcumin-zinc solid dispersion is superior to the curcumin solid dispersion.
  • the immobile state of the mouse in the tail-tail model reflects the desperate behavior of the animal.
  • the immobile state of the mouse in the forced swimming model also reflects the desperate behavior of the animal.
  • the curcumin solid dispersion group and the curcumin-zinc solid dispersion group can significantly shorten the forced swimming time in mice, which is significantly different from the blank control group, suggesting that the curcumin solid dispersion, curcumin-zinc solid dispersion It has an antidepressant effect, while the curcumin-zinc solid dispersion is superior to the curcumin solid dispersion.

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Description

姜黄素-锌化合物及其固体分散体的制备方法与应用 技术领域
本发明涉及医药组合物领域， 具体涉及姜黄素-锌化合物结构、 姜黄素 -锌化 合物及其面体分散体制备方法与医药及保健用途。
背景技术
总姜黄素 [包括姜黄素 (curcumin)、 去甲氧基姜黄素 (demethoxycurcumin)、 双 去甲氧基姜黄素 (bisdemethoxycurcumiii)]， 主要活性成分是姜黄素 (curcumiii)， 具 有很好的清除自由基、抗氧化、抗突变、延缓衰老、防辐射、抗病毒、防癌抗癌、 降血脂和降胆固醇、 抗动脉粥样硬化、 治疗抑郁等多种保健及药理作用。
虽然姜黄素具有广泛的药理学作用， 但极难溶于水， 同时生物利用度极低， 在碱性条件或光照条件下不稳定， 大大局限了其应用于药物开发。 给大鼠口服 1 g'kg—1姜黄素， 约 75%自粪便排出， 而尿中只有痕量的姜黄素， 测定血中姜黄素 含量和胆汁排泄结果表明肠吸收不好；动物药理实验给予姜黄素的方式采用腹腔 注射， 需 0.5%二甲基亚砜溶解， 而且溶液不稳定， 表明由于姜黄素的水溶性差、 口服吸收差、生物利用度低， 限制其合理应用在人体患者。姜黄素在碱性溶液中 容易降解， 在 pH7.4的磷酸盐缓冲液中， 姜黄素很快降解， 其 426nm的吸收值 5min后降低到约 50%， lOmin后只剩 10%， 最后溶液是无色的。 Atanu Barik有 报道应用姜黄素 -铜化合物用于清除自由基反应， 由于姜黄素 -铜化合物水溶性极 差、 同时生物利用度极低， 局限姜黄素 -铜化合物在整体水平发挥作用。 国内外 查新表明目前并无应用姜黄素-铜化合物的医药研究报道， 更无应用水溶性聚乙 烯吡咯垸酮 (PVP)于姜黄素-锌化合物制备固体分散体及其医药用途的报道。
由于生活水平的提高和医疗保健的不断进步，人均寿命越来越长，使世界人 口老龄化的趋势越来越明显。据中国老年协会专家指出，我国老年人口迅速增长，
1990年以来， 我国老年人口以平均每年 3.32%的速度增长， 到 2006年底， 全国 60岁以上老年人口总数已达到 1.2亿， 约占人中总数的 9.5%。 我国已成为 "老 年型"国家，随着老年人口增加，对老年性疾病的防治问题也越来越突出。因此， 研制开发新的抗衰老及防治常见老年性疾病的新药，已成为当今医药学界的一个 研究热点。
锌为体内多种酶的重要组成成份， 具有促进生长发育， 改善味觉， 加速伤口 愈合、 提高免疫功能、 抗氧化等作用，缺锌可引起的生长发育迟缓， 营养不良， 厌食症、 异食癖以及口腔溃疡。 '
老年性痴呆症、 记忆衰退、 高血脂症、 脑缺血、 动脉硬化、 血栓形成、 血 小板聚集、糖尿病、病毒性传染病等是常见的老年性疾病，而且许多情况下是多 种病症同时出现于同一病人身上。但是，现有的治疗这些疾病的药物普通存在着 适用症单一、疗效不尽理想的问题。 因此， 能否研究开发一种对上述多种老年性 疾病具有良好综合治疗效果的药物，这对方便老年性疾病的防治、保证老年人健 康将具有十分重要的意义。
发明内容
本发明的目的是针对现有老年性疾病治疗中普遍存在药物适用症单一、疗效 不尽理想的问题， 提供一种疗效显著、 用途广泛的姜黄素-锌化合物及其固体分 散体制剂。
本发明的另一个目的是提供上述姜黄素-锌化合物及其固体分散体的制备方 法。
本发明的进一步目的是提供上述姜黄素-锌化合物固体分散体在制备用于治 疗免疫功能低下、 胃溃疡、老年性痴呆症、肾衰、高血脂症、动脉硬化、糖尿病、 脑缺血、 记忆衰退、 心肌缺血、 炎症、 病毒感染、 肿瘤、 自由基损伤、 肝损伤、 抑郁症的药物或保健品中的应用。
为了实现上述目的， 本发明采用如下技术方案：
本发明利用姜黄素'分子中相邻的两个羰基与乙酸锌形成络.合物，并利用聚乙 烯吡咯烷酮 (PVP-k30)作为载体，利用姜黄素分子中的活泼氢与 PVP分子中的 羰基 0以氢键结合在一起， 形成姜黄素-锌化合物固体分散体， 大大提髙姜黄素- 锌化合物的水溶性， 发挥姜黄素与 Zn+离子的协同作用， 具有更强的药理作用。
姜黄素-锌化合物固体分散体中姜黄素 -锌与 PVP-k30的质量比为 1 : ：!〜 1 : 28, 优选比例为 1: 3〜1： 18， 最佳比例为 1 : 5〜1： 16。
将姜黄素和乙酸锌按摩尔比 1.· 1〜1： 5分别溶于有机溶剂 （如乙醇、 丙醇 等）， 于氮气保护或在无氧 45〜70°C环境下， 将含姜黄素的有机溶液滴加到含乙 酸锌的有机溶液中， 搅拌回流 3〜5小时， 冷却过滤， 收集沉淀物， 分别用 5〜 15°C无水乙醇洗涤沉淀物 4〜6次，沉淀物在真空干燥，得到姜黄素一锌化合物。 将姜黄素-锌化合物和聚乙烯吡咯烷酮按比例混合溶解于无氷乙醇溶液中， 通氮 气保护， 经高压均质机均质， 过滤， 滤液喷雾干燥得姜黄素 -锌化合物固体分散 体。
姜黄素-锌化合物固体分散体可提高姜黄素-锌化合物的水溶性， 疗效高于姜 黄素的药理作用，在制备用于治疗免疫功能低下、胃溃疡、老年性痴呆症、肾衰、 高血脂症、高胆固醇症、 动脉硬化、 糖尿病、脑缺血、 记忆衰退、 心肌缺血、 炎 症、病毒感染、 肿瘤、 自由基损伤、肝损伤、抑郁症的药物或保健品中具有重要 的作用。
姜黄素-锌化合物固体分散体可与普通常规的药用敷料配合（如十二'烷基硫 酸钠 (SDS)、微晶纤维素、硬酯酸续等)， 经常规方法而制得； 可根据需要制成适 当的剂型， 如片剂、粉剂、颗粒剂、 胶囊剂、 膏剂、 糖浆剂、 针剂、输液、 或栓 剂等。通常以口服方式使用， 当然也可以采用皮肤外用给药等其它给药方式； 姜 黄素 -锌化合物每天使用剂量一般为约 1〜1000毫克， 成年人常用量为每天 20〜 900毫克， 最常用剂量为 50〜700毫克。 每天一次或分数次使用。
本发明与现有技术相比， 具有如下有益效果：
1.姜黄素-锌化合物为新化合物，提高提内含锌抗氧化酶活性，发挥姜黄素与锌离 子的协同作用， 大大提高了原有姜黄素的药理作用。
2.姜黄素 -锌化合物固体分散体提高姜黄素 -锌化合物水溶性， 解决了姜黄素生物 利用度低、 吸收差等缺点， 本发明的姜黄素 -锌化合物固体分散体用途广泛， 在 治疗各种老年性疾病中疗效显著；克服了现有老年性疾病治疗中普遍存在药物适 用症单一、 疗效不尽理想的问题。
3.本发明的制备方法简单， 应用前景广阔。
具体实施方式
实施例 1 姜黄素-锌化合物的制备
姜黄素， 99.6%纯度， 由中山大学生命科学学院中药与海洋药物实验室提供。 二水合乙酸锌 （简称： 乙酸锌）， 分析纯， 由广州化学试剂批发部提供。
将 4.8克姜黄素溶于 300ml无水乙醇中， 2.9克乙酸锌溶于 150ml无水乙醇 中， 将乙酸锌乙醇溶液滴加到姜黄素乙醇溶液中， 边滴加边搅拌， 回流加热 3小 时， 至沉淀完全， 抽滤收集沉淀物， 重新用 300ml 5Ό无水乙醇震荡悬浮沉淀物， 5000rpm离心 5min， 收集沉淀物，再用 300ml无水乙醇震荡悬浮， 5000rpm离心 5min， 得 4.9克姜黄素 -锌化合物。
实施例 2 姜黄素固体分散体、 姜黄素-锌化合物固体分散体的制备
姜黄素（Curcumin), 姜黄素 -锌化合物，纯度为 99.6%， 由中山大学生命科 学学院中药与海洋药物实验室提供。 聚乙烯吡咯綜酮 PVP-k30(简称 PVP)， 进口 分装， 由广州化学试剂批发部提供。
将 10.0克姜黄素溶于 400ml含有 80.0克 FVP的无水乙醇溶液中，通氮气保 护， 经高压均质机均质， 喷雾干燥得姜黄素-锌固体分散体 86.7克。
将 10.0克姜黄素-锌化合物溶于 400ml含有 80.0克 PVP的无水乙醇溶液中， 通氮气保护， 经高压均质机均质， 喷雾干燥得姜黄素-锌固体分散体 86.2克。 实施例 3 对小鼠急性毒性试验 实验样品： 姜黄素-锌固体分散体（质量比姜黄素 -锌： PVP: SDS=1 : 6)， 由中山大学生命科学学院中药与海洋药物实验室提供。
实验动物： MH系小鼠 18-22g， 60只， 雌雄各半， 由广东省医学实验动物中心 提供。
方法与结果：
选取健康 NIH系小鼠， 体重 20 ± 2g， 20只， 雌雄各半， 对小鼠禁食不禁水 12h后， 对应每组小鼠 1次灌胃含有姜黄素-锌固体分散体（200mg/0.5ml)溶液 0.8ml, 连续观察 7d， 小鼠活动正常、 敏捷， 未引起死亡或异常反应， 限于给受 试样品体积不能再增大， 故进行最大耐受量试验。
另选择健康 NIH系小鼠 20只，体重 20 ± 2g，雌雄各半，对小鼠禁食不禁水 12h后， 对应每组小鼠 1 次灌胃装姜黄素-锌固体分散体（200mg/0. 5ml)溶液 0. 5ml, 每隔 3h灌胃一次， 共 4次， 此后连续观察 7d， 此期间让小鼠自由进食 和饮水， 观察期间小鼠活动正常， 未引起死亡或异常反应。
小鼠限于给受试样品体积不能再增大不能测出 LD5。。故改作最大耐受量试验， 结果如下：小鼠灌服姜黄素 -锌固体分散体最大耐受量不少于 g.kg—1体重，相当 给予姜黄素-锌最大耐受量不少于 5. 7g.kg— 1体重， 在给受试样品观察期间， 小鼠 活动敏捷， 毛皮光滑， 未见死亡。
由此可知， 姜黄素-锌固体分散体（质量比姜黄素 -锌： PVP=1 : 6) 的最大 耐受量不少于 40g.kg— 1体重， 相当给予姜黄素-锌最大耐受量不少于 5. 7g，kg—'体 重， 属于实际无毒级， 服用安全。 实施例 4 对心肌缺血的保护作用及清除自由基作用
一、 实验样品
姜黄素-锌固体分散体 (姜黄素 -锌： PVP， 质量比为 1 : 6)， 姜黄素固体分散 体 (姜黄素： PVP， 质量比为 1 : 6)， 制备方法同实施例 2， 由中山大学生命科学 学院中药与海洋药物实验室提供。 聚乙烯吡咯垸酮 PVP k30(简称 PVP)， 进口分 装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌胃给药。
SD大鼠，广东省医学实验动物中心提供； MS-302多媒体生物信号记录分析 系统（广东生命科学学院）； WZ-50D型微量注射泵（浙江医科大学医学仪器厂） 二、 实验方法
试验前测定 SD大鼠心电图和胸导联，弃去 T波、 ST段位移有异常和心律失 常者。 选取合格大鼠， 体重 210.1±20.0g， 雌雄兼用， 每组 8只大鼠， 随机分为 空白对照组、姜黄素固体分散体组、姜黄素-锌 固体分散体组， 空白对照组大鼠 灌胃给予等量 PVP溶液， 其佘各组大鼠分别对应灌胃给予姜黄素、 姜黄素 -锌含 量为 80mg Kg体重， 30min后乙醚麻醉，记录心电图后颈背部皮下注射异丙肾上 腺素 (Isoproterenol, ISO) 2mg kg， 30min后再记录心电图 1次， 24h和 48h后再 一次灌胃受试品 30min后，各组按上述方法重复给 ISO并记录心电图。 72h后再 一次灌胃受试品 30min后， 大鼠眼眶取血并离心分离血清， 处死大鼠，剪取心尖 以 4Ό生理盐水制成 10%的心肌组织勾浆备用。
对∑ST段位移幅度的影响：
测量第一次给 ISO前、 第一、 二、 三次皮下注射 ISO后 30min各鼠 ST值， 计算 EST的 ιην数均值作为心肌损伤程度的指标并做 t检验。对血清中 CK、LDH 和 MDA的影响：
用试剂盒测血清中的丙二醛 (MDA)、 乳酸脱氢酶（LDH)和肌酸激酶 (CK) 的变化。
三、 结果
1 对∑ST段的影响
ST段的变化幅度可基本反映心肌缺血的严重程度， 而∑ST段可作为缺血程 度的定量指标。 皮下注射 ISO诱发大鼠心肌缺血后， ST段明显抬高， 显示大鼠 心肌有严重的心肌缺血损伤。 姜黄素-锌 固体分散体对能降低皮下注射 ISO 引 起的 ST段的异常抬高程度 (∑ST平均位移幅度）， 优于姜黄素固体分散体。 如 表 1。 表 1 对异丙肾上腺素致大鼠心肌损伤模型中∑ST的影响 (x±s, n=8)
Group ∑ST/mv
30min 24h 48h 空白对照组 1. 40±0. 28 1. 25±0. 28 1. 04+0. 23 姜黄素 -锌组 0. 92±0. 27*## 0. 81+0. 16* 0. 76±0. 22* 姜黄素组 1. 26+0. 15 1. 05±0. 19 0. 89±0. 20
¾空白对照组相比， *Ρ<0. 05， *ΦΡ<0. 01; 与姜黄素组比较， # Ρ<0.05， 对血清中 CK：、 LDH和 MDA的影响：
大鼠皮下注射 ISO后， 血清中的 CK、 LDH和 MDA均明显上升， 显示由 于心肌组织的损伤而使心肌细胞中的 CK和 LDH漏出到血液中， 血清中的脂质 过氧化物 (Lipid per-oxidation, LPO)大量增加。 结果显示受试化合物可明显抑制 CK和 LDH从心肌细胞内向血清中漏出， 显著抑制 Iso致大鼠血清中 MDA的异 常升高， 对自由基损伤具有保护作用。 结果表明： 姜黄素-锌固体分散体对 ISO致心肌缺血、 自由基损伤具有保护 作用,优于姜黄素固体分散体。
对 Iso致心肌损伤大鼠血清的影响（x±s, n=8)
组别 CK LDH MDA
(U/L) (U/L) (nmol/ml) 空白对照组 382. 1+30. 7 337. 4±35. 2 58. 4+5. 8
姜黄素 -锌组 289. 3+25. 2***## 240. 6±21. 9***麵 28. 6+6. 2***## 姜黄素组 340. 5+20. 9** 298. 9+26. 4*** 39. 1+5. 2*** 与空白对照组相比， ***Ρ<0.001 ; 与姜黄素组比较， # Ρ<0.05，
画 Ρ<0· 001
实施例 5 对脑缺血、 记忆衰退模型的保护作用
一、 实验材料
姜黄素-锌固体分散体 (姜黄素 -锌： PVP, 质量比为 1 : 10)， 姜黄素固体分散 体 (姜黄素： PVP, 质量比为 1 : 10)， 制备方法同实施例 2, 由中山大学生命科学 学院中药与海洋药物实验室提供。 聚乙烯吡咯烧酮 PVP k30(简称 PVP), 进口分 装， 由广州化学试剂批发部提供。使用时样品溶于水中分别用灌胃给药。 NIH系 小鼠， 由广东省医学实验动物中心提供。
剂量实验方法与结果：
NIH小鼠， 18-22g， 以戊巴比妥钠 60mg.kg— 麻醉， 进行手术。 将小鼠仰 卧位固定， 颈部正中切开皮肤约 7mm， 分离两侧颈总动脉， 并套以 " 0"号丝线， 缺血时用动脉夹夹闭双侧颈总动脉 2min， 造成大脑缺血状态， 缺血结束， 松开 动脉夹， 恢复血液再灌注， 短时间歇 5min， 如此反复二次。 阻断颈总动脉血流 后， 缝合切口。加上尾端放血， 放血量为总血量的 10%以下。术毕从腹腔内注入 适量的生理盐水以补充血容量。 假手术组麻醉后分离双侧颈总动脉但不阻断血 流， 亦不放血。
双侧颈总动脉完全阻断后， 首先出 ¾'惊厥， 体温降低， 呼吸减慢， 最后翻正 反射消失。实验动物以体温降低和翻正反射消失为缺血阳性指标，无典型缺血指 征者弃之不用。在缺血过程中、 或手术后均会有动物死亡。恢复血液灌注后， 动 物翻正反射逐渐恢复， 呼吸加快， 3-5h基本恢复正常活动。手术后的动物隨机分 为 4组： 假手术组、 姜黄素固体分散体组、 姜黄素 -锌固体分散体组、 空白对照 组， 每组 10只。 每天给药一次， 空白对照组小鼠灌胃给予等量 PVP， 其余各组 小鼠分别对应给予姜黄素、 姜黄素-锌含量为 90mg/Kg体重， 按表 3剂量灌胃， 连续给药 15天。 15天后进行记忆试验。 对脑缺血再灌注损伤小鼠脑记忆避暗实验的作用 X ± S,
Figure imgf000008_0001
组别 剂量 到达暗箱的潜伏期 (秒） 错误次数
(mg-kg"1) 学 习 记忆 (n) 假手术组 ― 37. 6 土 15. 2** 58. 3土 20. 5*** 3. 5 ± 2. 0*** 空白对照组 —一 17. 2 ± 10. 2 28. 5 ± 12. 5 7. 9 ± 1. 6 姜黄素 -锌组 90 33. 8 ± 6. 2**# 49. 3 ± 14. 7**# 4. 1 ± 2. 3**# 姜黄素组 90 26. 3 ± 5. 8 37. 4 ± 9. 4 6. 3 ± 2. 1 与空白对照组相比， <0.05, * <0.01，^Ρ<0.001 ; 与姜黄素组比较， #P<0.05 从表 3的结果显示， 假手术组与空白组相比， 进入暗箱的潜伏期明显延长， 受电击次数明显减少 ( 0. 01)，说明脑缺血再灌注对小鼠的学习记忆功能造成了 明显损害。 与空白对照组相比， 姜黄素 -锌固体分散体能够明显延长进入暗箱的 潜伏期， 同时能显著降低错误次数， 对脑缺血、记忆衰退具有保护作用。姜黄素 -锌固体分散体对脑缺血、 记忆衰退具有保护作用，优于姜黄素固体分散体。 实施例 6 对机体免疫功能的影响
一、 实验材料
姜黄素-锌固体分散体 (姜黄素 -锌： PVP, 质量比为 1: 8), 姜黄素固体分散 体 (姜黄素： PVP， 质量比为 1 : 8)， 制备方法同实施例 2， 由中山大学生命科学 学院中药与海洋药物实验室提供。 聚乙烯毗咯烷酮 PVP k30(简称 PVP)， 进口分 装， 由广州化学试剂批发部提供。使用时样品溶于水中分别用灌胃给药。 NIH系 小白鼠， S.D.系大白鼠， 由广东省医学实验动物中心提供； 家鸡， 由中山大学菜 市场购得。
二、 实验方法
对小鼠胸腺和脾脏的作用
选择健康 NIH系小鼠 40只， 18〜22g， 雌雄各半， 随机选取 10只作为正常 对照组， 雌雄各半， 其余 30只实验时腹腔注射环磷酰胺 60mg/Kg体重， 随机分 成 3组， 每组 10只， 雌雄各半， 即空白对照组、姜黄素固体分散体组、 姜黄素- 锌固体分散体组。灌胃给受试样品，空白对照组及正常对照组小鼠给予同量 PVP， 姜黄素组、 姜黄素 -锌固体分散体组小鼠分别对应给予含姜黄素或姜黄素-锌 50mg/ g体重相应制剂受试样品， 连续给受试样品 30d， 第 30d各组小鼠禁食一 天， 但提供饮水。 第 31d， 处死各组小鼠， 称量各组小鼠的体重， 胸腺及脾脏重 量， 计算胸腺指数及脾脏指数， 比较各组间的差异。 三、 实验结果
表 4 对小鼠胸腺和脾脏的作用 （ ±SD， n=10) 项目 剂量 胸腺指数 脾脏指数 组别 （mg/kg) (mg/10g体重) (mg/10g体重） 空白对照组 -- 13. 21 ±4. 35 47. 24±8. 35 正常对照组 一 28. 14±5. 65*** 75. 31 ± 12. 25*** 姜黄素一锌组 50 22. 84±5. 37***# 61. 24± 15. 24***# 姜黄素组 50 17. 62±4· 52 48. 27 ±11· 34** 与空白对照组相比， **Ρ〈0. 01， ***Ρ〈0. 001。 与姜黄素组比较, #Ρ〈0. 05
与空白对照组相比， 受试物能够对小鼠胸腺指数及脾脏指数显著升高,具有 提高免疫功能作用。姜黄素锌 固体分散体对小鼠胸腺指数及小鼠脾脏指数升高 优于姜黄素固体分散体。 实施例 7 对肾功能衰竭的作用
一、实验材料
1.1动物： Η小鼠， 18-22g， 雌雄各半， 由广东省医学实验动物中心提供。 1.2姜黄素-锌固体分散体 (姜食素 -锌： PVP, 质量比为 1 : 8)， 姜黄素固体分散体 (姜黄素： PVP， 质量比为 1 : 8), 制备方法同实施例 2， 由中山大学生命科学学 院中药与海洋药物实验室提供。聚乙烯吡咯烷酮 PVPk30(简称 PVP)，进口分装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌胃给药。 二、 实验方法
内毒素致动物急性肾衰模型
选取小鼠 40只，随机分为 4组，分别为正常对照组、病理模型组、姜黄素组、 姜黄素-锌固体分散体组， 每组 10只。 正常对照组、 病理模型组小鼠分别灌胃 给予等量 PVP溶液， 姜黄素固体分散体组、 姜黄素-锌 固体分散体组小鼠分别 对应给予含姜黄素或姜黄素-锌 50mg Kg体重相应制剂受试样品，然后立即尾静 脉注射内毒素 0.06 g kg (正常对照组注射等量的等量 PVP溶液)。 5 h后，小鼠摘 眼球取血，测定血清尿素氮 (BUN)和肌肝 (Cr)浓度。取各小鼠一只肾分别制成 100 ml- L 1匀浆， 测定谷胱甘肽过氧化物酶 (GSH-Px)活性。
三、 实验结果
与模型组相比， 姜黄素-锌 固体分散体能够有效抑制因内毒素致小鼠血清 BUN. Cr值升高， 提高 GSH-Px酶活性， 对内毒素致动物急性肾衰具有保护作 用， 优于姜黄素固体分散体。
Figure imgf000010_0001
正常对照组 ― 17.5±2.4*** 247.5±23.2*** 50.2±22.4*** 病理模型组 ― 70.2± 20.1 420.3 ±27.8 28.7± 10.6 姜黄素 -锌组 50 35.3 ± 11.4**## 283.0±25.4***囊 34.5 ± 9.2 姜黄素组 50 48.2±7.2*** 362.4±31. 7*** 22.4 ±5. 9* 与病理模型组相比， **P<0.01, ***P<0.001.与姜黄素组比较, ##P<0.01 实施例 8 对治疗老年性痴呆症的药理作用
一、实验样品： 姜黄素-锌固体分散体 (姜黄素 -锌： PVP， 质量比为 1 : 8), 姜黄 素固体分散体 (姜黄素： PVP， 质量比为 1 : 8)， 制备方法同实施例 2， 由中山大 学生命科学学院中药与海洋药物实验室提供。 聚乙烯吡咯烷酮 PVP k30(简称 PVP)， 进口分装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌 胃给药。 MH系小鼠 18-22g， 60只， 雌雄各半， 由广东省医学实验动物中心； 实验药品：致老年痴呆药物 Amyloid β-蛋白 (Αβ 1-42片段)，分子量 4514.1 , Sigma Chemical Co. 出品， 产品号 [107761-42-2]。 二、 方法与结果：
Nffl系小鼠， 40只， 雌雄各半， 随机分为 4组， 每组 10只， 分别为正常对 照组、空白模型组、姜黄素固体分散体组、姜黄素 -锌固体分散体组，每组 10只。 正常对照组、 空白模型组小鼠分别灌胃给予等量 PVP溶液， 姜黄素固体分散体 组、 姜黄素-锌 固体分散体组小鼠分别对应给予含姜黄素或姜黄素-锌 50mg/Kg 体重相应制剂受试样品。各组 NIH小鼠（正常组除外），每只小鼠腹腔注射 0.45 %戊巴比妥钠 0.1 ml / 10g体重麻醉。 碘酒、 酒精消毒头顶皮肤后， 纵向正中切 开颅顶皮肤，暴露颅骨，在 bregma点后 2.0mm，左右旁开 2.5mm处用注射器钻 一小孔，以 5μ1注射器缓速注入 2μ1 Αβ溶液，浓度为 7μ§/μ1， 30分钟内注射完毕， 留针 5分钟， 取出注射器， 缝合皮肤。造模次日开始给药，连续 30d每天灌胃一 次。
跳台法实验- 跳台法实验的原理是： 反应箱底铺有通 36V电的铜栅， 动物受到电击。 其 正常反应是跳上箱内绝缘的平台以避免伤害性剌激。多数动物可能再次或多次跳 至铜栅上， 受到电击又迅速跳回平台， 如此训练 5min， 并记录每只小鼠受到电 击的次数， 即错误次错误次数， 以此作为学习成绩。 24h后重复作试验， 此即记 忆保持试验。 记录受电击的动物数， 第一次跳下平台的潜伏期和 3min的错误次 数。具体做法是：给药末次后次日开始训练。将动物放入反应箱内适应环境 3min, 然后立即通以 36V的交流电。 动物受到电击， 其正常反应是跳回平台， 以躲避 伤害性剌激。小鼠双足同时接触铜栅为触电， 视为错误反应。多数动物可能再次 或多次跳至铜栅上， 受到电击又迅速跳回平台上， 如此训练 5min (同一试验中 所有小鼠训练时间相同），记录每鼠后 5min的错误次数，以此作为学习成绩。 24h 后重复作试验， 即记忆保持试验。 记录动物第一次跳下平台的潜伏期和 3min内 的错误总数， 同时计算动物出现错误反应的频率。
表 6 对 Αβ所致小鼠痴呆模型跳台法实验的作用 （ SD， n=10) 项目 24h后
5min内受
第一次跳下平台潜 3min内错误频率 组别 mg/kg 电击次数 (n)
伏期 (s)
空白组 4. 1 ± 1. 7 81· 5± 16· 7 4. 1 ± 1. 2
1. 5± 1. 2***# 117. 4±21. 4 # 1. 9 + 0. 8***
2· 8± 1· 4 95. 8土 11. 5 2. 5±1. 2* 正常组 1. 4± 1. 0***# 120. 6 ±24. 7***# 1. 5±0. 7***# 与空白对照组比较， *Ρ<0.05, **Ρ<0.01, ***Ρ<0.001与姜黄素组比较 Ρ<0.05， 老年性痴呆症是以大脑神经细胞死亡，神经纤维缠结，大脑神经细胞和脑血 管出现老年性斑块为特征。 往小鼠的海马区注射 Αβ蛋白造成大脑神经细胞和脑 血管形成老年性斑块，从而形成痴呆模型。姜黄素锌 固体分散体对老年痴呆模 型小鼠脑记忆的获得、巩固均表现出明显的增强作用， 提示具有抗痴呆作用，优 于姜黄素固体分散体。 实施例 9对动脉硬化的药理作用
―、 实验样品： 姜黄素-锌固体分散体 (姜黄素 -锌： PVP， 质量比为 1 : 8), 姜黄 素固体分散体 (姜黄素： PVP， 质量比为 1 : 8), 制备方法同实施例 2, 由中山大 学生命科学学院中药与海洋药物实验室提供。 聚乙烯吡咯垸酮 PVP k30(简称 PVP), 进口分装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌 胃给药。 实验动物： SD系雄性大鼠， 由广东省医学实验动物中心提供。 二、 实验方法： 选择健康 S.D.系大鼠， 体重 275i20g， 雄性， 共 30只， 随机分 组， 分别为空白模型组、姜黄素固体分散体组、姜黄素-锌 固体分散体组， 每组 10只。 空白模型组大鼠灌胃给予等量 PVP溶液， 姜黄素固体分散体组、姜黄素- 锌 固体分散体组大鼠分别对应给予含姜黄素、 姜黄素-锌 80mg/Kg体重相应制 剂受试样品。 按表 9连续给受试样品 15d， 第 16d禁食 12h， 给受试样品 lh后， 用 4%戊巴比妥钠 42mg/kg腹腔注射麻醉， 仰卧体位固定动物手术台上， 进行颈 动脉-颈静脉环插管手术； 将大鼠气管分离， 插入一短塑料管， 随时将气管分泌 物从套管中吸出。剖离左颈总动脉， 右侧颈外静脉， 取一段聚乙烯软质塑料细管 (长管为 7.3cm， 内径 0.15cm, 管厚为 0.03cm)， 内放一根 8cm长的医用缝合线， 塑料管两端用小管将肝素钠水溶液 (50U/ml)充满聚乙烯管。将聚乙烯管的一端插 入右侧颈外静脉， 而另一端插入左颈总动脉， 结扎固定。 准备好后， 打开血流， 血流从左颈总动脉流至聚乙烯管内， 返回右外侧静脉管， 准确开放血流 15min 后， 立即中断血液， 迅速取出丝线， 用电子分析天平称总湿重。 总湿重量减去丝 线重得血栓湿重。
选择健康 SD系大鼠， 体重 275±20g， 雄性， 共 30只， 随机分组， 分为空白 模型组、 姜黄素固体分散体组、姜黄素 -锌固体分散体组， 每组 10只。 空白模型 组大鼠灌胃给予等量 PV 溶液， 姜黄素固体分散体组、 姜黄素-锌 固体分散体 组大鼠分别对应给予含姜黄素、姜黄素-锌 80mg/Kg体重相应制剂受试样品。按 表 9连续给受试样品 15d， 第 16d禁食 12h， 给受试样品 0.5h后， 用戊巴比妥钠 溶液 (30mg/kg体重)腹腔注射麻醉后解剖分离颈总动脉， 插入聚乙烯管取血， 用 3.8%枸椽酸钠作抗凝剂 (抗凝剂： 血体积为 1 : 9)， 800rpm离心 5min， 吸取上层 乳白色的富血小板血浆 (PRP)， 剩余部分再以 3000rpm离心 10min， 取上清液即 贫血小板血浆 (PPP)， 用 PPP调 PRP， 使血小板记数为 60〜90万 /mm3。
按比浊法， 为了消除不同血小板数量的影响， 用同一血样的 PRP和 PPP作 为纵生标 (血小板聚集率)的零点和顶点， 以 PPP为 100%， PRP为 0%，这样的相 对测量就排除了血小板数量的影响。 每支比浊管加入 20μ1 PRP， 用 20μ1诱导剂 (4mM AA, 或 0.2M/L pH 7.4磷酸缓冲液配制 3.0μΜ ADP), 诱导血小板聚集， lOmin内测定血小板聚集率。 对大鼠血栓形成的抑制作用 （ X土 SD, n=10) 剂量 血栓湿重 血栓抑制率 组别 、 (mg/kg) (mg) (%)
空白对照组 一 102. 4± 12. 5
姜黄素 -锌组 80 58. 7 ±7. 9***#ί# 42. 7
姜黄素组 80 80. 4±8. 5*** 21. 5
与空白对照组比： ***ρ<0.001 ; 与姜黄素组比较,灘 PO.001
Figure imgf000013_0001
姜黄素-锌 PVP组 80 49. 8 ±7. 6***### 46. 1 ±7. 2***## 姜黄素组 80 60. 5±6. 7*** 62. 8+8. 3*** 与空白对照组相比： ***p<0.001 ; 与姜黄素组比较 # P<0.01,# *P<0.001 由大鼠血栓形成及血小板聚集实验结果可知， 姜黄素-锌固体分散体具有明 显的抗血栓形成作用及血小板聚集作用，提示其具有抗动脉硬化作用，优于姜黄 素固体分散体。 实施例 10 对大鼠脂代谢紊乱治疗性给药模型的药理作用
―、 实验样品： 姜黄素-锌固体分散体 (姜黄素 -锌： PVP, 质量比为 1 : 8)， 姜黄 素固体分散体 (姜黄素： FVP， 质量比为 1 : 8), 制备方法同实施例 2， 由中山大 学生命科学学院中药与海洋药物实验室提供。 聚乙烯吡咯烷酮 PVP k30(简称 PVP), 进口分装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌 胃给药。
二、 实验动物： SD系雄性大鼠， 由广东省医学实验动物中心提供.
选择健康 S.D.系大白鼠， 雄性， 200±20g， 在实验环境下给普通词料喂养， 观察 6d， 无发现异常。 然后剪尾取血， 分离血清， 测定其各项血脂指标， 淘汰 异常血脂指标的动物， 选取血脂指标正常者， 共 50只， 供给食物和饮水， 这批 大鼠每天上午定时灌服高脂食物 (80%猪油 +4%胆酸钠 +4%胆固醇 +12%蛋黄 粉) 20ml/kg体重， 连续灌服 30d。 第 31d禁食 12h， 剪尾取血， 分离血清， 测定 甘袖三酯 TC含量， 挑出高于 600mg/dl的大鼠 30只， 随机分为 3组， 即高脂对 照组、姜黄素固体分散体组、姜黄素 -锌 固体分散体组。 高脂对照组大鼠分别灌 胃给予等量 PVP溶液， 姜黄素组、 姜黄素-锌 固体分散体组大鼠分别对应给予 含姜黄素或姜黄素-锌 50mg/Kg体重相应制剂受试样品。 连续给受试样品 25d， 第 26d禁食 12h， 将大鼠取血测定各项血脂指标。 对大鼠脂代谢率紊乱治疗性给药血脂指标的影响 （n=10, i土 SD) 项 目 剂量 甘油三脂 (TG) 总胆固醇 (TC) 高密度脂蛋白 组 别 mg/ g (mg/dl) (mg/dl) 醇 (HDL-C) (mgy 高脂对照组 —一 166. 3±24. 5 845. 7 ±80. 6 30. 4±8. 2 姜黄素 -锌组 50 83. 4± 12. 7***囊 384. 2 ±57. 2 灘 72. 3 ±7. 4**** 姜黄素组 50 134. 2上19. 1** 542. 7 ±68. 9*** 59. 2士 6. 3** 与高脂对照组相比： **p<0.01， ***p<0.001； 与姜黄素组比较，
?<0.01，，<0崖 由大鼠脂代谢紊乱预防性给药及治疗性给药模型实验结果表明：与模型组相 比， 姜黄素-锌 固体分散体能够对高血脂症、 高胆固醇症具有明显的治疗作用， 优于姜黄素固体分散体。 实施例 11对糖尿病模型的药理作用
一、 实验样品： 姜黄素-锌固体分散体 (萎黄素 -锌： PVP, 质量比为 1: 8)， 姜黄 素固体分散体 (姜黄素： PVP, 质量比为 1 : 8)， 制备方法同实施例 2， 由中山大 学生命科学学院中药与海洋药物实验室提供。 聚乙烯吡咯烷酮 PVP k30(简称 PVP), 进口分装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌 胃给药。 实验试剂： 四氧嘧啶 (Alloxan), 由 Sigma Chemical Co生产。
二、 实验动物： MH系小鼠， 由广东省医学实验动物中心提供；
三、 方法与结果： 选择健康 IH系小鼠， 雄性， 18〜22g， 50只， 随机选择 10 只小鼠作为正常对照组， 另外 40只禁食 12h后， 腹腔注射四氧嘧啶 250mg/ g 体重， 6h后灌服 2.5g g的葡萄糖液， 使小鼠安全渡过低血糖相， 随后小鼠提供 食物和饮水， 36h后再禁食 12h, 从小鼠眼眶静脉取血， 测定腹腔注射四氧嘧啶 小鼠的血糖值， 选择血糖值大于 200mg/dl的小鼠 30只， 隨机分成 3组， 高血糖 模型组、 姜黄素组、姜黄素-锌 固体分散体组， 每组 10只。 正常对照组、 高血 糖模型组小鼠分别灌胃给予等量 PVP溶液， 姜黄素固体分散体组、 姜黄素 -锌固 体分散体组小鼠分别对应给予含姜黄素或姜黄素-锌 40mg/Kg体重相应制剂受 试样品。 连续给受试样品 12 d。 末次给受试样品前禁食 12h， 小鼠给受试样品后 lh眼眶静脉取血， 采用血糖测定试剂盒测定血糖含量。 表 10 对四氧嘧啶所致小鼠糖尿病模型的药理作用 （ 士 SD， n=10 ) 组别 剂量（mg/Kg体 血糖值 (mM/L) 降糖率 （％)
S)
正常对照组 一 6. 41 ± 1. 20*** ― 高血糖对照组 一 29. 31 ±5. 41 ― 姜黄素 -锌组 40 21. 45± 1. 37***### 26. 8 姜黄素组 40 25. 48 ± 1. 36* 13. 6 与高血糖对照组相比： **p<0.01，***p<0.001与姜黄素组比较 ,*^<0.01 由四氧嘧啶所致小鼠糖尿病模型所致小鼠糖尿病模型实验结果可知，与模型 组相比，姜黄素 -锌 固体分散体具有明显的降血糖作用，优于姜黄素固体分散体。 实施例 12 对乙肝病毒的药理作用
一、 实验样品： 姜黄素-锌固体分散体 (姜黄素 -锌： PVP， 质量比为 1: 8)， 姜黄 素固体分散体 (姜黄素： PVP， 质量比为 1 : 8)， 制备方法同实施例 2， 由中山大 学生命科学学院中药与海洋药物实验室提供。 聚乙烯吡咯垸酮 PVP k30(简称 PVP)， 进口分装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌 胃给药。 实验动物： 新西兰兔， 由中山医科大学实验动物中心提供， 自由饮水， 普通兔饲料喂养。
实验试剂： HBsAg标准品， 由中国药品生物制品检定所提供， HBeAg酶联诊 断试剂盒， 上海实业科华生物工程有限分司生产。 HbsAg强阳性血清， 由广州 中医药大学第一附属医院检验科提供， 为 20份高效价 HBsAg 强阳性血清 (HBsAg、 HBeAg, 抗 -HBc、 HBVDNA均为阳性）混合而成， 置 -20°C冰箱保 存备用。
二、 方法与结果：
含药血清的制备： 选取 24只体重为 2.0 ± 0.2kg的新西兰兔， 随机分成空白 模型组、姜黄素固体分散体组、姜黄素 -锌固体分散体组， 每组 8只， 雌雄各半。 空白模型组新西兰兔分别灌胃给予等量 PVP溶液， 姜黄素固体分散体组、 姜黄 素 -锌固体分散体组新西兰兔分别对应给予含姜黄素或姜黄素-锌 200mg/Kg体 重相应制剂受试样品。每天 1次， 连续 8天。于最后一次给药后 lh， 心脏采血， 无菌分离血清， 部分血清经 56Ό 30min灭活， 部分不灭活， 用 0.45 μπι的微孔 滤膜过滤除菌， 置 -20°C冰箱保存备用。
HBsAg阳性血清中 HBsAg含量的标定： 将 HBsAg强阳性血清用含 20%小 牛血清的生理盐水 1:2、 1:4、 1:8、 1:16......1:16384的倍比稀释， 隨后用 ELISA 法检测各样品和 1 g L、 2 g/L、 5 g/L HBsAg标准品的 P N值， 重复做 3次， 每次均做双份平行实验。 最后根据 HBsAg浓度 --P/N值标准曲线求出 HBsAg强 阳性血清中的 HBsAg浓度， 置 -20°C冰箱保存备用。
空白血清灭活与否对 HBeAg的影响： 取灭活空白兔血清、 未灭活空白兔血 清、含 20%小牛血清的生理盐水各 200μ1分别与 50 μ1500 μ g/LHBsAg阳性血清 在 37Ό下作用 4h，每个样品均做 10孔,然后用 HBeAg酶联诊断试剂盒检测作用 后 HBeAg的 P 值， 并进行 t检验分析。
含药血清、 空白兔血清各 200 μ 1分别与 50 μ 500 μ g/L HBsAg阳性血清在 37Ό下作用 4h, 每个样品均做 10？ L， 然后用 HBeAg酶联诊断试剂盒检测作用 后 HBeAg的 P/N值， 并进行 t捡验分析。
空白血清灭活与否对 HbeAg的影响： 灭活空白兔血清、 未灭活空白兔血 清分别与 500 μ g LHBsAg阳性血清在 37°C下作用 4h，检测作用后 HBeAg的 P/N 值， 结果未灭活空白兔血清对 HBeAg具有抑制作用（P<0.05)，且未灭活空白兔 血清和灭活空白兔血清对 HBeAg的影响有显著性差异（P<0.05),故在以下的实 验中将家兔的空白血清和含药血清均进行 56°C 30min灭活处理。
空白血清灭活与否对 HBsAg的影响 （ ±SD) 组别 孔数 P/N值 含 20%小牛血清的生理盐水 10 5. 02 + 0. 54 灭活空白兔血清 10 4. 96 +0. 41△ 未灭活空白兔血清 10 4. 27 士 0. 39** 注- 与含 20%小牛血清的生理盐水比较 AP<0. 05 ; 与灭活空白兔血清比较
***P<0. 001。 表 12 含药血清对 HBeAg的抑制作用 （X 土 SD) 组别 孔数 P/N值 灭活空白兔血清 10 5. 82 土 0. 44 姜黄素-锌 PVP组 10 3. 57士 0. 43***### 姜黄素组 10 4. 56 ±0. 41*** 注： 与灭活空 兔血清比较 APX).05 ***P<0.001； 与姜黄素组比较, # P<0.05, # P<0.01, ###P<0.001 与空白血清相比， 姜黄素 -锌 固体分散体对乙肝病毒具有明显的抑制作用， 优于姜黄素固体分散体。
实施例 13 对乙酸烧灼诱导大鼠胃溃疡模型作用
实验样品姜黄素-锌固体分散体 (姜黄素 -锌： PVP, 质量比为 1 : 8： ), 姜黄 素固体分散体 (姜黄素： PVP， 质量比为 1: 8)， 制备方法同实施例 2， 由中山大 学生命科学学院中药与海洋药物实验室提供。 聚乙烯吡咯烷酮 PVP k30(简称 PVP), 进口分装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌 胃给药。 '
实验动物： SD系雄性大鼠， 由广东省医学实验动物中心提供。
选择健康 SD大鼠 30只， 雌雄各半，体重 200±10g，实验前大鼠禁食不禁水 24h, 用 30mg/kg戊巴比妥钠麻醉， 无菌条件于剑突下 2cm处打开腹腔， 将内径 5mm, 长 30mm的玻璃管垂直放于胃体部浆膜面上， 向管内加入乙酸 0.02mL lmin后用棉签蘸出乙酸， 用无菌生理盐水冲洗两次， 逢合切口。 随机分成 3组， 每组 10只 （雌雄各半)， 即： 随机分成空白模型组、姜黄素固体分散体组、姜黄 素-锌 固体分散体组， 每组 10 只， 雌雄各半。 空白对照组大鼠灌胃给予 PVP500mg/Kg体重（排除敷料对胃黏膜的影响）， 姜黄素固体分散体组、姜黄素 -锌 固体分散体组大鼠分别对应给予含姜黄素或姜黄素-锌 200mg/Kg体重相应 制剂受试样品。连续给药 14d,第 15d禁食不禁水 24h后，眼眶静脉丛取血 3mL 其中 2 mL注入含 10%EDTA- Na2 30/iL和抑肽酶的试管中， 混匀， 4°C 3000rmp 离心 10min， 分离血浆， -20°C保存待测； 余下 1 mL注入玻璃管中， 室温静置一 段时间后分离血清， -20°C保存待测。 血清 NO和血浆 ET测定按说明书操作。
脱颈椎处死大鼠， 打开腹腔结扎幽门贲门取胃， 向胃中注入 4%甲醛溶液 5mL, 固定 30min后沿胃大弯剪开， 将胃外翻， 洗去食物残渣。溃疡成圆形或椭 圆形，测量溃疡处的最长径和最短径， 以他们的均值作为溃疡指数。在各组间进 行统计比较， 按公式 1-1计算溃疡抑制率。
/、 对照组溃疡直径均值一实验组溃疡直径均值
溃疡恩合百分率 （％) = 对照组溃疡直径均值 Xl00 % 与模型组相比， 姜黄素-锌 固体分散体对该模型胃溃疡具有显著的抑制作 用， 优于姜黄素固体分散体。
表 13对乙酸烧灼性胃溃疡的作用 （ 土 SD， n=10) 组别 剂量 (mg kg) 溃疡指数 愈合率（％) 模型组 (给予辅料 PVP) 500 9.76+0.69 ― 姜黄素 -锌组 50 3.72 ±0.35***珊 61.9 姜黄素组 50 5.26+0.10*** 46.1 与模型组比较， ***p < 0.001; 与姜黄素组比较， #P<O.O5， #mp<o.001 对血清 NO与血浆 ET含量的影响：
与模型组相比， 姜黄素-锌固体分散体具有弁高血清 NO 7j平作用， 降低血 桨 ET K平作用， 优于姜黄素固体分散体。
表 14对各组血清 NO和血浆 ΕΤ含量的影响 （ ±SD， n=10)
组别 齐 U量 (mg kg) 血清 ΝΟ( μ mol/mL) 血浆 ET (pg/mL) 模型组 (给予辅料 PVP) 500 31.4±6.5 213.1 ±25.3 姜黄素 -锌组 50 52.5±11.4***# 125.7 ±14.2***** 姜黄素组 50 4·0.4±7.6* 156.7 ±20.9*** 与模型组比较， ***ρ < 0.001; 与姜黄素组比较， ###Ρ<0. 001 实施例 14 对大鼠棉球肉芽肿的炎症实验
实验样品： 姜黄素-锌固体分散体 (姜黄素 -锌： PVP, 质量比为 1: 8), 姜黄 素固体分散体 (姜黄素： PVP， 质量比为 1 : 8)， 制备方法同实施例 2， 由中山大 学生命科学学院中药与海洋药物实验室提供。 聚乙烯吡咯烷酮 PVP k30(简称 PVP), 进口分装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌 胃给药。
实验动物： SD系雄性大鼠， 由广东省医学实验动物中心提供
选取体重健康的雄性 SD大鼠 30只， 体重 180-200g， 随机分成 3组， 分别 为病理模型组、姜黄素固体分散体组、姜黄素 -锌固体分散体组，每组 10只大鼠。 大鼠在乙醚浅麻醉下腹部去毛并用 75%酒精和碘酒消毒。 在腹部正中做一个切 口， 将两个灭菌棉球 （每个棉球重 20mg, 高压灭菌， 各加入氨苄青霉素每个 lmg/O.lml, 50°C烘箱烤干）分别植入大鼠两侧腋窝皮下（或两侧腹股沟皮下）， 将切口缝合后用 75%酒精和碘酒消毒。于手术后当天开始灌胃给药，空白对照组 大鼠灌胃给予等量 PVP溶液， 姜黄素固体分散体组、 姜黄素 -锌固体分散体组分 别对应给予含姜黄素或姜黄素-锌 60mg Kg体重相应制剂受试样品。 每天一次， 连续 14d。 15d颈椎脱臼处死， 剥离并取出棉球肉芽组织， 然后在 60Ό烘箱内烘 12h后称干重， 相比各组肉芽肿胀重量。 实验结果进行统计学处理并比较组间差 大鼠棉球肉芽肿实验
植入棉球后， 大鼠伤口愈合良好。连续给药 14d后， 空白对照组大鼠腋下肉 芽肿非常明显。 与模型组相比， 姜黄素 -锌固体分散体能够明显抑制棉球肉芽肿 的增生， 具有抗炎作用， 优于姜黄素固体分散体。 表 15 对大鼠棉球肉芽肿的影响 (；±s)
组别 剂量 肉芽肿重量 抑制率
(fflg · Kg—1 ) (mg) ( % )
模型组 -- 125. 4±29. 5
姜黄素 -锌组 50 66. 8 ±7. 2***### 46. 7
姜黄素组 50 85. 4 ±9. 6*** 31. 9
与空白对照组相比， S"P<0.001 ; 与姜黄素组比较, ^ο.οοι 实施例 15 抗肿瘤实验
实验样品： 姜黄素-锌固体分散体 (姜黄素 -锌： PVP, 质量比为 1 : 8)， 姜黄 素固体分散体 (姜黄素： PVP, 质量比为 1 : 8)， 制备方法同实施例 2， 由中山大 学生命科学学院中药与海洋药物实验室提供。 聚乙烯吡咯烷酮 PVP k30(简称 PVP)， 进口分装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌 胃给药。实验动物： NIH纯系小白鼠， 由广东省医学实验动物中心提供， 艾氏腹 水瘤肿瘤、 移植性肉瘤 (S180)和移植性肝癌 (HepA)细胞， 由广州中山大学肿瘤 研究中心提供。 对小鼠移植性肉瘤 S180的作用
取小鼠 30只,于小鼠右腋下皮下接种肉瘤 S180细胞悬液 (1.0 X 107个 /L)0.2ml, 接种次日随机分为 3组， 分别为空白对照组、 姜黄素组、 姜黄素-锌固体分散体 组， 每组 10只小鼠， 空白对照组小鼠灌胃给予等量胶囊， 姜黄素组、 姜黄素- 锌 固体分散体组分别对应给予含姜黄素或姜黄素-锌 60mg Kg体重相应制剂 受试样品。 每天一次， 连续 10d， 停药次日处死动物， 称量体重、 胸腺及脾脏重 量,按下式计算抑瘤率- 抑瘤率 (％)=(1—给药组平均瘤重 /对照组平均瘤重) xl00%。
对 HepA小鼠生存期的影响：
取小鼠 30只， 于小鼠腹腔内接种 HepA细胞悬液 (L0X107个 /ml)0.2ml， 接 种次日随机分为 3组， 分别为空白对照组、姜黄素固体分散体组、姜黄素-锌固体 分散体组， 每组 10只小鼠， 空白对照组小鼠灌胃给予等量 PVP溶液， 姜黄素固 体分散体组、 姜黄素 -锌固体分散体组分别对应给予含姜黄素或姜黄素-锌 60mg/Kg体重相应制剂受试样品。 每天一次， 连续 10d， 记录自然死亡的天数， 按下式计算其生命延长率：
生命延长率 = (实验组平均生存天数 /对照组平均生存天数一 1)χΐ00%。
对小鼠艾氏腹水瘤的作用：
取小鼠 30只,于小鼠腹腔接种艾氏腹水瘤 (1.0X107个/ L)0.2ml,接种次日随机 分为 3组，分别为空白对照组、姜黄素固体分散体组、姜黄素-锌固体分散体组， 每组 10只小鼠，空白对照组小鼠灌胃给予等量 PVP溶液，姜黄素固体分散体组、 姜黄素 -锌固体分散体组分别对应给予含姜黄素或姜黄素-锌 60mg g体重相应 制剂受试样品。 每天一次， 连续 10d， 连续 10d， 记录自然死亡的天数，按下式计 算其生命延长率：
生命延长率 = (实验组平均生存天数 /对照组平均生存天数一 1) Χ 100%。抑瘤 率(《¾)=(1—给药组平均瘤重 /对照组平均瘤重) X 100%。 对小鼠移植性肉瘤 S180的抑制作用（ ±s, n=10 )
组别 剂量 瘤重 抑瘤率 胸腺指数 脾脏指数
mg/Kg (g) (%) (mg/10g体重） (mg/10g体重） 空白对照组 -- 1.50+0.46 一 21.7±3.4 72.5+8.2 姜黄素 -锌组 60 0.62±0.32*** 52.0 30.2±5.72***### 89.6±9.4**##
##
姜黄素组 60 0.84±0.40** 44.0 25.5+3.4** 76.4+5.9** 与空白对照组比较， ** P<0.01, *** P<0.001; 与姜黄素组比较， # P<0.05， P<0.01,###P<0. 001 对 HepA小鼠生存期的影响（ ±s, n=10) 组别 剂量 生存天数 生命延长率
/mg/ Kg (天） (%) 空白对照组 ― 13.5+2.7 一 姜黄素 -锌组 60 20.5+8.5***### 34.1 姜黄素组 60 16.2+3.8 20.0 与空白对照组比较， * P<0.05, ** P<0.01, *** P<0.001 与姜黄素组比 较,囊 P<0. 001 对艾氏腹水瘤小鼠生存期的影响（ s，n=10)
组别 剂量 生存天数 生命延长率
/mg/ Kg (天） (%)
空白对照组 ― 14.2+2.5 一
姜黄素 -锌组 60 19.4+5.2*## 36.6 姜黄素组 60 17.9+5.2 26.0 与空白对照组比较， * P<0.01;与姜黄素组比较 ,^ Ρ^.ΟΙ 与空白对照组相比， 姜黄素-锌固体分散体对 s180肉瘤有显著的抑制作用， 对艾氏腹水瘤小鼠、 HepA小鼠生存期延长， 优于姜黄素固体分散体。 实施例 16对四氯化碳所致大鼠肝损伤的保护作用
实验样品： 姜黄素-锌固体分散体 (姜黄素 -锌： PVP, 质量比为 1 : 6), 姜黄 素固体分散体 (姜黄素： PVP, 质量比为 1: 6)， 制备方法同实施例 2， 由中山大 学生命科学学院中药与海洋药物实验室提供。 聚乙烯吡咯烷酮 PVP k30(简称 PVP), 进口分装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌 胃给药。 实验动物： SD纯系大白鼠， 由广东省医学实验动物中心提供；
取健康 SD大鼠 40只，雄性，体重 230±25g， 随机分成 4组， 分别为正常对 照组、 空白对照组、 姜黄素固体分散体组、 姜黄素-锌 固体分散体组， 每组 10 只大鼠， 空白对照组、 正常对照组大鼠灌胃给予等量 PVP溶液， 姜黄素组、 姜 黄素-锌 固体分散体组分别对应给予含姜黄素或姜黄素-锌 60mg g体重相应 制剂受试样品。从开始实验之日起，正常对照组大鼠腹腔注射花生油 5ml/kg，其 余五组大鼠腹腔注射 25%CCl4-花生油 5nJ/kg， 连续给药 10天， 第 11天禁食， 不禁水 24h,给药 lh后取血离心 300Qrpmxl0min,测定血清中 ALT、 AST、 Alb 含量。
与空白对照组比较， 姜黄素-锌固体分散体具有抗肝损伤作用， 抑制血清中
ALT和 AST的含量， 同时显著的提高血清中 Alb的含量， 优于姜黄素固体分散 体。
表 19 对大鼠血清 ALT、 AST. Alb的影响 （mean±SD， n =10)
组别 齐 !j量 mg/kg ALT (U/ L) AST (U/L) Alb (g/L) 正常对照组 ― 38.44±9.36*** 145.63±15.25*** 51.25 ±4.92 * 空白对照组 ― 214.77±19.47 253.27±26.41 32.67 ±5.12 姜黄素 -锌组 60 94.52±25.29**** 172.30±21.80*** # 42.15 ±4.22**， 姜黄素组 60 129.82 ±28.26*** 195.62 ±27.92** 36.60 ±4.38*** 与空白对照组比较, *** P<0.001;与姜黄素组比较 ,β P<0.01
实施例 Γ7对强迫小鼠游泳不动时间的影响
实验样品： 姜黄素-锌固体分散体 (姜黄素 -锌： PVP， 质量比为 1 : 8), 姜黄 素固体分散体 (姜黄素： PVP, 质量比为 1 : 8), 制备方法同实施例 2， 由中山大 学生命科学学院中药与海洋药物实验室提供。 聚乙烯吡咯垸酮 PVP k30(简称
PVP), 进口分装， 由广州化学试剂批发部提供。 使用时样品溶于水中分别用灌 胃给药。 实验动物： 昆明纯系小白鼠， 由广东省医学实验动物中心提供；
雄性昆明小鼠 30只， 体重 18〜25g， 随机分成 3组， 每组 10只， 分别为正 常对照组、 空白对照组、姜黄素固体分散体组、姜黄素-锌 固体分散体组。 空白 对照组给予等量 PVP溶液，每日上午准时灌胃给药， 1次 /d，连续 8d。小鼠末次 灌胃给药后 lh， 将小鼠置于水深 10cm的玻璃皿 (高 30cm， 直径 18cm)， 小鼠游 泳 2πώι后， 立即开始观察， 观察持续 4min， 累计此 4min内小鼠在水中停止挣 扎， 或呈漂浮状态， 仅有细小的肢体运动以保持头部浮在水面的持续时间 (不动 时间)。 比较给药组和对照组在后 4min内的累计不动时间。 对小鼠强迫游泳不动时间的影响 ±S.D.， n=10) 组别 剂量 (mg kg) 不动时间 (S) 改善率 (％)
空白对照组 ― 120.12±28.43 一
姜黄素 -锌组 50 70.51+20.24**** 41.02
姜黄素组 50 92.43±23.04* 23.05
与空白对照组比较， * P<0.01, ***P<0.001; 与姜黄素组比较, # P<0-05 对小鼠悬尾不动时间的影响
雄性昆明小鼠， 体重 18〜25g， 共 30只， 随机分成 3组， 每组 10只， 分别为空 白对照组、姜黄素固体分散体组、姜黄素-锌 固体分散体组。 空白对照组给予等 量 PVP溶液， 每日上午准时灌胃给药， 1次 /d，连续 8d。小鼠末次灌胃 lh后将小鼠 尾端 2cm处固定于水平面上， 使其呈倒挂状态， 头部离台面 30cm， 四周以板隔离 动物视线， 首先让小鼠适应 2min, 记录小鼠在后 6min内的不动时间。
对小鼠悬尾不动时间的影响 （ 士 S.D., n=10)
组别 齐 (J量 (m.g/kg) 不动时间 (S) 改善率 (％) 空白对照组 ― 141.75±31.24 一 姜黄素 -锌组 50 74.26±23.41***# 47.61 姜黄素组 50 105.92±29.28* 25.28 与空白对照组比较， * Ρ<0.01 , *** P<0.001; 与姜黄素组比较, # P<0.05
小鼠在强迫游泳模型中出现的不动状态也反映了动物的绝望行为。姜黄素固 体分散体组、 姜黄素 -锌固体分散体组能显著缩短小鼠强迫游泳不动时间， 与空 白对照组，具有显著性差异， 提示姜黄素固体分散体、姜黄素 -锌固体分散体具有 抗抑郁的作用， 而姜黄素 -锌固体分散体的效果优于姜黄素固体分散体。
小鼠在悬尾模型中出现的不动状态反映了动物的绝望行为。小鼠在强迫游泳 模型中出现的不动状态也反映了动物的绝望行为。姜黄素固体分散体组、姜黄素 -锌固体分散体组能显著缩短小鼠强迫游泳不动时间，与空白对照组，具有显著性 差异， 提示姜黄素固体分散体、 姜黄素-锌固体分散体具有抗抑郁的作用， 而姜 黄素 -锌固体分散体的效果优于姜黄素固体分散体。

Claims

权 利 要 求 书
1.一种姜黄素 -锌化合物， 其结构式如 （ I ) 所示 :
Figure imgf000024_0001
( I )
Me:代表 C¾。
2.权利要求 1所述姜黄素-锌化合物的制备方法， 其特征在于将姜黄素和乙 酸锌按摩尔比 1 : 1〜1： 5分别溶于有机溶剂， 于氮气保护或在无氧 45〜70Ό环 境下， 将含乙酸锌的有机溶液滴加到含姜黄素的有机溶液中， 搅拌回流 3〜5小 时， 冷却过滤， 收集沉淀物， 分别用 5〜15°C无水乙醇洗漆沉淀物 4〜6次， 沉 淀物在真空干燥， 得到姜黄素 -锌化合物。
3.如权利要求 2所述的制备方法， 其特征在于所述有机溶剂为乙醇、 丙醇等 溶解乙酸锌的溶液。
4.一种姜黄素-锌化合物固体分散体， 其特征在于由姜黄素 -锌化合物、聚乙 錄吡咯烷酮组成， 姜黄素 -锌化合物和聚乙烯吡咯烷酮的质量比为 1 : 1〜1： 28。
5.如权利要求 4所述的姜黄素-锌化合物固体分散体， 其特征在于所述姜黄 素 -锌化合物和聚乙烯吡咯垸酮的质量比为 1 : 3〜1： 18。
6.如权利要求 4所述的姜黄素-锌化合物固体分散体， 其特征在于所述姜黄 素 -锌化合物和聚乙烯吡咯烷酮的质量比为 1: 5〜1： 16。
7.权利要求 4所述姜黄素-锌化合物固体分散体的制备方法，其特征在于包括 如下歩骤： 将姜黄素 -锌化合物、 PVP按比例混合溶解于无水乙醇中， 通氮气保 护， 经高压均质机均质， 过滤， 滤液喷雾干燥得姜黄素-锌化合物固体分散体。
8.权利要求 4所述姜黄素-锌化合物固体分散体在制备用于治疗免疫功能低 下、 胃溃疡、老年性痴呆症、 肾衰、高血脂症、高胆固醇症、动脉硬化、糖尿病、 脑缺血、 记忆衰退、 心肌缺血、 炎症、 病毒感染、 肿瘤、 自由基损伤、 肝损伤、 抑郁的药物或保健品中的应用。
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