WO2011088715A1 - Use of albiflorin for anti-parkinson's disease - Google Patents

Abstract

Use of albiflorin in preparation of medicaments or health care product for preventing, alleviating and/or treating parkinson's disease. The trials of the present invention prove that albiflorin has distinguished anti-trembling effect and low toxic and side effect.

Description

 Anti-Parkinson's use of paeoniflorin

 The present invention relates to a medicament or health food for preventing, alleviating and/or treating a neurodegenerative disease, and more particularly to a drug or health food resistant to Parkinson's disease. Background technique

 Degenerative diseases of the nervous system are a general term for a group of diseases caused by degenerative degeneration of chronic progressive central nervous tissue. Pathologically, neuronal degeneration and loss of brain and/or spinal cord were observed. The main diseases include Parkinson's disease (PD), Alzheimer's disease (AD), Huntington disease (HD) amyotrophic lateral s clerosis (ALS) ) Wait.

 Parkinson's Disease (PD), also known as 'shock paralysis, is a major type of neurodegenerative disease, an age-related neurodegenerative disease with reduced exercise, increased muscle tone, and Tremor is the main symptom.

 Parkinson's disease is basically a disease of middle and old people; its incidence increases with age; the prevalence of people over 60 years old is 1%. Parkinson's disease is very slow, often with motor symptoms, such as tremors, and the sides are not symmetric. Later, some patients will have psychiatric symptoms such as depression, dementia, personality changes and so on. The survival time of untreated patients is 1 to 30 years, mostly 8 to 10 years. The major lesions in Parkinson's disease are characterized by selective loss of dopaminergic neurons in the melanocytes, decreased dopamine content in the striatum, and the presence of Lewy bodies in the presence of substantia nigra and blue spots. The etiology has not yet been fully understood. At present, most scholars believe that environmental and genetic factors play an important role in the pathogenesis of Parkinson's disease.

 The drug treatment of Parkinson's disease is still dominated by levodopa-type drug replacement therapy, but these preparations have large side effects. Although they can significantly improve the clinical symptoms of patients in a short period of time, they cannot slow down the progress of the disease. And with the prolonged application time, the effect is gradually weakened, and the side effects become more and more serious, which accelerates the death of residual dopaminergic neurons. Therefore, it is necessary to explore and develop new drugs for the treatment of Parkinson's disease.

Albiflorin is a monoterpenoid with a molecular formula of C ₂₃ H ₂₈ O _u and a molecular weight of 480.46. Its molecular structure is as shown in formula ( I ). It is a natural active substance derived from the genus Ranunculaceae. Paeonia lactiflora Pall or Paeonia veitchii Lynch root, Peony Rsuffrsticosa Andrz Root.

 (I)

 Paeoniflorin has a lactone ring structure, but has no hemiacetal structure, which is converted into two products under anaerobic conditions, namely paeoni lactone and paeoni lactone 8 . The structure diagram of paeoni lactone and B is as follows:

芍药内酯 A 芍药内酯 B

Paeoniflorin A paeoni lactone B

 Modern pharmacological studies have shown that paeoniflorin has analgesic, sedative, anticonvulsant effects, effects on the immune system, effects on smooth muscle, anti-inflammatory effects, anti-pathogenic microorganisms, liver protection, clinically used mainly for anti-epilepsy, Analgesic, detoxification, vertigo, treatment of rheumatoid arthritis, treatment of bacterial dysentery and enteritis, treatment of viral hepatitis, senile diseases, anti-sulphate flocculation and dissolution of mucus. The use of paeoniflorin for the preparation of a medicament for the prevention, alleviation and treatment of Parkinson's disease has not been reported.

 The inventors have extracted the paeoniflorin, an active ingredient of Parkinson's disease, from a peony medicinal material by a large amount of modern scientific research, and the content thereof is more than 50%, and the paeoniflorin and its The corresponding pharmaceutical preparations have been used to alleviate and treat the pharmacodynamics and pharmacology of Parkinson's disease. The results show that the pharmacological effects of the paeoniflorin monomer are clear, the efficacy of treating Parkinson's disease is remarkable, the side effects are low, and the safety is high. Provide a highly effective and low-toxic drug for patients with Parkinson's disease. Summary of the invention

The primary object of the present invention is to provide a paeoniflorin, a paeoni lactone metabolite, a paeoniflorin composition, Medicament containing paeoniflorin or medicinal extract containing paeoniflorin has the efficacy of preventing, alleviating and treating neurodegenerative diseases, especially the performance and efficacy against Parkinson's disease, and exists in view of the above prior art The problem of providing paeoniflorin or a pharmaceutically acceptable salt thereof or a solvate thereof or a medicinal extract containing an effective amount of paeoniflorin, namely, in the prevention, conditioning and treatment of Parkinson's disease New applications in medicines or health foods.

 In order to achieve the above object, an aspect of the present invention provides a use of a paeoniflorin for the preparation of a medicament for preventing, ameliorating and/or treating a neurodegenerative disease, a health care product.

 Wherein, the neurodegenerative disease is Parkinson's disease.

 In the process of screening natural active ingredients for treating Parkinson's disease, the inventors found that one of the chemical components in the extract of traditional Chinese medicine, the paeoniflorin, has a strong pharmacological effect, and the paeoniflorin is found in the human body. Two metabolites, paeoni lactone A and paeoni lactone B, also have therapeutic effects in the treatment and prevention of Parkinson's disease.

 Here, the "peonylide" refers to a racemate, a stereoisomer or a mixture of stereoisomers mixed in an arbitrary ratio of paeoniflorin.

 In particular, the "peiolactone" further includes two metabolites of paeoniflorin, paeoni lactone.

A (paeoni lactone A) and Paeoni lactone B.

 The paeoniflorin has a purity of 50%, preferably 80%, more preferably 90%. The content of the paeoniflorin is 50%, preferably 80%, more preferably 90%. Wherein the drug consists of paeoniflorin and a pharmaceutically acceptable carrier.

 In particular, pharmaceutically acceptable carriers are generally approved by health care professionals for this purpose and as inactive ingredients of the agent. A compilation of pharmaceutically acceptable carriers can be found in the Handbook of Pharmaceutical excipients, 2nd edition, edited by A. Wade and P. J. Weller; American Pharmaceutical Association, Washington and The Pharmaceutical

Press, London, 1994) found in the reference books.

 In particular, the carrier includes excipients such as starch, water, etc.; a lubricant such as magnesium stearate or the like; a disintegrating agent such as microcrystalline cellulose; a filler such as lactose; a binder; Such as pregelatinized starch, dextrin, etc.; sweeteners; antioxidants; preservatives, flavoring agents, spices, etc.

Wherein the drug is in the form of a tablet, a capsule, a pill, a powder, a granule, a syrup, a solution, an injection, a spray, an aerosol, a patch or the like. Wherein the drug is administered by gastrointestinal administration and parenteral administration.

 In particular, the parenteral administration route is selected from the group consisting of injectable administration, respiratory administration, dermal administration, mucosal administration or intraluminal administration.

 Among them, non-gastrointestinal administration agents are selected as injections, sprays, aerosols, patches, and the like.

 In particular, the parenteral administration preparation comprises a tablet, a capsule, a powder, a granule, a pill, a solution or a syrup.

 Another aspect of the present invention provides a use of a paeoniflorin composition for the preparation of a medicament for preventing, ameliorating and/or treating a neurodegenerative disease, a health care product.

 The neurodegenerative disease described therein is Parkinson's disease.

 Wherein the paeoniflorin composition is a pharmaceutically acceptable salt of paeoniflorin or a solvate of paeoniflorin.

 In particular, the pharmaceutically acceptable salt of the paeoniflorin is a salt which is physiologically acceptable, especially when applied as a medicament to humans and/or mammals.

 Wherein the salt comprises a salt obtained by the addition reaction of paeoniflorin with an acid; and the solvate of the paeoniflorin is a hydrate of paeoniflorin.

 In particular, the acid is selected from one or more of hydrochloric acid, fumaric acid, maleic acid, citric acid or succinic acid, and the acids mentioned are for illustrative purposes only and are not intended to be limiting.

 According to still another aspect of the present invention, a medicinal material containing a peony lactone or a medicinal material extract containing a peony lactone for use in the preparation of a medicament for preventing, alleviating and/or treating a neurodegenerative disease, a health care product.

 The neurological degeneration described therein is Parkinson's disease.

 Wherein, the medicinal material containing paeoniflorin is selected from the group consisting of peony or peony bark.

 In particular, the peony is white or red peony, preferably white peony.

 In particular, the peony lactone glycoside in the medicinal material extract containing the peony lactone has a purity of 6%.

 In particular, the paeoniflorin-containing extract of the paeoniflorin-containing medicinal material has a purity of 6 to 49%, preferably 10 to 40%, more preferably 14 to 24%.

 Another aspect of the present invention provides a use of a paeoniflorin metabolite for the preparation of a medicament for preventing, ameliorating and/or treating a neurodegenerative disease, a health care product.

 Wherein the metabolite of paeoniflorin is paeoni lactone and paeoni lactone 8.

In particular, the neurodegenerative disease is Parkinson's disease. According to still another aspect of the present invention, a medicament for preventing, alleviating and/or treating Parkinson's disease, comprising a paeoniflorin.

 Wherein the paeoniflorin has a purity of 50%, preferably 80%, more preferably 90%. Wherein the medicament further comprises a pharmaceutically acceptable carrier.

 A further aspect of the present invention provides a medicament for preventing, alleviating and/or treating Parkinson's disease, comprising at least one of the following: a paeoniflorin metabolite, a paeoniflorin composition, and a paeoniflorin-containing Medicinal material or medicinal extract containing peony lactone.

 Wherein the drug consists of one of a paeoniflorin metabolite, a paeoniflorin composition, a paeoniflorin-containing medicinal material or a paeoniflorin-containing medicinal material extract and a pharmaceutically acceptable carrier.

 In particular, the peony lactone glycoside in the medicinal material extract containing the peony lactone has a purity of 6%.

 In particular, the paeoniflorin-containing extract of the paeoniflorin-containing saccharide has a purity of 6% to 49%, preferably 10% to 40%, more preferably 14% to 24%.

 Wherein the metabolite of paeoniflorin is paeoni lactone and paeoni lactone^

 Among them, the paeoniflorin composition selects a pharmaceutically acceptable salt of paeoniflorin and a solvate of paeoniflorin.

 In particular, the pharmaceutically acceptable salt of the paeoniflorin is a salt which is physiologically acceptable, especially when applied as a medicament to humans and/or mammals.

 Wherein the salt comprises a salt obtained by the addition reaction of paeoniflorin with an acid.

 In particular, the acid is selected from one or more of hydrochloric acid, fumaric acid, maleic acid, citric acid or succinic acid, and the acids mentioned are for illustrative purposes only and are not intended to be limiting.

 Wherein the solvate of the paeoniflorin is a hydrate of paeoniflorin

 Wherein, the medicinal material containing paeoniflorin is selected from the group consisting of paeoniflorin or peony bark, preferably white peony.

 In particular, the carrier includes an excipient such as starch, water or the like; a lubricant such as magnesium stearate or the like; a disintegrating agent such as microcrystalline cellulose; a filler such as lactose; a binder; Such as pregelatinized starch, dextrin, etc.; sweeteners; antioxidants; preservatives, flavoring agents, spices, etc.

 The medicament may be formulated into various dosage forms such as tablets, capsules, pills, powders, granules, syrups, solutions, injections, sprays, aerosols, patches, and the like, by methods well known in the art.

According to still another aspect of the present invention, there is provided a health food for preventing, alleviating and/or treating Parkinson's disease, which comprises one of the following: paeoniflorin, paeoniflorin metabolite, paeoniflorin composition,芍 A medicinal material of a drug lactone or a medicinal material extract containing a peony lactone.

 Wherein the paeoniflorin has a purity of 6%.

 In particular, the paeoniflorin-containing extract of the paeoniflorin-containing medicinal material has a purity of 6% to 49%, preferably 10% to 40%, more preferably 14% to 24%.

 The present invention also provides a method of treating a Parkinson's disease, comprising administering to a subject a therapeutically effective amount of a pharmaceutical composition of the paeoniflorin, wherein the therapeutically effective amount is 0.6 to 4 mg/kg · d, preferably 1~3. 5~3mg/kg · d. More preferably, it is 1. 2~3mg/kg · d.

 The term "therapeutically effective amount" as used herein, unless otherwise indicated, is the amount of the drug in need of an effective effect; "therapeutically effective amount" is adjusted and varied, and is ultimately determined by the medical personnel, the factors considered including the route of administration. And the nature of the preparation, the recipient's weight, age, etc., as well as the nature and severity of the condition being treated.

 The invention has the following distinct advantages:

 1. The present invention has discovered a new medicinal value for the known compound paeoniflorin or a pharmaceutically acceptable salt thereof or a solvate thereof, and is used for anti-Parkinson's disease (peoside lactone glycoside) The DA content of the striatum, the number of rotations of the 6-0HDA model rats, and the shortening of the climbing time of the MPTP model mice are significant, and can be prepared as a medicine or health food for preventing, regulating and/or treating Parkinson's disease. Thus, it has opened up a new field for the application of medicinal herbs such as peony.

 2. The series of experimental studies of the present invention prove that paeoniflorin has a significant preventive and therapeutic effect on Parkinson's disease, and the main active ingredients of paeoniflorin and paeoniflorin extract are more effective than paeoniflorin. .

 3. The anti-Parkinson's disease mechanism of the present invention has clear action mechanism, remarkable curative effect, small toxic and side effects, good safety, long-term use, high drug-forming property and good medicinal prospect.

 4. The raw materials of the invention have rich sources, low cost, safe clinical use, simple preparation process, can be made into various dosage forms, and have small dosage and convenient use, so it is easy to promote.

5. The invention can be used for preparing a medicament for preventing and treating Parkinson's disease by using a single component of paeoniflorin active ingredient, and also using paeoniflorin and other active ingredients (for example, paeoniflorin, schisandrol or other antioxidants, etc. Compound) A combination of multiple targets for the preparation of a compound drug against Parkinson's disease. Specific embodiment The invention is further illustrated below in conjunction with specific embodiments. However, the examples are intended to be illustrative only and not to limit the scope of the invention. The experimental methods in which the specific experimental conditions are not indicated in the following examples are usually carried out according to conventional conditions or according to the conditions recommended by the manufacturer.

 The beneficial effects of the medicament of the present invention are further illustrated by experimental examples which include pharmacodynamic experiments of the medicament of the present invention.

Experimental Example 1 Effect of paeoniflorin on rat striatum DA

1.1 Experimental materials

 Wistar rats, male, weighing 210 ± 20 g, were provided by the Viton Lihua Animal Experimental Center.

 Paeoniflorin (purity 96.77%) was purchased from Shanghai Eternal Biotechnology Co., Ltd.

 Dopamine (DA), purchased from Sigma.

 Acetonitrile, methanol (both chromatographically pure, Fisher Scientific).

 Waters510 pump, M464 electrochemical detector DL-822 chromatography workstation (Dalian Institute of Chromatography Center), MSE150 ultrasonic pulverizer.

1. 2 Experimental methods

 Rats were randomly divided into 5 groups, 5 rats in each group, namely saline control group, high dose of paeoniflorin (12 mg/kg), divided into 2 groups (ie 30 min group, 60 min group), paeoniflorin The low dose (6 mg/kg) group was divided into 2 groups (gp30min group, 60min group).

 According to lmL/100g (body weight), each group of rats was intraperitoneally injected with 10% urethane for anesthesia. After anesthesia, 0.5 mL/lOOg (body weight) was administered to the femoral vein for intravenous saline and high dose of paeoniflorin ( 2. 4mg/mL) and low-dose paeoniflorin (1. 20 mg / mL), 5 rats at each time point 30 min, 60 min after administration, take the brain, remove the cerebellum and brain stem, filter paper Blood was removed, and the bilateral striatum was carefully separated on ice, weighed, and immediately frozen in liquid nitrogen.

 The method of perchloric acid (HC104) solution is prepared by adding 1 mg of striatum to each gram of striatum and pre-cooling to 0 to 4 ° C for a concentration of 0. 2moI / L perchloric acid (HC104) solution. Contains lg/mL of dihydroxybenzoic acid (DHBA). The striatum was homogenized by MSE150 ultrasonic pulverizer. The homogenate was centrifuged at 02000 °C for 12000 rpm for 10 min. The supernatant was injected for 20 μί, and the liquid chromatographic determination was carried out. The internal standard method was used to calculate the sample. The content of dopamine (DA) is shown in Table 1.

Table 1 Determination of DA content in rat striatum Group U quantity (mg/kg) Number of samples DA ( gg - ¹ ) Blank control group 5 17. 4467 ± 1. 1212 Paeonilide high dose group (30min group) 12 5 19. 7432 ± 0· 2314* * High dose group of paeoniflorin (60min group) 12 5 19. 8218 ± 0. 2481** Low dose group of paeoniflorin (30min group) 6 5 18. 3825 ± 0. 4299* Low dose of paeoniflorin Group (60min group) 6 5 18. 4721 ± 0· 2331* Note: Compared with the blank control group, *Ρ<0. 05, **Ρ<0. 01

 Experimental results show that:

 1. Compared with the blank control group without injection of paeoniflorin, the content of DA in rat striatum was significantly increased after administration of paeoniflorin (Ρ<0.01, Ρ<0.05), indicating that paeoniflorin It has the effect of increasing the DA content in the striatum and is dose dependent in a certain degree.

 2. Patients with Parkinson's disease have pathological changes in the substantia nigra cells in the midbrain, resulting in a significant decrease in dopamine (DA) content in the striatum, resulting in a decrease in the function of inhibiting acetylcholine and abnormal globus pallidus in the brain. Active, causing Parkinson's disease. In addition, due to the decrease in DA content, extrapyramidal dysfunction is caused, resulting in a Parkinson's disease. By increasing the content of dopamine (DA) in the striatum, it can replace the therapeutic effect and maintain the normal function of the body. Therefore, the paeoniflorin has the medicinal effect of preventing, alleviating or treating Parkinson's disease, and can be used for Prevention and treatment of Parkinson's disease. Experimental Example 2 Effect of paeoniflorin on rotational behavior of striatum 6-OHDA induced damage model rats

 The disappearance and degeneration of cytochrome in the substantia nigra is the main pathological change of Parkinson's disease (PD), and 6-hydroxydopamine (6-0HDA) has selective damage to dopaminergic neurons. The damage caused by unilateral injection of 6-hydroxydopamine in the nigral striatum of the rat brain can cause asymmetry in the hemispheres of the animal and dopamine receptor (DA-R) hypersensitivity. Apomorphine is a post-synaptic membrane D2 receptor agonist. When the substantia nigra pars compacta (SNpc) dopaminergic neurons are deficient in more than 90%, hypersensitivity of the postsynaptic membrane will occur, and apomorphine-induced model mice will appear. The rotational behavior to the injury side, the number of rotations is inversely related to the number of tyrosine hydroxylase positive neurons in the SNpc. Paeoniflorin can increase the DA content of the nigrostria and has anti-free kinase activity. In this experiment, the paeoniflorin can significantly reduce the number of rotations of the model animals.

 2. 1 Experimental materials

SD male rats, weighing 240g~260g, purchased from Beijing Weitong Lihua Company, certificate number: SCXK (Beijing) 2002-0003.

 Apomorphine, purchased from EHSY Western Region China Product Service Center, purity 98. 5%; Lot number: L6170132. Paeoniflorin (96.77%), purchased from Shanghai Eternal Biotechnology Co., Ltd.

 Positive drug: Water-soluble Madopar, purchased from Shanghai Roche Pharmaceutical Co., Ltd.

2. 2 Experimental methods

 2. 2. 1 Preparation of the lateral PD model

 SD rats were placed on the locator with 10% chloral hydrate solution and intraperitoneal injection of 450 mg/kg (body weight). The tooth rod was 2 mm higher than the ear rod, and the injection site was on the right side of the substantia nigra and midbrain abdomen. Side covered part (VTA). 5mm, The depth of the surface of the skull is 8. 5mm, the depth of the surface of the skull is 8. 5mm, and the depth of the surface of the skull is 8. 5mm. 5μ1 6-0HDA (concentration 2μ / μ1, dissolved in sterile saline containing 0.2% ascorbic acid), injected at lg/min, needle for 2 minutes before needle extraction, then 1mm /min speed to withdraw the needle, seal the skull joint with filling teeth, suture the scalp; muscle injection of penicillin 100,000 units. The rotational behavior of the animals was induced by intraperitoneal injection of apomorphine (0.5 mg/kg) two weeks after surgery, and the animals at 7 laps/min were qualified.

 2.2.2 Grouping and processing

 Rats with successful modeling were randomly divided into three groups, namely, paeoniflorin group, metoprolol group and model group, with 6 rats in each group. Animals were observed for 1, 2, 3, and 4 weeks of behavioral changes.

 After 2 weeks of routine cage culture, the paeoniflorin group began to receive paeoniflorin (12 mg/kg) at 8 and 20 o'clock every day. The model control group was intragastrically administered with normal saline. The Medopa group was 10 mg/kg. Dopa solution is administered by stomach. For two weeks in a row, the dose was increased to 10 times after two weeks for 2 consecutive weeks.

 Parkinson's (PD) rats were intraperitoneally injected with apomorphine at a dose of 0.5 mg/kg for 4 weeks. The rats were induced to rotate and the behavioral changes were observed. The observation time was 40. minute. The results are shown in Table 2.

 Rotation test of PD rats

 Compared with the model group, *Ρ<0. 05

 Experimental results show that:

1. Compared with the model group, after 4 weeks of treatment, both the paeoniflorin and the positive drug Madopar can be significantly reduced. The number of rotations of the rats (P < 0.05), and the effect of paeoniflorin was basically equivalent to that of the positive drug Madopar.

2. Paeoniflorin can significantly reduce the number of rotations in model rats, indicating that paeoniflorin can increase the number of dopaminergic neurons in SNpc, while Parkinson's disease is mainly caused by the loss of dopaminergic neurons in the substantia nigra. The central nervous system is dysfunctional, so the paeoniflorin can be used to prevent and treat Parkinson's disease. Experimental Example 3 Effect of paeoniflorin on climbing time and spontaneous activity of MPTP-induced Parkinson's disease model mice

MPTP is an inhibitor of mitochondrial respiratory chain complex I. After injection into the body, it can be catalyzed by midbrain monoamine oxidase B to produce MPP ⁺ , which is a selective neurotoxin that specifically damages dopaminergic neurons in the substantia nigra. C57BL mice can produce pathological and neurophysiological changes similar to those of clinical PD patients after application of larger doses of MPTP, which is a better animal model. The crawl time reflects the animal's ability to coordinate movement. If the animal's movement coordination is good, the time required for the climbing of the rod is short, and the poor coordination is long. The number of autonomous activities reflects the animal's ability to exercise at will. Parkinson's disease mice reduced the number of autonomous activities due to muscle stiffness and tremors.

3.1 Experimental materials

 C57BL mice, male, weighing 20 ± 2 g, were provided by the Viton Lihua Animal Experimental Center.

 Paeoniflorin (96.77%), purchased from Shanghai Eternal Biotechnology Co., Ltd.

 i-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), purchased from Beijing Jingyao Yaobang Pharmaceutical Technology Co., Ltd.

 Positive drug: Medoc, purchased from Shanghai Roche Pharmaceutical Co., Ltd.

 Stainless steel measuring rod: Take a stainless steel tube with a length of 0.5m and a diameter of lcm, wrapped with tape, and place a wooden ball with a diameter of 2.5cm at the top of the rod;

 CS-2 mouse autonomous activity program.

3.2 Experimental methods

 3.2.1 Establishment of mouse model of Parkinson's disease caused by MPTP:

 MPTP was directly dissolved in 0.9% sterile physiological saline to prepare an MPTP solution at a concentration of 2 mg/ml. C57BL mice were intraperitoneally injected with MPTP 20 mg/kg/day for 8 consecutive days to establish a MPTP-induced Parkinson's disease mouse model.

3.2.2 grouping and administration The model mice were randomly divided into 4 groups, 15 in each group, namely the model group, the blank control group, the paeoniflorin group, and the positive drug control group.

 The mice in the model group were intraperitoneally injected with MPTP 20 mg/kg/day for 8 consecutive days to prepare model mice. The blank control group was injected intraperitoneally with the model group for the same dose of normal saline for 8 days. The paeoniflorin group was used for the abdominal cavity of mice. Injection of MPTP 20mg/kg/day, continuous injection for 8 days, and intragastric administration of paeoniflorin (12mg/kg); positive drug control group mice were injected with MPTP 20mg/kg/day, continuous injection for 8 days, simultaneous gavage Metopbar (65 mg/kg) was given.

 3.2.3 Determination of mouse climbing time

 The mouse climbing time was determined by the experimental method of Nobutaka Arai: The experimental tool was a self-made stainless steel rod with a length of 0.5 m, a diameter of lcm (which was wrapped with a tape), and a wooden ball of 2.5 cm in diameter at the top. The day before the animal was given MPTP, the pole training was carried out first, and the animals were guided to climb from the top of the pole to the sole of the rod within 10 seconds, each training 4 times. For the formal measurement, the first climbing time of the mouse before the application of MPTP was first determined, and the second climbing time of the animal was measured within 1 to 2 hours after the last MPTP administration.

 The model group, the blank control group, the paeoniflorin group, and the positive drug control group were each subjected to a mouse climbing rod experiment. The specific determination methods are as follows:

 Hold the tail of the mouse, place its head down on the top of the rod (which is based on the mouse's hind limbs), let it climb naturally, record the mouse from the top of the pole to the forefoot contact platform It takes time. The measurement time was 120 seconds, the mouse could not hold the rod, and the person who completely slipped naturally recorded the climbing time of 120 seconds. The difference in the time of the climbing time before and after MPTP administration in each group of mice was calculated (when the climbing time was shorter than that of the untreated, the climbing time was recorded as 0). The results are shown in Table 3.

The effect of MPTP model mouse climbing time (±s, n=15)

 Compared with the model group: *Ρ<0.05; **Ρ<0.01

 Experimental results show that:

1. The time difference between the two groups in the model group was significantly longer than that in the normal control group (Ρ<0.01), but relative In the model group, the time difference between the two groups of mice in the paeoniflorin group and the positive drug (Medopa) group was significantly shorter (P<0.05), and the time difference between the two rods in the paeoniflorin group was more positive. The drug metoprol is short, indicating that the effect of paeoniflorin is superior to the positive drug metoprolol.

 2. Paeoniflorin can significantly shorten the climbing time of model mice, indicating that it can strongly inhibit the damage of MPTP on dopaminergic neurons in the substantia nigra, and help restore the motor coordination of mice, thus effectively Prevent, alleviate and/or treat Parkinson's disease.

3.2.4 Determination of spontaneous activity of mice

 The model group, the blank control group, the paeoniflorin group, and the positive drug control group mice were each taken for 15 spontaneous activity experiments. The specific measurement method is as follows:

 When the computer of the CS-2 mouse autonomous activity program enters the measurement program, the mouse is placed in the autonomous activity measurement box of the CS-2 type autonomous activity program, and 4 mice are simultaneously measured each time. One of the activity boxes was automatically recorded by the computer, and the number of activities in each mouse within 5 minutes was measured for statistical processing. The results are shown in Table 4. Effects on the autonomic activity of mice in the MPTP model (±s, n=15)

 Compared with the model group: *Ρ<0.05; **Ρ<0.01

 Experimental results show that:

 1. Compared with the blank control group, the autonomic activity of the mice in the model group was significantly reduced (Ρ<0.01), while the autonomic activity of the mice in the paeoniflorin group and the positive drug Medopa group increased significantly compared with the model group. (Ρ<0.01).

2. Paeoniflorin can increase the number of spontaneous activities of mice, which is beneficial to improve the ability of mice to exercise at random, thus effectively alleviating symptoms such as muscle stiffness and tremor in Parkinson's disease model mice. It has prevention, relief and/or treatment. The role of Parkinson's disease. Experimental Example 4 Effect of peony extract containing paeoniflorin on climbing time of MPTP-induced Parkinson's disease model mice 4.1 Experimental materials

 C57BL mice, male, weighing 20 ± 2 g, were provided by the Viton Lihua Animal Experimental Center.

 A white peony extract containing paeoniflorin (purity of 45%) was purchased from Shanghai Eternal Biotechnology Co., Ltd. i-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), purchased from Beijing Jingyao Yaobang Pharmaceutical Technology Co., Ltd.;

 Positive drug: Medoc, purchased from Shanghai Roche Pharmaceutical Co., Ltd.

 Stainless steel measuring rod: Take a stainless steel tube with a length of 0.5m and a diameter of lcm, wrapped with tape, and place a wooden ball with a diameter of 2.5cm at the top of the rod;

4.2 Experimental methods

 4.2.1 Establishment of mouse model of Parkinson's disease caused by MPTP:

 MPTP was directly dissolved in 0.9% sterile physiological saline to prepare an MPTP solution at a concentration of 2 mg/ml. C57BL mice were intraperitoneally injected with MPTP 20 mg/kg/day for 8 consecutive days to establish a MPTP-induced Parkinson's disease mouse model.

 4.2.2 Grouping and administration

 The model mice were randomly divided into 4 groups, 15 in each group, namely the model group, the blank control group, the paeoniflorin extract group, and the positive drug control group.

 The mice in the model group were intragastrically injected with distilled water for 3 weeks, and then injected intraperitoneally with MPTP 20 mg/kg/day for 8 days to prepare model mice. The blank control mice were intragastrically administered with distilled water for 3 weeks. The rats in the peony extract group were given an equal dose of normal saline for 8 days; the peony extract group was intragastrically administered with the extract of Radix Paeoniae Alba (24 mg/kg, wherein the purity of the paeoniflorin in the extract was 45%). After 3 weeks, the intraperitoneal injection was performed. MPTP 20mg/kg/day, continuous injection for 8 days; positive drug control group mice were treated with distilled water for 3 weeks, intraperitoneal injection of MPTP 20mg/kg/day, continuous injection for 8 days, and intragastric administration of Madopar ( 65mg/kg).

 4.2.3 Determination of mouse climbing time

 The mouse climbing time was determined by the experimental method of Nobutaka Arai: The experimental tool was a self-made stainless steel rod with a length of 0.5 m, a diameter of lcm (which was wrapped with a tape), and a wooden ball of 2.5 cm in diameter at the top. The day before the animal was given MPTP, the pole training was carried out first, and the animals were guided to climb from the top of the pole to the sole of the rod within 10 seconds, each training 4 times. For the formal measurement, the first climbing time of the mouse before the application of MPTP was first determined, and the second climbing time of the animal was measured within 1 to 2 hours after the last MPTP administration.

The specific measurement method is as follows: Hold the tail of the mouse, place the head down on the top of the rod (placed with the hind limbs of the mouse) Subject to the ball, let it climb naturally, recording the time it takes for the mouse to stand on the top of the pole to the double forelimbs. The measurement time was 120 seconds. The mouse could not hold the rod. If it was completely slippery, the crawling time was recorded as 120 seconds. The difference in the time of climbing before and after MPTP administration in each group of mice was calculated (when the climbing time was shorter than that of untreated after MPTP, the climbing time was recorded as 0). The results are shown in Table 5. Table 5 Effect of Peony extract containing paeoniflorin on climbing time of MPTP model mice (±s, n=15)

 Compared with the model group: *P<0.05; **P<0.01

 Experimental results show that:

 1. The time difference between the two groups in the model group was significantly longer than that in the normal control group (P<0.01), while in the model group, the purity was 55% in the paeoniflorin extract group and the positive drug Madopar. The time difference between the two groups was significantly shortened (P<0.05), and the effect of paeoniflorin extract was slightly better than that of the positive drug.

 2, the peony-containing lactone extract can significantly shorten the mouse climbing time, indicating that it can better avoid the damage of MPTP on dopaminergic neurons in the substantia nigra, and help restore the motor coordination of the mouse, thus It can effectively prevent, alleviate and/or treat Parkinson's disease. Experimental Example 5 Acute Toxicity Test of Paeoniflorin

 The SFP-grade ICR mice (provided by the Vital River Animal Experimental Center) were intragastrically administered with paeoniflorin (purity > 98%, purchased from WAKO, Japan) at a dose of 8.4 g/kg. The administration time was 14 days.

 Observe the respiratory properties and rate, behavior of the mice (especially including vomiting), movement, fur color tension, abdominal shape, stool hardness, body weight, etc.

 After 14 days of administration, no abnormal symptoms and no death were observed in the animals; the body weight of the mice in the administration group was weighed on the 7th and 14th day, respectively, and there was no significant difference compared with the control group (P >0.05).

The results of the acute toxicity test showed that the mice were intragastrically administered with paeoniflorin. At the dose of up to 8.4 g/kg body weight, the drug had no acute toxicity damage to the traditional Chinese medicine tissues and organs of the animals, and the administration of the medicament of the present invention was still safe. Example 1 Preparation of an extract of paeoniflorin

 After pulverizing the chalk, the mixture was heated and refluxed three times with a 70% by volume aqueous solution of ethanol, and the weight of the ethanol solution having a solvent concentration of 70% for three times was 5 times, 4 times, and 3 times the weight of the chalk. (eg 1 kg of chalk, add 5 kg of 70% aqueous ethanol), recover the ethanol, dilute the extract to 4 volumes (eg 1 kg of chalk, dilute the extract volume of 4 liters), filter to clear solution A, stand-by.

 D-101 type macroporous resin was soaked in 95% ethanol overnight, wet packed, steamed and washed to near alcohol-free, and then clear liquid A on D-101 type macroporous resin adsorption column, 1 column volume / hour ( BV/H) Flow rate adsorption, first rinse with 4 column volumes of water, then rinse with 10% ethanol, then elute with 30% ethanol, collect the 2-5 column volume of ethanol eluent, concentrate and dry (Temperature ≤ 70 ° C, relative vacuum ≤ -0.06 Mpa), after smashing through 80 mesh sieve, the content of 30~35% of the paeoniflorin extract, the creaming rate is about 3 to 3.5%.

Example 2 Preparation of paeoniflorin extract by water percolation method

 1) Break the chalk into coarse particles (≤10mm), use 4 times the amount of water as solvent (for example, 1 kg of chalk, soaked in 4 kg of water), immerse for 2 hours, put into the percolator, soak for 1 hour. , percolation, flow rate 0.03ml / g.min, collect 10 times the amount of osmosis liquid (for example, 1 kg of white peony, collect 10 kg of osmosis solution), take the osmosis liquid at atmospheric pressure to concentrate and concentrate The weight ratio is 2: 1 (measured at 70 ° C), and the percolate is obtained.

 2) D-101 macroporous resin was soaked in 95% ethanol overnight, packed in a wet manner, washed with steam and washed until nearly alcohol-free, and set aside.

 3) The D-101 type macroporous resin adsorption column on the percolate A, the ratio of the weight of the macroporous resin in the macroporous resin adsorption column to the weight of the medicinal herbs is 1.5:1, and the loading flow rate is 0.033 ml/min. Discard the effluent, then rinse the resin column with 3 times the amount of resin, the flow rate is 0.033ml/min, discard the effluent, then rinse the resin column with 4 times the amount of resin 50% ethanol, the flow rate is 0.033ml/min, collect 4 The amount of resin in the effluent is taken up. Wash with water until the ethanol content of the effluent is 0%. Repeat the loading with 50% ethanol eluting the effluent. The effluent is concentrated under reduced pressure (temperature ≤ 70 ° C, relative vacuum ≤ -0.06 MPa), to a relative density of 1.30 1.35 (60 ° C measured) thick paste, dried, crushed through 80 mesh sieve, that is, the content For the 15~303⁄4 paeoniflorin extract, the creaming rate is about 4~5%.

The above-mentioned extract of peony (white peony or red peony) with paeoniflorin as the main active active ingredient of anti-Parkinson, if necessary, after adding the auxiliary material, filling the capsule or preparing the tablet, the preferred oral administration of the present invention is obtained. Drug Preparation; or the above-mentioned extract containing the higher purity paeoniflorin (white or red peony) and other anti-Parkinson active active ingredients (such as paeoniflorin, schisandrin, etc.), together with the preparation of the preparation Compound medicine for Parkinson's disease. Example 3 Preparation of Paeoniflorin Capsules

 100 g of pistol lactone having a purity of 96.77% was added to 80 g of starch and 20 g of starch silica gel, mixed and directly poured into gelatin hard capsules to prepare capsules each containing 10 mg of paeoniflorin. Example 4 Preparation of Paeoniflorin Tablets

 100g of penicillin lactone with a purity of 96.77%, crushed through a 100 mesh sieve and mixed with 700g of starch passed through a 100 mesh sieve, then added with an appropriate amount of starch slurry and stirred uniformly, and sieved through a 16 mesh iron sieve, 60°. Dry below C, granules, add appropriate amount of magnesium stearate, mix and feed into a tableting machine for tableting, and prepare tablets containing 10 mg of paeoniflorin. Example 5 Preparation of Paeoniflorin Sodium Chloride Infusion

 Take a purity of 98.5% Paeoniflorin 10g, add 90g of sodium chloride, add water for injection, stir to dissolve, add water for injection to 1000ml, then filter with 0. 22 μ πι microporous membrane, fill and seal, Sterilize. Example 6 Preparation of Drug Lactone Suspension

 200 g of the paeoniflorin extract having the content of paeoniflorin prepared in Example 2 was pulverized to 200 mesh, added to 100 g of swollen sodium carboxymethylcellulose (CMC), stirred uniformly, and added with distilled water to 10 L. The mixture was stirred to prepare a suspension, and each ml of the suspension contained 10 mg of paeoniflorin.

Claims

Claim  1. Use of paeoniflorin for the preparation of a medicament for the prevention, alleviation and/or treatment of a neurodegenerative disease or condition. 2. Use of paeoniflorin for the preparation of a health product for preventing, alleviating and/or treating a neurodegenerative disease or condition. 3. Use according to claim 1 or 2, characterized in that the neurodegenerative disease is Parkinson's disease. 4. Use according to claim 1 or 2, characterized in that the paeoniflorin has a purity of 50%. 5. Use according to claim 1 wherein the medicament consists of paeoniflorin and a pharmaceutically acceptable carrier. 6. The use according to claim 1, wherein the medicament is in the form of a tablet, a capsule, a pill, a powder, a granule, a syrup, a solution, an injection, a spray, an aerosol, a patch. . 7. A medicament for preventing, alleviating and/or treating Parkinson's disease, which comprises at least one of the following substances: paeoniflorin, paeoniflorin metabolite, paeoniflorin composition, paeoniflorin A medicinal material of a lactone or a medicinal extract containing a peony lactone. The medicament according to claim 7, characterized in that the peony lactone glycoside in the medicinal material extract containing the peony lactone is 6%.  9. The medicament according to claim 7, wherein the paeoniflorin composition is selected from the pharmaceutically acceptable salts of paeoniflorin and the solvate of paeoniflorin. 10. The medicament according to claim 7, wherein the medicinal material containing paeoniflorin is selected from the group consisting of paeoniflorin or peony bark. 11. The medicament according to claim 7, wherein the paeoniflorin metabolite is selected from the group consisting of paeoni lactone A and paeoni lactone.  12. A health care product for preventing, alleviating and/or treating Parkinson's disease, which comprises at least one of the following substances: paeoniflorin, paeoniflorin metabolite, paeoniflorin composition, A medicinal material of paeoniflorin or a medicinal material extract containing paeoniflorin. The nutraceutical according to claim 12, wherein the medicinal extract of the peony-containing lactone glycoside has a purity of 6%.  A nutraceutical according to claim 12, wherein said paeoniflorin composition is a pharmaceutically acceptable salt of paeoniflorin or a solvate of paeoniflorin. A health care product according to claim 12, wherein said medicinal material containing peony lactone is selected from medicinal herbs or peony bark. The nutraceutical according to claim 12, wherein said paeoniflorin metabolite is selected from the group consisting of paeoniflorin and paeoni lactone.