Pharmaceutical composition containing cistanche deserticola extract and microbial preparation

Abstract

The invention relates to the technical field of medicines, and particularly discloses a pharmaceutical composition containing a cistanche deserticola extract and a microbial preparation. The pharmaceutical composition containing the cistanche deserticola extract is prepared from the following raw materials in parts by mass: 1 to 3 parts of the cistanche deserticola extract, 0.1 to 0.5 part of glutathione, 1 to 2 parts of dopamine hydrochloride and 2 to 6 parts of montmorillonite. The microbial preparation comprises the following raw materials in parts by mass: 5-12 parts of the pharmaceutical composition containing the cistanche extract, 1-2 parts of compound microorganisms, 10-20 parts of starch and 1-2 parts of superfine silica powder. The invention also discloses application of the pharmaceutical composition containing the cistanche deserticola extract or / and the microbial preparation in preparation of drugs for treating Parkinson's disease. The traditional Chinese medicine composition not only can effectively improve the motor function of Parkinson's disease patients and delay the progress of Parkinson's disease, but also is remarkable in effect, high in safety, simple in preparation method and suitable for large-scale popularization and application.

Description

Technical Field

[0001] The invention relates to the technical field of medicines, and in particular to a medicine composition containing a Cistanche deserticola extract and a microbial preparation. Background Art

[0002] Parkinson's disease is a neurodegenerative disease characterized by the degeneration or disappearance of dopaminergic neurons in the substantia nigra and striatum of the midbrain, and the abnormal accumulation of α-synuclein in cells, which leads to the formation of Lewy bodies, resulting in clinical manifestations such as resting tremor, muscle rigidity, bradykinesia, and postural gait disorders.

[0003] Parkinson's disease is common in middle-aged and elderly people, and the prevalence rate in people over 65 years old is about 1.7%. Currently, with the aging of the population, the prevalence and incidence of Parkinson's disease have doubled with age, seriously endangering the health and quality of life of middle-aged and elderly patients, and has become one of the most serious issues facing geriatric medicine.

[0004] Cistanche is the dried fleshy stem with scaly leaves of the plant Cistanche of the Orobanchaceae family. It is one of the commonly used Chinese medicines. Modern pharmacological research has shown that in addition to its kidney-tonifying and aphrodisiac effects, as well as its intestinal moistening and laxative effects, it also has anti-cancer, anti-aging, and memory-improving effects, and no adverse reactions have been found. In recent years, Cistanche has been widely used in various medicines.

[0005] CN101293016A discloses the use of Cistanche deserticola extract in the preparation of a medicament for treating Parkinson's disease. Cistanche deserticola total glycosides can be used to prepare a medicament for treating Parkinson's disease, which can effectively improve the motor function of Parkinson's disease patients and delay the progression of Parkinson's disease, bringing new hope to the treatment of Parkinson's disease with traditional Chinese medicine. However, its efficacy is not obvious, so it is particularly important to develop a preparation with significant efficacy, high safety, and synergistic effect with the assistance of Cistanche deserticola Chinese medicine composition. Summary of the invention

[0006] The purpose of the present invention is to solve the shortcomings of the prior art and to provide a pharmaceutical composition and a microbial preparation containing a Cistanche deserticola extract.

[0007] A pharmaceutical composition containing a Cistanche deserticola extract comprises the following raw materials by weight: 1-3 parts of the Cistanche deserticola extract, 0.1-0.5 parts of glutathione, 1-2 parts of dopamine hydrochloride and 2-6 parts of montmorillonite.

[0008] Preferably, the Cistanche deserticola extract is prepared by the following specific steps: crushing the dried Cistanche deserticola, sieving, adding ether to immerse for 5-10 hours, ultrasonic extraction at 40-50°C for 1-2 hours, filtering, ultrafiltration, concentrating the filtrate to a relative density of 1.15-1.18, and vacuum drying.

[0009] Preferably, the mass ratio of Cistanche deserticola to ether is 1:2-4.

[0010] Preferably, the ultrasound frequency is 20-30 kHz.

[0011] Preferably, ultrafiltration is performed using an ultrafiltration membrane with a molecular weight cutoff of 5000Da.

[0012] The method for preparing the pharmaceutical composition containing the Cistanche deserticola extract comprises the following steps:

[0013] S1, dispersing montmorillonite in hydrochloric acid and ultrasonically treating it for 10-30 minutes, centrifuging, washing, vacuum drying, adding Cistanche deserticola extract and water and ultrasonically treating it for 1-3 hours to obtain a loaded extract;

[0014] S2. Add dopamine hydrochloride and glutathione to the loaded extract, adjust the pH value of the system to 7.5-8.2, stir for 1-2 hours, let stand for 2-4 hours, centrifuge, wash, and vacuum dry.

[0015] Preferably, in S1, the concentration of hydrochloric acid is 1-2 mol / L, and the frequency of ultrasonic treatment after the montmorillonite is dispersed in the hydrochloric acid is 10-15 kHz.

[0016] Preferably, in S1, the frequency of ultrasonic treatment after adding Cistanche deserticola extract and water is 5-10 kHz.

[0017] A microbial preparation, the raw materials of which include, by mass, 5-12 parts of the above-mentioned medicinal composition containing Cistanche deserticola extract, 1-2 parts of composite microorganisms, 10-20 parts of starch, and 1-2 parts of micropowder silica gel.

[0018] Preferably, the composite microorganisms include: Bifidobacterium bifidum, Lactobacillus plantarum, Butyrivibrio fibrisolvens, Enterococcus faecalis.

[0019] The above strains are widely available and can be purchased from commercial sources, such as China General Microbiological Culture Collection (CGMCC), China Type Culture Collection (CCTCC), American Type Culture Collection (ATCC), German Microbiological Culture Collection (DSMZ), etc. and other commercial sources.

[0020] Preferably, the viable counts of Bifidobacterium bifidum, Lactobacillus plantarum and Butyrivibrio fibrinolyticus in the composite microorganisms are 2-7×10 9cfu / g, and the number of live Enterococcus faecalis in the composite microorganisms accounts for 10-20% of the total live number of the composite microorganisms.

[0021] The preparation method of the microbial preparation comprises the following steps: uniformly mixing the medicinal composition containing the Cistanche deserticola extract, the composite microorganism and water, then adding starch and micro-powder silica gel, granulating and vacuum drying.

[0022] The use of the above-mentioned pharmaceutical composition containing the Cistanche deserticola extract, or / and the above-mentioned microbial preparation in the preparation of drugs for treating Parkinson's disease.

[0023] Beneficial effects:

[0024] 1. The invention uses Cistanche deserticola as raw material, extracts by ether ultrasonic, filters and concentrates. The extraction process is simple and the production cycle is short. The process is stable. The efficacy experiment proves that the extract has significant anti-inflammatory effect, stable efficacy and high safety.

[0025] 2. With the deepening of intestinal microecological research, it is found that intestinal microbial disorder is correlated with Parkinson's motor symptoms. After the natural montmorillonite layer structure is acidified and peeled off, it not only has high adsorption activity for Cistanche deserticola extract, but also further coats the surface with polydopamine, which can effectively adhere to the polarity of microorganisms, effectively enhance the stability and anti-interference of microorganisms in the body, and improve the loading and protection performance of microorganisms.

[0026] The experiment found that: the present invention uses Bifidobacterium bifidum, Lactobacillus plantarum, Butyrivibrio fibrinolyticus, and Enterococcus faecalis to compound and adhere to the polydopamine layer, and then cooperates with the effect of loading Cistanche deserticola extract in the polydopamine layer, which not only shows a significant anti-inflammatory effect, but also has an anti-inflammatory effect that is better than using Cistanche deserticola extract or composite microorganisms alone, especially the best effect of dopamine hydrochloride and glutathione synergistic self-oxidation coating, which can not only improve gastrointestinal dysfunction, but also effectively alleviate Parkinson's non-motor symptoms, with significant effects.

[0027] 3. The present invention prepares a medicament for treating Parkinson's disease by compounding the Cistanche deserticola extract with microorganisms, organic components, etc., which can not only effectively improve the motor function of Parkinson's patients and delay the progression of Parkinson's disease, but also has significant effects, high safety, and a simple preparation method, and is suitable for large-scale promotion and application. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 This is a comparison chart of the areas under the curve of behavioral activity in the rotarod test of mice in the CK group, MC group, S5R group, S5W group, D1W group, D2W group, and D3W group at the 5th and 8th weeks.

[0029] Figure 2This is a comparison of the total time spent by mice in the CK group, MC group, S5R group, S5W group, D1W group, D2W group, and D3W group in the pole climbing test at the 5th and 8th weeks.

[0030] Figure 3 Comparison of dopaminergic neurons in the substantia nigra and dopaminergic nerve fibers in the striatum of mice in the CK group, MC group, S5R group, S5W group, D1W group, D2W group, and D3W group.

[0031] Figure 4 This is a comparison chart of small intestinal conductivity and serum endotoxin content of mice in CK group, MC group, S5R group, S5W group, D1W group, D2W group, and D3W group. DETAILED DESCRIPTION

[0032] The present invention will be further explained below in conjunction with specific embodiments.

[0033] Example 1

[0034] A pharmaceutical composition containing a Cistanche deserticola extract comprises raw materials including: 10 g of the Cistanche deserticola extract, 1 g of glutathione, 10 g of dopamine hydrochloride and 20 g of montmorillonite.

[0035] The Cistanche deserticola extract is prepared by the following specific steps: the dried Cistanche deserticola is crushed, passed through a 100-mesh sieve, added with 2 times the weight of ether and soaked for 5 hours, ultrasonically extracted at a temperature of 40°C for 1 hour with an ultrasonic frequency of 20kHz, filtered, ultrafiltered with an ultrafiltration membrane with a molecular weight cutoff of 5000Da, the filtrate is collected, concentrated at a temperature of 50°C to obtain an extract with a relative density of 1.15, and vacuum dried.

[0036] The method for preparing the pharmaceutical composition containing the Cistanche deserticola extract comprises the following steps:

[0037] S1. Disperse montmorillonite in 200 g of 1 mol / L hydrochloric acid, perform ultrasonic treatment for 10 min at an ultrasonic frequency of 10 kHz, centrifuge, wash once with deionized water, vacuum dry, add Cistanche deserticola extract and 200 g of water, perform ultrasonic treatment for 1 h at an ultrasonic frequency of 5 kHz, and obtain a loaded extract;

[0038] S2. Add dopamine hydrochloride and glutathione to the loaded extract, adjust the pH value of the system to 7.5-8.2, stir at a speed of 100 r / min for 1 hour, let stand for 2 hours, centrifuge, wash once with deionized water, and vacuum dry.

[0039] A microbial preparation, whose raw materials include: 5g of the above-mentioned medicinal composition containing Cistanche deserticola extract, 1g of composite microorganism, 10g of starch, and 1g of micropowder silica gel.

[0040] Among the composite microorganisms, the number of viable Bifidobacterium bifidum was 2×109 cfu / g, the number of viable Lactobacillus plantarum is 2×10 9 cfu / g, the number of viable bacteria of Butyrivibrio fibrinolyticus is 2×10 9 cfu / g, and the viable count of Enterococcus faecalis was 6.67×10 8 cfu / g,

[0041] The number of live Enterococcus faecalis in the composite microorganisms accounts for 10% of the total number of live bacteria in the composite microorganisms.

[0042] The preparation method of the microbial preparation comprises the following steps: uniformly mixing the pharmaceutical composition containing the Cistanche deserticola extract, the composite microorganism and water, stirring at a speed of 50 r / min for 1 hour, adding starch and micro-powder silica gel, granulating and vacuum drying.

[0043] Example 2

[0044] A pharmaceutical composition containing a Cistanche deserticola extract comprises raw materials including: 30 g of the Cistanche deserticola extract, 5 g of glutathione, 20 g of dopamine hydrochloride and 60 g of montmorillonite.

[0045] The Cistanche deserticola extract is prepared by the following specific steps: the dried Cistanche deserticola is crushed, passed through a 120-mesh sieve, added with 4 times the weight of ether and soaked for 10 hours, ultrasonically extracted at a temperature of 50°C for 2 hours with an ultrasonic frequency of 30kHz, filtered, ultrafiltered with an ultrafiltration membrane with a molecular weight cutoff of 5000Da, the filtrate is collected, concentrated at a temperature of 60°C to obtain an extract with a relative density of 1.18, and vacuum dried.

[0046] The method for preparing the pharmaceutical composition containing the Cistanche deserticola extract comprises the following steps:

[0047] S1. Disperse montmorillonite in 500 g of 2 mol / L hydrochloric acid, perform ultrasonic treatment for 30 min at an ultrasonic frequency of 15 kHz, centrifuge, wash with deionized water for 3 times, vacuum dry, add Cistanche deserticola extract and 400 g of water, perform ultrasonic treatment for 3 h at an ultrasonic frequency of 10 kHz, and obtain a loaded extract;

[0048] S2. Add dopamine hydrochloride and glutathione to the loaded extract, adjust the pH value of the system to 7.5-8.2, stir at a speed of 200 r / min for 2 hours, let stand for 4 hours, centrifuge, wash 3 times with deionized water, and vacuum dry.

[0049] A microbial preparation, whose raw materials include: 12g of the above-mentioned medicinal composition containing Cistanche deserticola extract, 2g of composite microorganisms, 20g of starch, and 2g of micropowder silica gel.

[0050] Among the composite microorganisms, the number of viable Bifidobacterium bifidum was 7×10 9cfu / g, the number of viable Lactobacillus plantarum was 7×10 9 cfu / g, and the number of viable bacteria of Butyrivibrio fibrinolyticus was 7×10 9 cfu / g, and the viable count of Enterococcus faecalis was 5.25×10 9 cfu / g.

[0051] The preparation method of the microbial preparation comprises the following steps: uniformly mixing the pharmaceutical composition containing the Cistanche deserticola extract, the composite microorganism and water, stirring at a speed of 200 r / min for 2 hours, adding starch and micro-powder silica gel, granulating and vacuum drying.

[0052] Example 3

[0053] A pharmaceutical composition containing a Cistanche deserticola extract comprises raw materials including: 15 g of Cistanche deserticola extract, 4 g of glutathione, 13 g of dopamine hydrochloride and 50 g of montmorillonite.

[0054] The Cistanche deserticola extract is prepared by the following specific steps: the dried Cistanche deserticola is crushed, passed through a 120-mesh sieve, added with 2.5 times the weight of ether and soaked for 9 hours, ultrasonically extracted at a temperature of 43°C for 100 minutes with an ultrasonic frequency of 21kHz, filtered, ultrafiltered with an ultrafiltration membrane with a molecular weight cutoff of 5000Da, the filtrate is collected, concentrated at a temperature of 58°C to obtain an extract with a relative density of 1.16, and vacuum dried.

[0055] The method for preparing the pharmaceutical composition containing the Cistanche deserticola extract comprises the following steps:

[0056] S1. Disperse montmorillonite in 400 g of 1.2 mol / L hydrochloric acid, perform ultrasonic treatment for 25 min at an ultrasonic frequency of 11 kHz, centrifuge, wash twice with deionized water, vacuum dry, add Cistanche deserticola extract and 350 g of water, perform ultrasonic treatment for 1.5 h at an ultrasonic frequency of 9 kHz, and obtain a loaded extract;

[0057] S2. Add dopamine hydrochloride and glutathione to the loaded extract, adjust the pH value of the system to 7.5-8.2, stir at a speed of 120 r / min for 100 min, let stand for 2.5 h, centrifuge, wash twice with deionized water, and vacuum dry.

[0058] A microbial preparation, whose raw materials include: 10g of the above-mentioned medicinal composition containing Cistanche deserticola extract, 1.3g of composite microorganisms, 18g of starch, and 1.3g of micropowder silica gel.

[0059] Among the composite microorganisms, the number of viable Bifidobacterium bifidum was 6×10 9 cfu / g, the number of viable Lactobacillus plantarum is 4×10 9 cfu / g, the number of viable bacteria of Butyrivibrio fibrinolyticus was 4×109 cfu / g, while the viable count of Enterococcus faecalis was 2.47×10 9 cfu / g.

[0060] The preparation method of the microbial preparation comprises the following steps: uniformly mixing the pharmaceutical composition containing the Cistanche deserticola extract, the composite microorganism and water, stirring at a speed of 100 r / min for 100 min, adding starch and micro-powder silica gel, granulating and vacuum drying.

[0061] Example 4

[0062] A pharmaceutical composition containing a Cistanche deserticola extract comprises raw materials including: 25g of Cistanche deserticola extract, 2g of glutathione, 17g of dopamine hydrochloride and 30g of montmorillonite.

[0063] The Cistanche deserticola extract is prepared by the following specific steps: the dried Cistanche deserticola is crushed, passed through a 120-mesh sieve, added with 3.5 times the weight of ether and soaked for 7 hours, ultrasonically extracted at a temperature of 47°C for 80 minutes with an ultrasonic frequency of 27kHz, filtered, ultrafiltered with an ultrafiltration membrane with a molecular weight cutoff of 5000Da, the filtrate is collected, concentrated at a temperature of 52°C to obtain an extract with a relative density of 1.17, and vacuum dried.

[0064] The method for preparing the pharmaceutical composition containing the Cistanche deserticola extract comprises the following steps:

[0065] S1. Disperse montmorillonite in 300 g of 1.8 mol / L hydrochloric acid, perform ultrasonic treatment for 15 min at an ultrasonic frequency of 13 kHz, centrifuge, wash twice with deionized water, vacuum dry, add Cistanche deserticola extract and 250 g of water, perform ultrasonic treatment for 2.5 h at an ultrasonic frequency of 6 kHz, and obtain a loaded extract;

[0066] S2. Add dopamine hydrochloride and glutathione to the loaded extract, adjust the pH value of the system to 7.5-8.2, stir at a speed of 180 r / min for 80 min, let stand for 3.5 h, centrifuge, wash twice with deionized water, and vacuum dry.

[0067] A microbial preparation, whose raw materials include: 6g of the above-mentioned medicinal composition containing Cistanche deserticola extract, 1.7g of composite microorganisms, 12g of starch, and 1.7g of micropowder silica gel.

[0068] Among the composite microorganisms, the number of viable Bifidobacterium bifidum was 5×10 9 cfu / g, the number of viable Lactobacillus plantarum was 6×10 9 cfu / g, the number of viable bacteria of Butyrivibrio fibrinolyticus was 3×10 9 cfu / g, while the viable count of Enterococcus faecalis was 2.47×10 9 cfu / g.

[0069] The preparation method of the microbial preparation comprises the following steps: uniformly mixing the pharmaceutical composition containing the Cistanche deserticola extract, the composite microorganism and water, stirring at a speed of 140 r / min for 80 min, adding starch and micro-powder silica gel, granulating and vacuum drying.

[0070] Example 5

[0071] A pharmaceutical composition containing a Cistanche deserticola extract comprises raw materials including: 20 g of the Cistanche deserticola extract, 3 g of glutathione, 15 g of dopamine hydrochloride and 40 g of montmorillonite.

[0072] The Cistanche deserticola extract is prepared by the following specific steps: the dried Cistanche deserticola is crushed, passed through a 120-mesh sieve, added with 3 times the weight of ether and soaked for 8 hours, ultrasonically extracted at a temperature of 45°C for 90 minutes with an ultrasonic frequency of 24kHz, filtered, ultrafiltered with an ultrafiltration membrane with a molecular weight cutoff of 5000Da, the filtrate is collected, concentrated at a temperature of 55°C to obtain an extract with a relative density of 1.16, and vacuum dried.

[0073] The method for preparing the pharmaceutical composition containing the Cistanche deserticola extract comprises the following steps:

[0074] S1. Disperse montmorillonite in 350 g of 1.5 mol / L hydrochloric acid, perform ultrasonic treatment for 20 min at an ultrasonic frequency of 12 kHz, centrifuge, wash twice with deionized water, vacuum dry, add Cistanche deserticola extract and 300 g of water, perform ultrasonic treatment for 2 h at an ultrasonic frequency of 7.5 kHz, and obtain a loaded extract;

[0075] S2. Add dopamine hydrochloride and glutathione to the loaded extract, adjust the pH value of the system to 7.5-8.2, stir at a speed of 150 r / min for 90 min, let stand for 3 h, centrifuge, wash twice with deionized water, and vacuum dry.

[0076] A microbial preparation, whose raw materials include: 8g of the above-mentioned medicinal composition containing Cistanche deserticola extract, 1.5g of composite microorganisms, 15g of starch, and 1.5g of micropowder silica gel.

[0077] Among the composite microorganisms, the number of viable Bifidobacterium bifidum was 5.5×10 9 cfu / g, the number of viable Lactobacillus plantarum is 5×10 9 cfu / g, and the number of viable bacteria of Butyrivibrio fibrinolyticus was 3.5×10 9 cfu / g, while the viable count of Enterococcus faecalis was 2.47×10 9 cfu / g.

[0078] The preparation method of the microbial preparation comprises the following steps: uniformly mixing the pharmaceutical composition containing the Cistanche deserticola extract, the composite microorganism and water, stirring at a speed of 100-140 r / min for 80-100 min, adding starch and micro-powder silica gel, granulating and vacuum drying.

[0079] Comparative Example 1

[0080] A microbial preparation, whose raw materials include: 8g of the pharmaceutical composition containing the Cistanche deserticola extract obtained in Example 5, 1.5g of composite microorganisms, 15g of starch, and 1.5g of micropowder silica gel.

[0081] Among the composite microorganisms, the number of viable Bifidobacterium bifidum was 5.5×10 9 cfu / g, the number of viable Lactobacillus plantarum is 5×10 9 cfu / g, and the number of viable bacteria of Butyrivibrio fibrinolyticus was 3.5×10 9 cfu / g, while the viable count of Enterococcus faecalis is 5×10 9 cfu / g.

[0082] The preparation method of the microbial preparation comprises the following steps: uniformly mixing the pharmaceutical composition containing the Cistanche deserticola extract, the composite microorganism and water, stirring at a speed of 100-140 r / min for 80-100 min, adding starch and micro-powder silica gel, granulating and vacuum drying.

[0083] Comparative Example 2

[0084] A pharmaceutical composition containing a Cistanche deserticola extract, the raw materials of which include: 20 g of the Cistanche deserticola extract obtained in Example 5, 15 g of dopamine hydrochloride, and 40 g of montmorillonite.

[0085] The method for preparing the pharmaceutical composition containing the Cistanche deserticola extract comprises the following steps:

[0086] S1. Disperse montmorillonite in 350 g of 1.5 mol / L hydrochloric acid, perform ultrasonic treatment for 20 min at an ultrasonic frequency of 12 kHz, centrifuge, wash twice with deionized water, vacuum dry, add Cistanche deserticola extract and 300 g of water, perform ultrasonic treatment for 2 h at an ultrasonic frequency of 7.5 kHz, and obtain a loaded extract;

[0087] S2. Add dopamine hydrochloride to the loaded extract, adjust the pH value of the system to 7.5-8.2, stir at a speed of 150 r / min for 90 min, let stand for 3 h, centrifuge, wash twice with deionized water, and vacuum dry.

[0088] A microbial preparation, whose raw materials include: 8g of the above-mentioned medicinal composition containing Cistanche deserticola extract, 1.5g of composite microorganisms, 0.31g of glutathione, 15g of starch, and 1.5g of micropowder silica gel.

[0089] Among the composite microorganisms, the number of viable Bifidobacterium bifidum was 5.5×10 9 cfu / g, the number of viable Lactobacillus plantarum is 5×10 9 cfu / g, and the number of viable bacteria of Butyrivibrio fibrinolyticus was 3.5×10 9 cfu / g, while the viable count of Enterococcus faecalis was 2.47×10 9 cfu / g.

[0090] The preparation method of the microbial preparation comprises the following steps: uniformly mixing the pharmaceutical composition containing the Cistanche deserticola extract, the composite microorganism, glutathione and water, stirring at a speed of 100-140 r / min for 80-100 min, adding starch and micro-powder silica gel, granulating and vacuum drying.

[0091] Comparative Example 3

[0092] A microbial preparation, whose raw materials include: 2.05g of the Cistanche extract obtained in Example 5, 0.31g of glutathione, 1.54g of dopamine hydrochloride, 4.1g of montmorillonite, 1.5g of composite microorganisms, 15g of starch, and 1.5g of micropowder silica gel.

[0093] Among the composite microorganisms, the number of viable Bifidobacterium bifidum was 5.5×10 9 cfu / g, the number of viable Lactobacillus plantarum is 5×10 9 cfu / g, and the number of viable bacteria of Butyrivibrio fibrinolyticus was 3.5×10 9 cfu / g, while the viable count of Enterococcus faecalis was 2.47×10 9 cfu / g.

[0094] The preparation method of the above-mentioned microbial preparation comprises the following steps:

[0095] (1) Add dopamine hydrochloride and glutathione to 300 g of water, adjust the pH value of the system to 7.5-8.2, stir at a speed of 150 r / min for 90 min, let stand for 3 h, centrifuge, wash twice with deionized water, and vacuum dry.

[0096] (2) The product obtained in step (1), montmorillonite, Cistanche deserticola extract, composite microorganisms and water are uniformly mixed, stirred at a speed of 100-140 r / min for 80-100 min, starch and micro-powder silica gel are added, granulated, and vacuum dried.

[0097] The pharmaceutical composition containing the Cistanche deserticola extract and the microbial preparation obtained in Example 5, and the microbial preparations obtained in Comparative Examples 1-3 were used to conduct mouse experiments, as follows:

[0098] Male C57BL / 6 mice aged 7-8 weeks (SPF grade, weight 22-24 g) were selected and kept at a temperature of 23±2°C, a humidity of 55±5%, a 12-h light-dark cycle, and free access to sterile water and food (using 60 Co irradiation sterilization). The experiment started after one week of adaptive feeding.

[0099] The mice were randomly divided into 7 groups (5 replicates in each group, 5 mice in each replicate), namely CK group (daily gavage with sterile water), MC group (daily gavage with sterile water), S5R group (daily gavage with the pharmaceutical composition containing Cistanche extract obtained in Example 5, 50 mg / kg), S5W group (daily gavage with the microbial preparation obtained in Example 5, 50 mg / kg), D1W group (daily gavage with the microbial preparation obtained in Comparative Example 1, 50 mg / kg), D2W group (daily gavage with the microbial preparation obtained in Comparative Example 2, 50 mg / kg), and D3W group (daily gavage with the microbial preparation obtained in Comparative Example 3, 50 mg / kg).

[0100] The MC group, S5R group, S5W group, D1W group, D2W group, and D3W group were intraperitoneally injected with probenecid (250 mg / kg mouse body weight) and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (25 mg / kg mouse body weight) every Wednesday and Sunday morning for 5 consecutive weeks before the experiment; the control group was injected with the above-mentioned volume of normal saline.

[0101] The body weight of each group of mice was continuously monitored, and it was found that there was no significant difference in the body weight of mice among the groups (P>0.05), indicating that the use of the modeling drug, the pharmaceutical composition containing the Cistanche extract and the microbial preparation obtained in Example 5, and the microbial preparation obtained in Comparative Examples 1-3 had no significant effect on the changes in mouse weight.

[0102] At the end of the 5th and 8th week of the experiment, the motor coordination ability of mice was measured by a mouse fatigue rotarod instrument. Each test can accommodate up to 6 mice, each mouse has an independent runway, and the computer software connected to the instrument can automatically record the time the mouse spends on the rod. A total of 7 increasing rotation speeds (8, 10, 12, 16, 20, 23, 25 r / min) were set, and the measurement time of each speed was 150 s. There was at least 10 minutes between each two rotation speeds to reduce the stress and fatigue of the mice.

[0103] The rotation speed was used as the abscissa and the time the animal stayed on the rotarod was used as the ordinate to plot the cumulative behavioral activity score of each animal and convert it into the area under the curve (AUC) for statistical analysis.

[0104] like Figure 1 As shown in the figure, the AUC of the S5W group was the highest, which was better than the other groups except the CK group (P < 0.05), and there was no significant difference with the CK group (P > 0.05). The AUC of the D1W group at week 5 was higher than that of the MC group (P < 0.05), while the AUC at week 8 was similar to that of the MC group, with no significant difference (P > 0.05). This may be because Enterococcus faecalis, as an opportunistic pathogen, has too high a viable count, which destroys the balance of intestinal flora.

[0105] The pole climbing test was conducted at the end of the fifth and eighth weeks of the experiment. The pole climbing test device was a 50 cm high, 1 cm diameter wooden stick fixed on a wooden base, with a 2.5 cm diameter ball on the top. The stick was wrapped with two layers of gauze to prevent the mouse from slipping during the crawling process. After the mice were adapted, they were placed on the top of the ball and the total time it took for the mouse to climb from the top of the pole to the bottom of the pole and land on its hind legs was recorded. The test was performed three times, with at least 10 minutes between each test. If the mouse paused or climbed in the opposite direction, it was retested.

[0106] like Figure 2 As shown, the total time of the CK group was the shortest; while the total time of the S5W group was the second shortest, which had no significant difference with the CK group (P>0.05), but was better than the other groups (P<0.05).

[0107] The total time of the S5R group was better than that of the MC group (P < 0.05), but was still inferior to those of the D1W group, the D2W group, and the D3W group. The total time of the D1W group in the 5th week was not significantly different from that of the S5R group (P > 0.05). The applicant believes that this may be due to the excessive number of live Enterococci, which destroyed the balance of intestinal flora.

[0108] After the end of the experiment, the mice were fasted but not watered for the first 12 hours. All mice were gavaged with 0.3 mL of 2.5% Evans blue solution (dissolved in 1.5% sodium carboxymethyl cellulose) 20 minutes before deep anesthesia with isoflurane. After deep anesthesia, the chest cavity was quickly opened, blood was collected from the heart, and the serum endotoxin content was detected; then the intact mouse brain tissue and colon tissue were quickly peeled off and fixed in 4% paraformaldehyde solution; finally, the small intestinal tissue from the pylorus to the upper end of the cecum was peeled off.

[0109] The mouse brain tissue was embedded in wax, and the substantia nigra and striatum were sliced ​​coronally with a thickness of 4 μm and stored at 4°C for later use. Five slices were selected from each tissue and each part for TH immunohistochemical staining, and then the expression of tyrosine hydroxylase (TH) was determined by enzyme-linked immunosorbent assay.

[0110] Since tyrosine hydroxylase is highly expressed in dopaminergic neurons, tyrosine hydroxylase-positive cells may represent dopaminergic neurons.

[0111] like Figure 3 As shown in the figure, compared with the other groups, the MC group had a large loss of dopaminergic neurons in the substantia nigra and a significant decrease in dopaminergic nerve fibers in the striatum; while the substantia nigra dopaminergic neurons and striatal dopaminergic nerve fibers in the S5R, S5W, D1W, D2W, and D3W groups were significantly increased compared with the MC group (P < 0.05).

[0112] The number of dopaminergic neurons in the substantia nigra and dopaminergic nerve fibers in the striatum of the S5W group was significantly increased compared with those of the S5R group, D1W group, D2W group, and D3W group (P<0.05).

[0113] Move the colon and small intestine tissues onto clean white paper, adjust them into a straight line respectively, do not stretch them to avoid unnecessary errors, and measure the length of the colon; use the same method to measure the distance from the gastric pylorus to the distal end of the Evans blue and the total length of the small intestine. The ratio of the two is the small intestine propulsion percentage of the Evans blue, that is, the small intestine conductivity.

[0114] Small intestinal conductivity and serum endotoxin content of each group Figure 4 As shown in the data, compared with other groups, the small intestinal conductivity of the MC group was significantly lower and the serum endotoxin was significantly increased; the serum endotoxin content of the CK group was the lowest, which was significantly better than those of the S5R group, D1W group, D2W group, and D3W group (P < 0.05), but there was no significant difference with the S5W group (P > 0.05); and the small intestinal conductivity of the S5W group was the highest, which was significantly better than those of the other groups (P < 0.05).

[0115] The applicant believes that the reason for the above results is that the present invention uses Cistanche deserticola as raw material, extracts by ether ultrasonic, filters, and concentrates, not only the extraction process is simple, the production cycle is short, and the process is stable, and the efficacy experiment proves that it has significant anti-inflammatory effect, stable efficacy, and high safety. The present invention also acidifies and erodes the natural montmorillonite layer structure and peels it off, not only the adsorption activity of Cistanche deserticola extract is high, but also polydopamine is further coated on the surface, which can effectively adhere to the polarity of microorganisms, effectively enhance the stability and anti-interference of microorganisms in vivo, and improve the load and protection performance of microorganisms. At the same time, the present invention uses Bifidobacterium bifidum, Lactobacillus plantarum, Butyrivibrio fibrinolyticus, and Enterococcus faecalis to compound and adhere to the polydopamine layer, and then cooperates with the Cistanche deserticola extract loaded in the polydopamine layer, which not only shows a significant anti-inflammatory effect, but also has an anti-inflammatory effect that is better than using the Cistanche deserticola extract or the composite microorganism alone, especially the best effect of the synergistic self-oxidation coating of dopamine hydrochloride and glutathione, which can not only improve gastrointestinal dysfunction, but also effectively alleviate Parkinson's symptoms, with a significant effect.

[0116] The above description is only a preferred specific implementation manner of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can make equivalent replacements or changes according to the technical scheme and inventive concept of the present invention within the technical scope disclosed by the present invention, which should be covered by the protection scope of the present invention.

Claims

1. A pharmaceutical composition containing a Cistanche deserticola extract, characterized in that: The raw materials include, by weight: 1-3 parts of Cistanche deserticola extract, 0.1-0.5 parts of glutathione, 1-2 parts of dopamine hydrochloride and 2-6 parts of montmorillonite.

2. The pharmaceutical composition containing the Cistanche deserticola extract according to claim 1, characterized in that: The Cistanche deserticola extract is prepared by the following specific steps: crushing the dried Cistanche deserticola, sieving, adding ether to immerse for 5-10 hours, ultrasonic extraction at 40-50°C for 1-2 hours, filtering, ultrafiltration, concentrating the filtrate to a relative density of 1.15-1.18, and vacuum drying.

3. The pharmaceutical composition containing the Cistanche deserticola extract according to claim 2, characterized in that: The mass ratio of Cistanche deserticola to ether is 1:2-4; the ultrasonic frequency is 20-30kHz.

4. The pharmaceutical composition containing the Cistanche deserticola extract according to claim 2, characterized in that: Ultrafiltration was performed using an ultrafiltration membrane with a molecular weight cutoff of 5000 Da.

5. A method for preparing a pharmaceutical composition containing a Cistanche deserticola extract according to any one of claims 1 to 4, characterized in that: The steps include: S1, dispersing montmorillonite in hydrochloric acid and ultrasonically treating it for 10-30 minutes, centrifuging, washing, vacuum drying, adding Cistanche deserticola extract and water and ultrasonically treating it for 1-3 hours to obtain a loaded extract; S2. Add dopamine hydrochloride and glutathione to the loaded extract, adjust the pH value of the system to 7.5-8.2, stir for 1-2 hours, let stand for 2-4 hours, centrifuge, wash, and vacuum dry.

6. The pharmaceutical composition containing the Cistanche deserticola extract according to claim 5, characterized in that: In S1, the concentration of hydrochloric acid is 1-2 mol / L, and the frequency of ultrasonic treatment after montmorillonite is dispersed in hydrochloric acid is 10-15 kHz; In S1, the frequency of ultrasonic treatment after adding Cistanche deserticola extract and water is 5-10 kHz.

7. A microbial preparation, characterized in that: The raw materials include, by mass: 5-12 parts of the pharmaceutical composition containing the Cistanche deserticola extract as claimed in any one of claims 1 to 4, 1-2 parts of composite microorganisms, 10-20 parts of starch, and 1-2 parts of micropowder silica gel.

8. The microbial preparation according to claim 7, characterized in that: The composite microorganisms include: Bifidobacterium bifidum, Lactobacillus plantarum, Butyrivibrio fibrisolvens, Enterococcus faecalis; The viable counts of Bifidobacterium bifidum, Lactobacillus plantarum and Butyrivibrio fibrinolyticus in the composite microorganisms were 2-7×10 9 cfu / g, and the number of live Enterococcus faecalis in the composite microorganisms accounts for 10-20% of the total live number of the composite microorganisms.

9. A method for preparing the microbial preparation according to claim 7 or 8, characterized in that: The method comprises the following steps: uniformly mixing the pharmaceutical composition containing the Cistanche deserticola extract as claimed in any one of claims 1 to 4, composite microorganisms and water, adding starch and micro-powder silica gel, granulating and vacuum drying.

10. Use of the pharmaceutical composition containing the Cistanche deserticola extract according to any one of claims 1 to 4, or / and the microbial preparation according to claim 7 or 8 in the preparation of a drug for treating Parkinson's disease.

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