Kidney-tonifying and blood-activating decoction modified formula

Abstract

The invention relates to the technical field of traditional Chinese medicines, in particular to a kidney-tonifying and blood-activating decoction modified prescription which is prepared from the following components in parts by mass: 10-20 parts of prepared rehmannia root, 10-20 parts of salted semen cuscutae, 5-15 parts of salted fructus psoraleae, 10-20 parts of fructus lycii, 5-15 parts of salted eucommia ulmoides, 5-15 parts of wine-prepared fructus corni, 5-15 parts of angelica sinensis, 5-15 parts of radix angelicae pubescentis, 3-9 parts of safflower carthamus, 14-16 parts of honey-fried astragalus membranaceus, 9-11 parts of roasted rhizoma atractylodis macrocephalae and 5-7 parts of honey-fried licorice root. The decoction process comprises the following steps: adding water to decoct twice, adding water of which the mass is 6 times of that of the kidney-tonifying and blood-activating decoction for the first time, soaking for 30 minutes, decocting for 1 hour, and filtering; adding water in an amount which is 4 times of the mass of the kidney-tonifying and blood-activating decoction for the second time, decocting for 45 minutes, and filtering; combining the two filtrates; the concentrating and drying process comprises the following steps: concentrating under reduced pressure, and then drying at 80 DEG C. Compared with an ancient kidney-tonifying and blood-activating decoction, the decoction process and the concentration drying process provided by the invention better meet the requirements of modern industrial production, and meanwhile, compared with the ancient kidney-tonifying and blood-activating decoction, the decoction has a better bone formation promoting effect.

Description

Technical Field

[0001] The invention relates to the technical field of traditional Chinese medicines, in particular to a modified recipe of Bushen Huoxue Decoction. Background Art

[0002] Bushen Huoxue Decoction (also known as "Bushen Huoxue Recipe") comes from the Qing Dynasty medical book "Shangke Dacheng". It is a classic Chinese medicine prescription that has the effects of nourishing the kidneys and strengthening the tendons, promoting blood circulation and relieving pain.

[0003] The existing public document "Observation on the Clinical Efficacy of Bushen Jiangu Prescription in the Treatment of Osteoporotic Thoracolumbar Compression Fractures" (Jilin Traditional Chinese Medicine, Vol. 43, No. 12, December 2023, 1415-1419) discloses a regimen of Bushen Huoxue Decoction combined with vertebroplasty for the treatment of osteoporotic thoracolumbar compression fractures. This regimen can improve the patient's treatment efficacy, relieve pain VAS, improve bone density, increase postoperative vertebral height, and reduce kyphosis angle. Patients benefit more than surgical treatment alone.

[0004] However, the proposal uses the ancient recipe of Bushen Huoxue Decoction from the Qing Dynasty medical book "Compendium of Traumatology". The ancient recipe of Bushen Huoxue Decoction is a traditional decoction with a relatively complicated decoction method, which does not meet the needs of modern industrial production. At the same time, the public documents have not conducted further research on the formula components of the ancient recipe of Bushen Huoxue Decoction. Summary of the invention

[0005] The object of the present invention is to provide a modified recipe of Bushen Huoxue Decoction to solve the problems raised in the above background technology.

[0006] To achieve the above object, the present invention provides the following technical scheme: a modified recipe of Bushen Huoxue Decoction, which is composed of 10-20 parts of Rehmannia glutinosa, 10-20 parts of salt Cuscuta seeds, 5-15 parts of salt Psoralea corylifolia, 10-20 parts of Lycium barbarum, 5-15 parts of salt Eucommia ulmoides, 5-15 parts of wine cornus fruit, 5-15 parts of Angelica sinensis, 5-15 parts of Angelica dahurica, 3-9 parts of safflower, 14-16 parts of honey-fried Astragalus, 9-11 parts of stir-fried Atractylodes macrocephala and 5-7 parts of honey-fried Licorice root, by weight;

[0007] The decoction process of the modified Bushen Huoxue Decoction is as follows: add water and decoct twice, add water 6 times the mass of the modified Bushen Huoxue Decoction for the first time, soak for 30 minutes, decoct for 1 hour, and filter; add water 4 times the mass of the modified Bushen Huoxue Decoction for the second time, decoct for 45 minutes, and filter; combine the two filtrates;

[0008] The concentration and drying process of the modified Bushen Huoxue Decoction is as follows: concentration under reduced pressure and then drying at 80°C.

[0009] Preferably, a modified recipe of Bushen Huoxue Decoction is composed, by mass, of 15 parts of Rehmannia glutinosa, 15 parts of salt Cuscuta seeds, 10 parts of salt Psoralea corylifolia, 15 parts of Lycium barbarum, 10 parts of salt Eucommia ulmoides, 10 parts of wine Cornus officinalis, 10 parts of Angelica sinensis, 10 parts of Angelica dahurica, 6 parts of Carthamus tinctorius, 14-16 parts of honey-roasted Astragalus membranaceus, 9-11 parts of stir-fried Atractylodes macrocephala, and 5-7 parts of honey-roasted Licorice.

[0010] Preferably, a modified recipe of Bushen Huoxue Decoction is composed, by mass, of 15 parts of Rehmannia glutinosa, 15 parts of salt Cuscuta seeds, 10 parts of salt Psoralea corylifolia, 15 parts of Lycium barbarum, 10 parts of salt Eucommia ulmoides, 10 parts of wine Cornus officinalis, 10 parts of Angelica sinensis, 10 parts of Angelica dahurica, 6 parts of Carthamus tinctorius, 15 parts of honey-roasted Astragalus membranaceus, 9-11 parts of stir-fried Atractylodes macrocephala, and 5-7 parts of honey-roasted Licorice.

[0011] Preferably, a modified recipe of Bushen Huoxue Decoction is composed, by mass, of 15 parts of Rehmannia glutinosa, 15 parts of salt Cuscuta seeds, 10 parts of salt Psoralea corylifolia, 15 parts of Lycium barbarum, 10 parts of salt Eucommia ulmoides, 10 parts of wine Cornus officinalis, 10 parts of Angelica sinensis, 10 parts of Angelica dahurica, 6 parts of Safflower, 15 parts of honey-roasted Astragalus, 10 parts of stir-fried Atractylodes macrocephala and 6 parts of honey-roasted Licorice.

[0012] The present invention provides a modified recipe of Bushen Huoxue Decoction, which has the following beneficial effects:

[0013] 1. According to experiments, the Bushen Huoxue Decoction provided by the present invention is subjected to the following decoction process: adding water and decocting twice, adding water 6 times the mass of the Bushen Huoxue Decoction for the first time, soaking for 30 minutes, decocting for 1 hour, and filtering; adding water 4 times the mass of the Bushen Huoxue Decoction for the second time, decocting for 45 minutes, and filtering; combining the two filtrates, the hyperoside transfer rate and the paste yield of this process are closest to the results of the traditional decoction method, and the filtrate obtained by decoction is concentrated under reduced pressure and dried (80°C), and the hyperoside transfer rate is higher than that of normal pressure concentration, which is more in line with the requirements of industrial production.

[0014] 2. According to the experiment, the bone volume / total volume (BV / TV), cancellous bone BMD, trabecular thickness (Tb.Th), and trabecular number (Tb.N) indicators of mice in the Bushen Huoxue Decoction modified prescription group were significantly increased, while the trabecular separation (Tb.Sp) was significantly decreased, and was close to the value of the positive drug group. It can be seen that the Bushen Huoxue Decoction modified prescription provided by the present invention can improve the bone mass reduction in estrogen deficiency model rats.

[0015] 3. The ALP activity test results show that the modified Bushen Huoxue Decoction provided by the present invention can effectively increase the ALP activity of femoral tissue.

[0016] 4. Detect the protein expression level in femoral tissues by Western Blot to evaluate the level of osteoblast-specific proteins. In the femoral tissues of the modified Bushen Huoxue Decoction group provided by the present invention, the protein expressions of Runx2 and ALP were significantly increased (P<0.05).

[0017] 5. Observe the effects of the Bushen Huoxue Decoction and the medicated serum containing the modified Bushen Huoxue Decoction provided by the present invention on the osteogenic differentiation of MC3T3 cells. The medicated serum containing the modified Bushen Huoxue Decoction showed better effects. MC3T3 cells were cultured with the medicated serum containing the Bushen Huoxue Decoction and the medicated serum containing the modified Bushen Huoxue Decoction provided by the present invention respectively. After 14 days, the alizarin red staining results showed that the modified Bushen Huoxue Decoction provided by the present invention further promoted the differentiation of MC3T3 cells into osteoblasts compared with the Bushen Huoxue Decoction.

[0018] 6. Make slight changes based on the original Bushen Huoxue Decoction, subtract myrrh, and add two spleen-strengthening drugs, astragalus membranaceus and atractylodes macrocephala. Licorice is used to relieve pain and harmonize various drugs. The specific formula is as follows: 15g of dodder seed (salt-fried), 15g of rehmannia root (prepared), 15g of wolfberry fruit, 10g of psoralea corylifolia (salt-fried), 10g of eucommia ulmoides (salt-fried), 10g of cornel fruit (wine-fried), 10g of angelica sinensis, 10g of angelica pubescens, 6g of safflower, 15g of astragalus membranaceus (honey-fried), 10g of atractylodes macrocephala (stir-fried), 6g of licorice (honey-fried). In the formula, dodder seed (salt-fried) is used to tonify the kidney and replenish essence as the monarch drug. Rehmannia root (prepared), wolfberry fruit, psoralea corylifolia (salt-fried), eucommia ulmoides (salt-fried), and cornel fruit (wine-fried) assist the monarch drug to strengthen the effect of tonifying the kidney and have the effect of strengthening tendons and bones as the minister drugs. Angelica sinensis, angelica pubescens, and safflower promote blood circulation to remove stasis and dredge collaterals to relieve pain. Astragalus membranaceus and atractylodes macrocephala replenish qi and strengthen the spleen as the assistant drugs. Licorice (honey-fried) harmonizes various drugs as the envoy drug. The combination of various drugs has the effects of tonifying the kidney, strengthening the spleen, and promoting blood circulation. Description of the Drawings

[0019] Figure 1 It is the chromatogram of the hyperoside reference substance solution in the present invention;

[0020] Figure 2 It is the chromatogram of the test solution in the present invention;

[0021] Figure 3 It is the chromatogram of the negative sample solution in the present invention;

[0022] Figure 4 It is the Micro-CT detection image of the femoral tissues of rats in five experimental groups;

[0023] Figure 5 It is the ALP activity detection image of the femoral tissues of rats in five experimental groups;

[0024] Figure 6 It is the detection image of the expression of osteoblast-specific proteins in the femoral tissues of rats in five experimental groups;

[0025] Figure 7Experimental figure for testing the ability of Bushen Huoxue Decoction group and modified Bushen Huoxue Decoction group to promote the differentiation of MC3T3 cells into osteoblasts. Detailed implementation manners

[0026] The following will clearly and completely describe the technical solutions in the embodiments of the present invention with reference to the accompanying drawings in the embodiments of the present invention.

[0027] 1.1 Sources of raw decoction pieces

[0028] Prepared Rehmannia Root (manufacturer - Bozhou Yonggang Decoction Pieces Factory Co., Ltd., origin - Jiaozuo, Henan, batch number - A230918), Semen Cuscutae (manufacturer - Bozhou Yonggang Decoction Pieces Factory Co., Ltd., origin - Bayannur, Inner Mongolia, batch number - A231024), Fructus Psoraleae (manufacturer - Bozhou Huqiao Pharmaceutical Co., Ltd., origin - Yunnan, batch number - 2308310612), Lycium Barbarum Fruit (manufacturer - Bozhou Huqiao Pharmaceutical Co., Ltd., origin - Ningxia, batch number - 2309080082), Eucommia ulmoides Oliv. (manufacturer - Bozhou Huqiao Pharmaceutical Co., Ltd., origin - Hubei, batch number - 23008040192), Fructus Corni (manufacturer - Bozhou Huqiao Pharmaceutical Co., Ltd., origin - Henan, batch number - 2305070222), Angelica sinensis (manufacturer - Bozhou Huqiao Pharmaceutical Co., Ltd., origin - Gansu, batch number - 2307020332), Heracleum hemsleyanum Diels (manufacturer - Bozhou Yonggang Decoction Pieces Factory Co., Ltd., origin - Chongqing, batch number - A230921), Carthamus tinctorius L. (manufacturer - Bozhou Huqiao Pharmaceutical Co., Ltd., origin - Xinjiang, batch number - 2309170012), Astragalus membranaceus (manufacturer - Zhejiang Zhenyuantang Traditional Chinese Medicine Decoction Pieces Co., Ltd., origin - Inner Mongolia, batch number - 20230701), Atractylodes macrocephala Koidz. (manufacturer - Bozhou Huqiao Pharmaceutical Co., Ltd., origin - Anhui, batch number - 2307170122), Glycyrrhiza uralensis Fisch. (manufacturer - Bozhou Yonggang Decoction Pieces Factory Co., Ltd., origin - Zhangye, Gansu, batch number - A230929).

[0029] 1.2 Research on extraction process

[0030] 1.2.1 Selection of extraction process

[0031] According to the "Traditional Chinese Medicine Law of the People's Republic of China", the "Drug Administration Law of the People's Republic of China", the "Announcement of the General Administration on the Implementation of Record-filing Management for Traditional Chinese Medicine Preparations Prepared by Medical Institutions Using Traditional Processes (No. 19, 2018)", and the "Implementation Rules for the Record-filing Management of Traditional Chinese Medicine Preparations Prepared by Medical Institutions in Jiangsu Province Using Traditional Processes (Trial)", based on the actual situation of the clinical use of this prescription, it is planned to conduct a study on the water decoction process of the traditional Chinese medicine decoction pieces components (prepared rehmannia root, semen cuscutae with salt, fructus psoraleae with salt, wolfberry fruit, eucommia ulmoides with salt, cornel fruit with wine, angelica sinensis, pubescent angelica root, safflower, astragalus membranaceus with honey, stir-fried atractylodes macrocephala, and licorice root with honey) in a modified recipe of Bushen Huoxue Decoction provided by the present invention with the transfer rate and extract yield of the index components as indicators.

[0032] 1.2.2 Selection of index components

[0033] In this prescription, semen cuscutae with salt is the monarch drug, which has the effects of tonifying the liver and kidney, consolidating essence and reducing urination, preventing miscarriage, improving eyesight, and stopping diarrhea; externally, it can dispel wind and remove freckles. It is used for liver and kidney deficiency, weakness of the waist and knees, impotence and spermatorrhea, enuresis and frequent urination, threatened abortion due to kidney deficiency, restlessness of the fetus in pregnancy, dizziness and tinnitus, diarrhea due to spleen and kidney deficiency; externally, it is used for vitiligo. In the "Semen Cuscutae" item in Part I of the Chinese Pharmacopoeia 2020 Edition, the content determination is carried out with hyperin. Therefore, hyperin is selected as the index component in this study.

[0034] 1.2.3 Study on the content determination of hyperin

[0035] 1.2.3.1 Instruments and materials

[0036] 1.2.3.1.1 Instruments

[0037] Instruments and materials: Agilent 1260 type high performance liquid chromatograph, Agilent 1260 vwd detector, Agilent Chem-Station chromatographic workstation (Agilent Company, USA); QUINTIX35-1CN electronic balance (sartorius); KH-500DE type ultrasonic cleaner (Kunshan Ultrasonic Instrument Co., Ltd.); DHG-9051A hot air circulation oven (Shanghai Linyiheng Scientific Instrument Co., Ltd.).

[0038] 1.2.3.1.2 Test drugs

[0039] Hyperin reference substance (National Institutes for Food and Drug Control, batch number 111521-202310, content calculated as 94.7%), acetonitrile (Werck Company, Germany), and water is ultrapure water.

[0040] 1.2.3.2 Methods and results

[0041] 1.2.3.2.1 Chromatographic conditions

[0042] Using octadecylsilane-bonded silica gel as the filler and Shim-pack GIST (4.6×250mm, 5μm) as the chromatographic column; using acetonitrile-0.1% phosphoric acid solution (17:83) as the mobile phase; the detection wavelength is 360nm.

[0043] 1.2.3.2.2 Preparation of reference substance solution

[0044] Weigh an appropriate amount of hyperoside reference substance accurately, dissolve it in 80% ethanol to prepare a solution containing 102.28μg of hyperoside per mL, and that's it.

[0045] 1.2.3.2.3 Preparation of test sample solution

[0046] Take a dose of the cut crude drugs of the modified Bushen Huoxue Decoction provided by the present invention, decoct it twice with water. Each time, add 6 times the mass (800mL) of water of a dose of the cut crude drugs of the modified Bushen Huoxue Decoction, soak for 30min, decoct for 30min, and filter; for the second time, add 6 times the mass (530mL) of water of a dose of the cut crude drugs of the modified Bushen Huoxue Decoction, decoct for 25min, filter and combine the filtrates, concentrate to less than 400mL, let it cool, and add water to 400mL. Accurately measure 5mL of the above solution, place it in a 10mL volumetric flask, dilute it to the mark with 80% ethanol, shake well, and filter through a 0.45μm filter membrane to obtain the test sample solution.

[0047] 1.2.3.2.4 Preparation of negative sample solution

[0048] According to the preparation method of the test sample solution, prepare the negative sample solution of Cuscuta chinensis Lam. cut crude drugs in the same way.

[0049] 1.2.3.2.5 Specificity investigation

[0050] Accurately measure 10μL of the reference substance solution, the test sample solution and the negative sample solution respectively, and determine according to the chromatographic conditions under item "1.2.3.2.1". From Figure 1 , Figure 2 , Figure 3 It can be seen that the negative sample has no interference at the main peak position, and the resolution between hyperoside and adjacent impurity peaks is greater than 1.5.

[0051] 1.2.3.2.6 Linear relationship investigation

[0052] Accurately weigh an appropriate amount of hyperin reference substance (with a content of 94.7%), place it in a 100 mL volumetric flask, add 80% ethanol to dissolve and dilute to the mark to prepare reference substance solutions with hyperin concentrations of 3.20 μg / mL, 6.39 μg / mL, 12.78 μg / mL, 25.57 μg / mL, 51.14 μg / mL, and 102.28 μg / mL respectively. According to the above chromatographic conditions, accurately pipette 10 μL of the reference substance solution and inject it into the liquid chromatograph. Using the injection volume of hyperin as the abscissa and the peak area as the ordinate, plot the standard curve, and obtain the regression equation as follows:

[0053] y = 2003.42549x - 1.50249, R 2 = 0.99996

[0054] It shows that there is a good linear relationship between the injection volume of hyperin within the range of 0.032 - 1.0228 μg and the peak area.

[0055] 1.2.3.2.7 Precision investigation

[0056] Accurately pipette 10 μL of the reference substance solution, determine it according to the chromatographic conditions under item "2.3.2.1", inject samples repeatedly 6 times, measure the peak area, and the result shows that the RSD% is 0.33% (n = 6), indicating good precision of the instrument.

[0057] 1.2.3.2.8 Repeatability investigation

[0058] For the same decoction solution prepared according to the method in "1.2.3.2.3", divide it into 6 parallel portions, accurately pipette 10 μL from each portion, determine it according to the chromatographic conditions under item "1.2.3.2.1", inject samples repeatedly 6 times, measure the peak area, and the result shows that the RSD% is 1.04%, indicating good repeatability of this method.

[0059] 1.2.3.2.9 Stability investigation

[0060] Accurately pipette 10 μL of the same test solution, determine the peak area according to the chromatographic conditions under item "1.2.3.2.1" at 0, 2, 4, 6, 8, 10, 12, 16, 20, 24, and 36 h respectively. The result shows that the RSD of the peak area of hyperin is 0.37%, indicating that the test solution is stable within 24 h.

[0061] 1.2.3.2.10 Spike recovery test

[0062] Take the decoction with known hyperoside content, accurately pipette 5 mL into a 10-mL volumetric flask, in duplicate for 9 portions. Then pipette 1 mL of the reference substance solution with a hyperoside concentration of 0.09576 mg / mL into the same volumetric flask, dilute to the mark with 70% ethanol, shake well, filter through a 0.45-μm filter membrane, in duplicate for 6 portions, and determine under the chromatographic conditions described in "2.3.2.1". The results are shown in Table 1.

[0063] Table 1: Results of the recovery test with added samples (n = 6)

[0064]

[0065]

[0066] 1.2.5 Study the transfer rate and extract yield of hyperoside by the traditional clinical decoction method

[0067] The modified Bushen Huoxue Decoction provided by the present invention is based on the traditional decoction used clinically. To enable the preparation to carry the safety and effectiveness of the traditional decoction, it is necessary to bridge with the traditional decoction method and study the extraction process. The components of traditional Chinese medicine are complex. Except for the index components, the transfer of components is affected by the extract yield, which is an important index for evaluating the extraction process. The transfer rate and extract yield of hyperoside were investigated by the traditional clinical decoction method, and the extraction process was further studied based on this.

[0068] 1.2.5.1 Study on the transfer rate and extract yield of hyperoside by the traditional decoction method

[0069] According to the clinical use characteristics of this formula and the "Administrative Specification for Traditional Chinese Medicine Decoction Rooms in Medical Institutions", the decoction method for this traditional decoction is as follows: Decoct the six kinds of cut crude drugs in proportion according to the prescription twice. For the first time, add 930 mL (7 times) of water, soak for 30 min, decoct for 1 h, and filter; for the second time, add 660 mL (5 times) of water, decoct for 45 min, and filter. Combine the filtrates of the two times, cool, measure the volume of the decoction, and conduct 9 parallel tests.

[0070] Transfer rate of hyperoside = W / M × 100%;

[0071] where W represents the mass of hyperoside in the traditional decoction, and M represents the mass of hyperoside in the cut crude drugs. The results are shown in Table 2.

[0072] Extract yield = W 干浸膏质量 / M 饮片质量 × 100%, and the results are shown in Table 3.

[0073] Table 2: Results of the transfer rate of hyperoside by the traditional decoction method (%)

[0074]

[0075]

[0076] Table 3: Results of extract yield by traditional decocting method (%)

[0077]

[0078] 1.2.6 Study on water decocting process

[0079] Fix the soaking time at 30 min, the number of decocting times at 2 times, the first decocting time at 1 h, and the second decocting time at 45 min. Taking the multiple of water addition as a single factor, set 3 levels: 6 times, 7 times, and 8 times for the first time; 4 times, 5 times, and 6 times for the second time. Prepare two parallel samples and examine the results of hyperoside transfer rate (%) as shown in Table 4 and extract yield (%) as shown in Table 5.

[0080] Table 4: Results of berberine hydrochloride transfer rate with different multiples of water addition (%)

[0081]

[0082]

[0083] Table 5: Results of extract yield with different multiples of water addition (%)

[0084]

[0085]

[0086] According to the test results, decoct with water twice. For the first time, add water with a mass 6 times that of the modified Bushen Huoxue Decoction, soak for 30 min, decoct for 1 h, and filter; for the second time, add water with a mass 4 times that of the modified Bushen Huoxue Decoction, decoct for 45 min, and filter; combine the filtrates of the two times. The hyperoside transfer rate and extract yield of this process are closest to those of the traditional decocting method.

[0087] 1.2.8 Conclusion

[0088] Through the study on the hyperoside transfer rate (%) and extract yield (%) of the traditional decocting method, and based on this, study the corresponding water decocting process. Finally, the extraction process of this preparation is determined as: decoct with water twice. For the first time, add water with a mass 6 times that of the modified Bushen Huoxue Decoction, soak for 30 min, decoct for 1 h, and filter; for the second time, add water with a mass 4 times that of the modified Bushen Huoxue Decoction, decoct for 45 min, and filter; combine the filtrates of the two times.

[0089] 1.3. Investigate the influence of temperature on concentration and drying processes

[0090] Weigh one dose of the decoction pieces of the modified formula of Bushen Huoxue Decoction provided by the present invention, and decoct it twice with water. For the first time, add water with a mass 6 times that of the modified formula of Bushen Huoxue Decoction, soak for 30 minutes, decoct for 1 hour, and filter; for the second time, add water with a mass 4 times that of the modified formula of Bushen Huoxue Decoction, decoct for 45 minutes, and filter; combine the filtrates of the two times. Parallel 9 portions and combine the filtrates of 9 portions. The sum of the volumes of the 9 filtrates / 9 is the volume of the decoction of each dose of decoction pieces. Take two portions according to the volume of the decoction of each dose of decoction pieces, and concentrate each portion to about 200 mL under normal pressure and wash it out with water to 400 mL; take another two portions according to the volume of the decoction of each dose of decoction pieces, and concentrate each portion to 200 mL under reduced pressure at 80 °C and wash it out with water to 400 mL. Calculate the transfer rate of hyperoside as shown in Table 6.

[0091] Table 6: Transfer rate of hyperoside by different concentration methods (%)

[0092]

[0093] As can be seen from Table 6, the transfer rate of hyperoside after concentration under reduced pressure and drying (80 °C) is higher than that under normal pressure concentration, and it also better meets the requirements of industrial production.

[0094] Example 1

[0095] A modified formula of Bushen Huoxue Decoction, calculated by mass parts, is composed of 10 parts of Rehmannia glutinosa, 10 parts of Cuscuta chinensis with salt, 5 parts of Psoralea corylifolia with salt, 10 parts of Lycium barbarum, 5 parts of Eucommia ulmoides with salt, 5 parts of Cornus officinalis with wine, 5 parts of Angelica sinensis, 5 parts of Heracleum hemsleyanum, 3 parts of Carthamus tinctorius, 14 parts of Astragalus membranaceus with honey, 9 parts of Atractylodes macrocephala stir-fried, and 5 parts of Glycyrrhiza uralensis with honey.

[0096] The decocting process of the modified formula of Bushen Huoxue Decoction is: decoct twice with water. For the first time, add water with a mass 6 times that of the modified formula of Bushen Huoxue Decoction, soak for 30 minutes, decoct for 1 hour, and filter; for the second time, add water with a mass 4 times that of the modified formula of Bushen Huoxue Decoction, decoct for 45 minutes, and filter; combine the filtrates of the two times;

[0097] The concentration and drying process of the modified formula of Bushen Huoxue Decoction is: conduct drying at 80 °C after concentration under reduced pressure.

[0098] Example 2

[0099] A modified formula of Bushen Huoxue Decoction, calculated by mass parts, is composed of 20 parts of Rehmannia glutinosa, 20 parts of Cuscuta chinensis with salt, 15 parts of Psoralea corylifolia with salt, 20 parts of Lycium barbarum, 15 parts of Eucommia ulmoides with salt, 15 parts of Cornus officinalis with wine, 15 parts of Angelica sinensis, 15 parts of Heracleum hemsleyanum, 9 parts of Carthamus tinctorius, 16 parts of Astragalus membranaceus with honey, 11 parts of Atractylodes macrocephala stir-fried, and 7 parts of Glycyrrhiza uralensis with honey.

[0100] The decoction process of the modified Bushen Huoxue Decoction is as follows: add water and decoct twice, add water 6 times the mass of the modified Bushen Huoxue Decoction for the first time, soak for 30 minutes, decoct for 1 hour, and filter; add water 4 times the mass of the modified Bushen Huoxue Decoction for the second time, decoct for 45 minutes, and filter; combine the two filtrates;

[0101] The concentration and drying process of the modified Bushen Huoxue Decoction is as follows: concentration under reduced pressure and then drying at 80°C.

[0102] Example 3

[0103] A modified recipe of Bushen Huoxue Decoction, comprising, by weight, 15 parts of Rehmannia glutinosa, 15 parts of salt Cuscuta seeds, 10 parts of salt Psoralea corylifolia, 15 parts of Lycium barbarum, 10 parts of salt Eucommia ulmoides, 10 parts of wine-brewed Cornus officinalis, 10 parts of Angelica sinensis, 10 parts of Angelica dahurica, 6 parts of Carthamus tinctorius, 15 parts of honey-fried Astragalus membranaceus, 10 parts of stir-fried Atractylodes macrocephala, and 6 parts of honey-fried Licorice root;

[0104] The decoction process of the modified Bushen Huoxue Decoction is as follows: add water and decoct twice, add water 6 times the mass of the modified Bushen Huoxue Decoction for the first time, soak for 30 minutes, decoct for 1 hour, and filter; add water 4 times the mass of the modified Bushen Huoxue Decoction for the second time, decoct for 45 minutes, and filter; combine the two filtrates;

[0105] The concentration and drying process of the modified Bushen Huoxue Decoction is as follows: concentration under reduced pressure and then drying at 80°C.

[0106] 12-week-old female SD rats were selected and underwent ovariectomy (OVX). One week after surgery, oral administration began, including:

[0107] The sham operation group and the modeling group were gavaged with an equal amount of normal saline;

[0108] The positive drug group was given alendronate sodium (2.5 mg / kg) by gavage;

[0109] The Bushen Huoxue Decoction group was given Bushen Huoxue Decoction (20 g / kg) by gavage; the Bushen Huoxue Decoction, by weight, consisted of 15 parts of Rehmannia glutinosa, 9 parts of Cuscuta australis, 9 parts of Psoralea corylifolia, 3 parts of Lycium barbarum, 3 parts of Eucommia ulmoides, 3 parts of Cornus officinalis, 3 parts of Angelica sinensis, 3 parts of Angelica dahurica, 3 parts of Herba Angelicae Pubescentis, 3 parts of Cistanche deserticola, 3 parts of Myrrha and 1.5 parts of Carthamus tinctorius;

[0110] The modified Bushen Huoxue Decoction group was given the modified Bushen Huoxue Decoction (20 g / kg) provided in Example 2 of the present invention by intragastric administration;

[0111] The drugs were administered continuously for 12 weeks. Blood was collected from each group 24 hours after the last administration, and the rats were killed to obtain bone tissue for Micro-CT examination.

[0112] See also Figure 4, Micro-CT examination results showed that compared with the sham operation group, osteoporosis occurred in the femurs of the model rats, and an osteoporosis model induced by estrogen deficiency was successfully established.

[0113] Compared with the model group, the indexes of bone volume / total volume (BV / TV), cancellous bone BMD, trabecular bone thickness (Tb.Th), and trabecular bone number (Tb.N) in the mice of the modified Bushen Huoxue Decoction group were significantly up-regulated, while the trabecular bone separation degree (Tb.Sp) was significantly decreased, and the values were close to those of the positive drug group.

[0114] These results all suggested that the modified Bushen Huoxue Decoction provided by the present invention could improve the bone mass reduction in estrogen deficiency model rats (#p < 0.05, ##p < 0.01 compared with the sham operation group; *p < 0.05, **p < 0.01 compared with the model group).

[0115] Please refer to Figure 5 , for the detection of ALP activity in the femur tissues of ovariectomized rats treated with Bushen Huoxue Decoction and the modified Bushen Huoxue Decoction respectively, the ALP activity in the femur tissues was higher after the intervention of the modified Bushen Huoxue Decoction.

[0116] The ALP activity detection results showed that the modified Bushen Huoxue Decoction increased the ALP activity in the femur tissues (#p < 0.05, ##p < 0.01 compared with the sham operation group. *p < 0.05, **p < 0.01 compared with the model group).

[0117] Please refer to Figure 6 , for the detection of the expression of osteoblast-specific proteins in the femur tissues of ovariectomized rats treated with Bushen Huoxue Decoction and the modified Bushen Huoxue Decoction respectively, the expression of osteoblast-specific proteins in the femur tissues was higher after the intervention of the modified Bushen Huoxue Decoction.

[0118] Western Blot was used to detect the protein expression level in the femur tissues to evaluate the level of osteoblast-specific proteins. Compared with the sham operation group, the protein expression levels of Runx2 and ALP in the femur tissues of the model group were significantly decreased (P < 0.05); compared with the model group, the protein expression levels of Runx2 and ALP in the femur tissues of the Bushen Huoxue Decoction group and the modified Bushen Huoxue Decoction group were significantly increased (P < 0.05) (#p < 0.05, ##p < 0.01 compared with the sham operation group; *p < 0.05, **p < 0.01 compared with the model group).

[0119] Please refer to Figure 7 , for the effects of the drug-containing sera of Bushen Huoxue Decoction and the modified Bushen Huoxue Decoction on the osteogenic differentiation of MC3T3 cells respectively, the drug-containing serum of the modified Bushen Huoxue Decoction had a better effect.

[0120] MC3T3 cells were cultured with the serum containing Bushen Huoxue Decoction and the serum containing the modified Bushen Huoxue Decoction respectively. After 14 days, the results of alizarin red staining showed that the modified Bushen Huoxue Decoction promoted the differentiation of MC3T3 cells into osteoblasts more effectively than the Bushen Huoxue Decoction.

Claims

1. A modified formula of Bushen Huoxue Decoction, characterized in that: By mass parts, it is composed of 10 - 20 parts of Rehmannia glutinosa Libosch. preparata, 10 - 20 parts of Cuscuta chinensis Lam. salted, 5 - 15 parts of Psoralea corylifolia L. salted, 10 - 20 parts of Lycium barbarum L., 5 - 15 parts of Eucommia ulmoides Oliv. salted, 5 - 15 parts of Cornus officinalis Sieb. et Zucc. wine-processed, 5 - 15 parts of Angelica sinensis (Oliv.) Diels, 5 - 15 parts of Heracleum hemsleyanum Diels, 3 - 9 parts of Carthamus tinctorius L., 14 - 16 parts of Astragalus membranaceus (Fisch.) Bunge honey-roasted, 9 - 11 parts of Atractylodes macrocephala Koidz. stir-fried, and 5 - 7 parts of Glycyrrhiza uralensis Fisch. honey-roasted.

2. The modified formula of Bushen Huoxue Decoction according to claim 1, wherein: By mass parts, it is composed of 15 parts of Rehmannia glutinosa Libosch. preparata, 15 parts of Cuscuta chinensis Lam. salted, 10 parts of Psoralea corylifolia L. salted, 15 parts of Lycium barbarum L., 10 parts of Eucommia ulmoides Oliv. salted, 10 parts of Cornus officinalis Sieb. et Zucc. wine-processed, 10 parts of Angelica sinensis (Oliv.) Diels, 10 parts of Heracleum hemsleyanum Diels, 6 parts of Carthamus tinctorius L., 14 - 16 parts of Astragalus membranaceus (Fisch.) Bunge honey-roasted, 9 - 11 parts of Atractylodes macrocephala Koidz. stir-fried, and 5 - 7 parts of Glycyrrhiza uralensis Fisch. honey-roasted.

3. The modified prescription of Bushen Huoxue Decoction according to claim 2, characterized in that: By mass parts, it is composed of 15 parts of Rehmannia glutinosa Libosch. preparata, 15 parts of Cuscuta chinensis Lam. salted, 10 parts of Psoralea corylifolia L. salted, 15 parts of Lycium barbarum L., 10 parts of Eucommia ulmoides Oliv. salted, 10 parts of Cornus officinalis Sieb. et Zucc. wine-processed, 10 parts of Angelica sinensis (Oliv.) Diels, 10 parts of Heracleum hemsleyanum Diels, 6 parts of Carthamus tinctorius L., 15 parts of Astragalus membranaceus (Fisch.) Bunge honey-roasted, 9 - 11 parts of Atractylodes macrocephala Koidz. stir-fried, and 5 - 7 parts of Glycyrrhiza uralensis Fisch. honey-roasted.

4. The modified prescription of Bushen Huoxue Decoction according to claim 3, characterized in that: By mass parts, it is composed of 15 parts of Rehmannia glutinosa Libosch. preparata, 15 parts of Cuscuta chinensis Lam. salted, 10 parts of Psoralea corylifolia L. salted, 15 parts of Lycium barbarum L., 10 parts of Eucommia ulmoides Oliv. salted, 10 parts of Cornus officinalis Sieb. et Zucc. wine-processed, 10 parts of Angelica sinensis (Oliv.) Diels, 10 parts of Heracleum hemsleyanum Diels, 6 parts of Carthamus tinctorius L., 15 parts of Astragalus membranaceus (Fisch.) Bunge honey-roasted, 10 parts of Atractylodes macrocephala Koidz. stir-fried, and 6 parts of Glycyrrhiza uralensis Fisch. honey-roasted.

5. The modified prescription of Bushen Huoxue Decoction according to claim 4, characterized in that, Its decocting process is as follows: Decoct with water twice. For the first time, add 6 times the mass of water of the modified Bushen Huoxue Decoction, soak for 30 min, decoct for 1 h, and filter; for the second time, add 4 times the mass of water of the modified Bushen Huoxue Decoction, decoct for 45 min, and filter; combine the filtrates of the two times.

6. The modified formula of Bushen Huoxue Decoction according to claim 4, characterized in that Its concentration and drying process is as follows: Concentrate under reduced pressure and then dry at 80 °C.

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