Preparation process of tendon-relaxing and detoxifying granules

Abstract

The invention relates to the technical field of pharmacy, and provides a preparation process of tendon-relaxing and detoxifying granules, which comprises the following steps: step 1, decocting medicines, and adding water for decocting; step 2, adding conventional decoction medicines, decocting, and then filtering to obtain filtrate for later use; step 3, taking medicine residues in the step 2, adding water, decocting, adding uncaria, decocting, and filtering to obtain filtrate for later use; step 4, mixing the medicine liquid in the step 2 and the medicine liquid in the step 3, concentrating under reduced pressure to obtain thick paste, drying under reduced pressure, crushing, and sieving with a 60-mesh sieve to obtain medicine powder; and 5, taking other auxiliary materials, uniformly mixing, adding the medicinal powder in the step 4, uniformly mixing, and granulating. By optimizing the extraction efficiency, the gastrodin content reaches 34.717 mg / g, the astilbin extraction rate reaches 5.944 mg / g, the particle performance is greatly improved, the forming rate is improved to 93.97% from 83.95%, the particle dissolution time is shorter than 5 minutes, and the requirements of pharmacopeia are met. The 80% ethanol wetting agent reduces the moisture absorption rate to 9.60% and improves the storage stability. And multi-component collaborative optimization is realized through an AHP-EWM weight method.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, specifically to the preparation process of muscle-relaxing and detoxifying granules. Background Technology

[0002] Parkinson's disease is a common neurodegenerative disease, most prevalent in middle-aged and elderly individuals. Its etiology is not fully understood, but it may be related to age, genetics, and environmental factors. Its core symptoms are motor disorders, including resting tremor, rigidity, bradykinesia, and postural instability. In addition, it is often accompanied by non-motor symptoms such as decreased sense of smell, sleep disturbances, constipation, and depression. While there is currently no cure, comprehensive treatment including medication, surgery, and rehabilitation can effectively improve symptoms and enhance patients' quality of life.

[0003] The Muscle-Soothing and Detoxifying Granules were developed by Chief Physician Wang Yali of the Affiliated Hospital of Shaanxi University of Traditional Chinese Medicine based on over 30 years of clinical experience. Its formula originates from the TCM principle of "soothing muscles and detoxifying, tonifying the kidneys and promoting blood circulation," and consists of 14 Chinese herbs including Gastrodia elata, Uncaria rhynchophylla, Paeonia lactiflora, raw dragon bone, and raw oyster shell. This formula is primarily designed for the TCM concept of "tremor syndrome," which corresponds to Parkinson's disease in modern medicine. It has a good therapeutic effect on both motor and non-motor symptoms of Parkinson's disease, with a total clinical effective rate of approximately 85%.

[0004] However, the preparation process and quality control of Shujin Jiedu Granules currently suffer from prominent problems. Specifically: The molding process is crude: the excipient ratio is simple, resulting in low granule formation rate, high hygroscopicity, and excessively long dissolving time, affecting the patient's experience; Extraction efficiency is low: insufficient decoction time for mineral drugs leads to low CaCO3 dissolution rate and incomplete extraction of active ingredients, making it difficult to guarantee efficacy; Quality control is incomplete: there are currently no quality standards for Shujin Jiedu Formula, and current methods rely only on single indicators or single component testing, neglecting the synergistic effects of multiple components in the compound, leading to unstable preparation quality. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides a preparation process for muscle-relaxing and detoxifying granules, which solves the problems of crude molding process, low extraction efficiency, and one-sided quality control.

[0006] To achieve the above objectives, the present invention provides the following technical solution: a preparation process for muscle-relaxing and detoxifying granules, comprising the following steps:

[0007] Step 1: Take the herbs to be decocted first, add water and boil;

[0008] Step 2: Add the usual decoction herbs, decoct, then filter and reserve the filtrate.

[0009] Step 3: Take the dregs from Step 2, add water and decoct, then add Uncaria rhynchophylla, decoct again, then filter and keep the filtrate for later use;

[0010] Step 4: Mix the medicinal liquids from Step 2 and Step 3, concentrate under reduced pressure to a thick paste, dry under reduced pressure, pulverize, and pass through a 60-mesh sieve to obtain medicinal powder;

[0011] Step 5: Take another excipient, mix it well, then add the powder from Step 4, mix evenly, and then granulate.

[0012] Preferably, the decoction ingredients in step one include: 280g-320g of raw dragon bone, 280g-320g of raw oyster shell, and 280g-320g of mother-of-pearl; the amount of water added is 14 times the total amount of the prescription, and the decoction time is 100 minutes.

[0013] Preferably, the conventional decoction herbs added in step two include: Gastrodia elata 110g-130g, Rehmannia glutinosa 140g-160g, Paeonia lactiflora 280g-320g, Buthus martensii 50g-70g, Bombyx mori 90g-110g, Spatholobus suberectus 280g-320g, Magnolia officinalis 110g-130g, Smilax glabra 280g-320g, Lycopodium clavatum 140g-160g, and Glycyrrhiza uralensis 20g-40g; the decoction time is 100 minutes.

[0014] Preferably, the amount of Uncaria rhynchophylla added in step three is 140g to 160g; the amount of water added is 12 times the total amount of the prescription; the decoction time of the dregs is 85 minutes; and the decoction is continued for 15 minutes after adding Uncaria rhynchophylla.

[0015] Preferably, the reduced pressure drying conditions in step four are 70°C and 0.06–0.08 MPa.

[0016] Preferably, the excipients in step five include dextrin and lactose, and the mass ratio of dextrin to lactose is 1:1, and the mass ratio of the drug powder to the excipients is 1:1; 80% ethanol is used as a lubricant during granulation, and the granules are dried at 60°C after granulation.

[0017] Preferably, the granules obtained in step five must meet the following requirements: molding rate ≥ 93%, moisture absorption rate ≤ 10% after 48 hours, and melting time < 5 minutes.

[0018] Preferably, the method further includes a quality control step: simultaneously identifying Gastrodia elata and Paeonia lactiflora using thin-layer chromatography, with dichloromethane-ethyl acetate-methanol-formic acid as the developing solvent in a mass ratio of 7.3:0.9:1.8:0.01, spraying with 5% vanillin-sulfuric acid solution, and heating at 105℃ for color development; and determining the content of gastrodin ≥30mg / g and paeoniflorin ≥5mg / g using high-performance liquid chromatography.

[0019] Preferably, the high performance liquid chromatography (HPLC) conditions are as follows: using an Inert Sustain AQ-C18 column, gradient elution with acetonitrile-0.1% formic acid solution as the mobile phase, detection wavelength 251 nm, column temperature 25 °C, and injection volume 50 μl.

[0020] Preferably, the dosages of the pre-decocted herbs, the conventionally decocted herbs, and Uncaria rhynchophylla are as follows: 300g of raw dragon bone, 300g of raw oyster shell, 300g of mother-of-pearl, 120g of Gastrodia elata, 150g of Uncaria rhynchophylla, 150g of Rehmannia glutinosa, 300g of Paeonia lactiflora, 60g of scorpion, 100g of Bombyx mori, 300g of Spatholobus suberectus, 120g of Magnolia officinalis, 300g of Smilax glabra, 150g of Lycopodium clavatum, and 30g of Glycyrrhiza uralensis.

[0021] This invention provides a preparation process for muscle-relaxing and detoxifying granules. It has the following beneficial effects:

[0022] 1. This invention optimizes extraction efficiency, resulting in a gastrodin content of 34.717 mg / g and an astilbin extraction rate of 5.944 mg / g. Granule performance is significantly improved, with the forming rate increasing from 83.95% to 93.97%, and the granule dissolution time being less than 5 minutes, meeting pharmacopoeia requirements. An 80% ethanol wetting agent reduces the moisture absorption rate to 9.60%, improving storage stability. Multi-component synergistic optimization is achieved using the AHP-EWM weighted method, resulting in a comprehensive score of 96.321.

[0023] 2. This invention uses TLC to simultaneously identify Gastrodia elata and Paeonia lactiflora, with dichloromethane-ethyl acetate-methanol-formic acid as the developing solvent at a mass ratio of 7.3:0.9:1.8:0.01. The samples are removed, dried, sprayed with 5% vanillin-sulfuric acid solution, and heated at 105℃ until the spots are clearly visible. HPLC quantitatively covers gastrodin and paeoniflorin, ensuring stable and controllable preparation quality. Physical fingerprint chromatograms show that the similarity between ten batches of samples and the control chromatogram is above 0.99, indicating that the quality of the Shujin Jiedu Granules preparation is stable and the quality control is relatively comprehensive. Attached Figure Description

[0024] Figure 1 These are the HPLC chromatograms of the components of this invention;

[0025] Figure 2 This is a response surface plot showing the influence of the interaction of various factors in this invention on the comprehensive score of the extraction process of the muscle-relaxing and detoxifying formula.

[0026] Figure 3 This is a flowchart of the present invention. Detailed Implementation

[0027] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0028] Example:

[0029] Please see the appendix Figure 1 - Appendix Figure 3 This invention provides a preparation process for muscle-relaxing and detoxifying granules, including the following steps:

[0030] Step 1: Take the herbs to be decocted first, add water and decoct; the herbs to be decocted first include: 280g-320g of raw dragon bone, 280g-320g of raw oyster shell, and 280g-320g of mother-of-pearl; the amount of water added is 14 times the total amount of the prescription, and the decoction time is 100 minutes;

[0031] Step 2: Add the usual decoction herbs, decoct, then filter and reserve the filtrate. The usual decoction herbs include: Gastrodia elata 110g-130g, Rehmannia glutinosa 140g-160g, Paeonia lactiflora 280g-320g, Buthus martensii 50g-70g, Bombyx mori 90g-110g, Spatholobus suberectus 280g-320g, Magnolia officinalis 110g-130g, Smilax glabra 280g-320g, Lycopodium clavatum 140g-160g, Glycyrrhiza uralensis 20g-40g; the decoction time is 100 minutes.

[0032] Step 3: Take the dregs from Step 2, add water and decoct, then add Uncaria rhynchophylla, decoct again, then filter and keep the filtrate for later use; the amount of Uncaria rhynchophylla is 140g-160g; the amount of water is 12 times the total amount of the prescription, the decoction time of the dregs is 85 minutes, and after adding Uncaria rhynchophylla, continue to decoct for 15 minutes.

[0033] Step 4: Mix the liquids from Step 2 and Step 3, concentrate under reduced pressure to a thick paste, dry under reduced pressure, pulverize, and pass through a 60-mesh sieve to obtain powder; the reduced pressure drying conditions are 70℃ and 0.06~0.08MPa.

[0034] Step 5: Take another excipient, mix it evenly, add it to the powder from Step 4, mix evenly, and then granulate. The excipients include dextrin and lactose, and the mass ratio of dextrin to lactose is 1:1, and the mass ratio of powder to excipients is 1:1. 80% ethanol is used as a lubricant during granulation, and the granules are dried at 60°C after granulation. The obtained granules must meet the following requirements: molding rate ≥93%, moisture absorption rate ≤10% after 48 hours, and dissolution time <5 minutes.

[0035] The procedure also includes quality control steps: Thin-layer chromatography (TLC) is used to simultaneously identify Gastrodia elata and Paeonia lactiflora, with dichloromethane-ethyl acetate-methanol-formic acid as the developing solvent (mass ratio 7.3:0.9:1.8:0.01), followed by spraying with 5% vanillin-sulfuric acid solution and heating at 105℃ for color development; high-performance liquid chromatography (HPLC) is used to determine the content of gastrodin (≥30 mg / g) and paeoniflorin (≥5 mg / g); the HPLC conditions are: Inert Sustain AQ-C18 column, acetonitrile-0.1% formic acid solution as the mobile phase for gradient elution, detection wavelength 251 nm, column temperature 25℃, and injection volume 50 μl.

[0036] The dosages for the first decoction, the regular decoction, and Uncaria rhynchophylla are as follows: 300g raw dragon bone, 300g raw oyster shell, 300g mother-of-pearl, 120g gastrodia elata, 150g Uncaria rhynchophylla, 150g rehmannia glutinosa, 300g white peony root, 60g scorpion, 100g silkworm pupae, 300g chicken blood vine, 120g magnolia bark, 300g smilax glabra, 150g lycopodium clavatum, and 30g licorice root.

[0037] I. Composition of the Muscle-Relaxing and Detoxifying Formula

[0038] The ingredients are: Gastrodia elata, Uncaria rhynchophylla, Paeonia lactiflora, raw dragon bone, raw oyster shell, scorpion, silkworm, chicken blood vine, mother-of-pearl, Rehmannia glutinosa, Magnolia officinalis, Smilax glabra, Lycopodium clavatum, and Glycyrrhiza uralensis.

[0039] II. Reagents and Drugs

[0040] The following Chinese medicinal herbs were purchased from Shaanxi Province: Gastrodia elata (batch number: 230117), Uncaria rhynchophylla (batch number: 241101), Paeonia lactiflora (batch number: 230721), raw dragon bone (batch number: 231206), raw oyster shell (batch number: 230413), Rehmannia glutinosa (batch number: 230619), Glycyrrhiza uralensis (batch number: 240202), Spatholobus suberectus (batch number: 230912), Smilax glabra (batch number: 220714), Lycopodium clavatum (batch number: 220809), Magnolia officinalis (batch number: 230721), Bombyx mori (batch number: 220312), Buthus martensii (batch number: 220423), and Mother-of-pearl (batch number: 241008). The samples tested at the Xi'an University of Traditional Chinese Medicine Hospital met the requirements of the 2020 edition of the Chinese Pharmacopoeia. Gastrodin reference standard (batch number: 110807-202010, content ≥96.7%), paeoniflorin (batch number: 110736-202447, content ≥96.7%), and astilbene reference standard (batch number: 111798-202306, content ≥93.6%) were all purchased from the China National Institutes for Food and Drug Control. Methanol and acetonitrile (chromatographic grade) were purchased from Merck & Co., Ltd., USA. Ultrapure water (self-made) was used. All other reagents were of analytical grade. Lactose and dextrin (pharmaceutical grade) were purchased from Zhenjiang Kangfu Biotechnology Co., Ltd.

[0041] Preparation and titration of EDTA disodium ethylenediaminetetraacetate (EDTA) titrant: Refer to the preparation and standardization of EDTA in Part IV of the 2020 edition of the Chinese Pharmacopoeia. Preparation: Dissolve 0.4653 g of EDTA disodium in an appropriate amount of water to make 250 ml, and shake well. Standardization: Accurately transfer 5 mL of calcium standard solution (0.1 mg / ml) into an Erlenmeyer flask, add 1 drop of concentrated hydrochloric acid, add 1 drop of methyl red indicator, and add 10% potassium hydroxide solution until the solution turns yellow. Then add 10 ml of 10% potassium hydroxide solution, and a small amount of calcein indicator. Titrate with EDTA disodium titrant (0.005 mol / L) until the solution changes from yellow-green fluorescence to orange. Correct the titration result with a blank test, and repeat in triplicate. Each 1 ml of disodium ethylenediaminetetraacetate titrant (0.005 mol / L) is equivalent to 0.5004 mg of calcium carbonate (CaCO3). Based on the consumption of this solution (V1) and the amount of calcium standard solution used (V2), the actual concentration of EDTA was calculated according to formula (1), and the results are shown in Table 1.

[0042]

[0043] Table 1 EDTA calibration results

[0044]

[0045] As shown in Table 1, the actual concentration of EDTA titrated was 0.0052 mol / L.

[0046] III. Extraction Process

[0047] In this formula, raw dragon bone, raw oyster shell, and mother-of-pearl are mineral drugs with hard textures, making it difficult to extract their active ingredients. Therefore, they need to be decocted first. Then, add gastrodia elata, white peony root, rehmannia root, licorice root, chicken blood vine, smilax glabra, lycopodium clavatum, magnolia bark, silkworm, and scorpion, and decoct. Finally, add uncaria (because uncaria contains rhubarb alkaloids, which are unstable at high temperatures).

[0048] 1. Examination of the decoction time of the first decoction

[0049] The determination index was set as follows: The raw dragon bone, raw oyster shell and mother-of-pearl, which are the "pre-decocted herbs" in this formula, were taken as the research objects. The content of CaCO3, the main component, was taken as the evaluation index. The content of CaCO3 was determined with reference to the content determination method of oyster shell in Part I of the 2020 edition of the Pharmacopoeia of the People's Republic of China.

[0050] Factor level setting: According to the "Technical Specifications for Decoction of Traditional Chinese Medicine" issued by the China Association of Traditional Chinese Medicine in 2021, raw oyster and mother-of-pearl should be decocted for 20-30 minutes first. Therefore, this experiment adopted a single-factor experiment to examine the decoction time of 20 minutes and 30 minutes respectively.

[0051] Single-factor experiment: Accurately weigh 1 / 10 of the prescription amount of the "pre-decocted medicine" raw dragon bone (3g), raw oyster shell (3g), and mother-of-pearl (3g), totaling 9g (M), place them in a decoction pot, add 10 times the amount of pharmaceutical water, and start timing after it boils slightly. Decoct for 20 and 30 minutes respectively, with 3 samples in parallel for each decoction time. After returning to room temperature, first filter with four layers of gauze, then filter by suction, and determine the CaCO3 content of the filtrate.

[0052] Determination of CaCO3: Accurately pipette 50 ml of the filtrate, add 2 drops of concentrated hydrochloric acid, heat to remove CO2, wait for it to return to room temperature, add 1 drop of methyl red indicator, add 10% potassium hydroxide solution until the solution turns yellow, then add 10 ml of 10% potassium hydroxide solution, add a small amount of calcein indicator, and titrate with disodium ethylenediaminetetraacetate titrant (0.005 mol / L) until the yellow-green fluorescence of the solution disappears and turns orange. Perform three parallel titrations and correct the titration results with a blank test. Calculate the CaCO3 content according to formula (2). The results are shown in Table 2.

[0053]

[0054] In the formula, T is the titer; V is the volume of titrant consumed; F is the ratio of the actual concentration of the titrant to the specified concentration; and M is the total mass of the decocted medicinal pieces.

[0055] Table 2. Titration results of CaCO3 content in the pre-decocted herbs.

[0056]

[0057]

[0058] Table 2 shows that: the first decoction should be boiled for 30 minutes. a The CO3 content was higher than 20 min, and the difference was highly significant (P = 0.0056 < 0.01). Therefore, it was chosen to simmer for 30 min first.

[0059] 2. Investigation into the extraction processes of medicinal herbs such as Gastrodia elata, Paeonia lactiflora, and Smilax glabra.

[0060] Evaluation index setting: Based on the prescription's explanation, the principal, assistant, and adjuvant herbs in the prescription were selected as the herbs to be analyzed. Combining the research results of the chemical components of each herb in the prescription, the content and yield of effective components such as gastrodin, paeoniflorin, and astilbin in the above-mentioned herbs were selected as evaluation indicators.

[0061] Factor level setting: Considering that factors such as soaking time, water volume, extraction time, and number of extractions affect the extraction of the above indicators, this experiment first uses single-factor experiments to find the optimal level of each factor, and then uses a 3-factor, 3-level star point design centered on the optimal level to screen the process.

[0062] Evaluation index setting: Based on the prescription's explanation, the principal, assistant, and adjuvant herbs in the prescription were selected as the herbs to be analyzed. Combining the research results of the chemical components of each herb in the prescription, the content and yield of effective components such as gastrodin, paeoniflorin, and astilbin in the above-mentioned herbs were selected as evaluation indicators.

[0063] Factor level setting: Considering that factors such as soaking time, water volume, extraction time, and number of extractions affect the extraction of the above indicators, this experiment first uses single-factor experiments to find the optimal level of each factor, and then uses a 3-factor, 3-level star point design centered on the optimal level to screen the process.

[0064] 2.1 Determination of the content of gastrodin, paeoniflorin, and astilbin

[0065] Chromatographic conditions: InertSustain AQ-C18 (250 mm × 4.6 mm, 5 μm) column was used; mobile phase: acetonitrile (A) - 0.1% formic acid solution as mobile phase (B), gradient elution (0–10 min, 98% B, 0.8 mL / min; 15–45 min, 96%–90% B, 1.0 mL / min; 55–60 min, 89%–89% B, 1.0–1.2 mL / min; 90–100 min, 83%–40% B, 1.2 mL / min; 100–110 min, 40% B, 1.2 mL / min); detection wavelength: 251 nm; column temperature: 25 ℃; injection volume: 10 μl.

[0066] Preparation of reference solutions: Accurately weigh appropriate amounts of each reference standard and add 50% methanol to prepare single-standard stock solutions containing 2.628 mg of gastrodin, 2.726 mg of paeoniflorin, and 1.588 mg of astilbene per 1 mL. Accurately pipette 1 mL of gastrodin reference stock solution, 0.5 mL of paeoniflorin reference stock solution, and 0.1 mL of astilbene reference stock solution into a 5 mL volumetric flask, dilute to the mark with 50% methanol, and shake well to obtain the mixed standard reference solution.

[0067] Preparation of the test solution: Take 1 / 10 of the total prescription amount, weigh accurately, extract according to the factor levels of single-factor experiment and star point experiment, and dilute the extract to a certain volume to obtain the test solution.

[0068] Preparation of negative solution: Take the other medicinal slices in the prescription except for Gastrodia elata, Paeonia lactiflora and Smilax glabra, and extract them according to the prescription ratio using the final optimized process.

[0069] Specificity test: Take 10 μL each of the reference solution, test solution, and negative solution, and inject them according to the above chromatographic conditions. The results are shown in the figure. Figure 1Therefore, the analytes showed good separation and no interference from negative samples, indicating good specificity of the method. Note: A - Mixed reference standard, B - Test sample, C - Negative sample; 1-Gastrodin, 2-Paeoniflorin, 3-Astilbene.

[0070] Linearity assessment: Precisely pipettes of 1, 2, 5, 10, 15, 20, 25, and 30 μl of the above mixed standard solution were injected into the liquid chromatograph, and the analysis was performed under the chromatographic conditions described above. Linear regression was performed with mass (μg) as the abscissa (x) and peak area as the ordinate (y). The linear equations, correlation coefficients, and linear ranges for each analyte are shown in Table 3. It can be seen that each component exhibits good linearity within its respective range.

[0071] Table 3 Results of linear relationship examination

[0072] Indicator Components Regression equation <![CDATA[R 2 ]]> Linear range / (μg) Gastrodin y = 0.5906x - 0.0097 0.9998 1.0512～13.14 paeoniflorin y = 1.701x - 0.0595 0.9999 0.5452～6.815 Astilbene y = 1.7217x - 0.0141 0.9991 0.03176～0.9528

[0073] Precision test: Take the above mixed standard solution, inject 15 μl, and determine 6 times under the above chromatographic conditions. Record the peak area of ​​the analyte. The RSD of the peak areas of gastrodin, paeoniflorin and astilbene were 0.49%, 0.44% and 0.84%, respectively. The results are shown in Table 4, indicating that the instrument has good precision.

[0074] Table 4 Precision Test Results

[0075]

[0076] Repeatability test: Six test solutions were prepared using the same sample according to the optimal extraction process. The RSDs of gastrodin, paeoniflorin and astilbin were determined under the above chromatographic conditions. The results are shown in Table 5, indicating that the method has good repeatability.

[0077] Table 5 Results of Repeatability Tests

[0078]

[0079] Stability test: The same test solution was injected at 0, 2, 4, 6 and 8 h, and the peak area of ​​the analyte was recorded. The RSD of the peak areas of gastrodin, paeoniflorin and astilbin were 2.86%, 2.94% and 2.29%, respectively. The results are shown in Table 6, indicating that the analyte in the test solution has good stability within 8 h.

[0080] Table 6. Stability Test Results

[0081]

[0082]

[0083] Spiking recovery test: Take 1 mL of the concentrated solution of the test sample with known content of each component, make 6 portions, and accurately add appropriate amounts of gastrodin, paeoniflorin, and astilbene reference standards in a 1:1 (m / m) ratio with the content of each component. Prepare the test solution according to the above method, and then inject and determine the sample under the above chromatographic conditions. Record the peak area and calculate the spiking recovery rate. The average spiking recovery rates of gastrodin, paeoniflorin, and astilbene were calculated to be 98.33%, 99.22%, and 98.87%, respectively, with RSDs of 1.25%, 1.38%, and 1.22%, respectively. The results are shown in Table 7, indicating that the method is accurate and feasible.

[0084] Table 7 Results of the recovery test

[0085]

[0086] 3. Determination of extract yield

[0087] The total volume of each test solution under the single-factor and star-point design items is recorded as TV. Accurately transfer 20 ml of each test solution to an evaporating dish dried to constant weight (constant weight weighing m1), evaporate to dryness in a water bath, place in an oven at 105℃ for 3 hours, take it out, cool in a desiccator for 30 minutes, and quickly weigh it until constant weight (m2). Calculate the paste yield (%) according to formula (3).

[0088] Extraction rate (%) = [(m2-m1) / 20×TV]×100% Formula (3)

[0089] 4. Determining the weights of relevant indicators for comprehensive scoring

[0090] A comprehensive scoring method combining subjective and objective evaluation was adopted to assign different weights to the measured indicators. Subjective weights were obtained by subjective weighting using the AHP method based on the theory of monarch, minister, assistant and guide in traditional Chinese medicine prescriptions, and objective weights were obtained by objective weighting using the entropy value method based on the results of single-factor experiments. The comprehensive weights were obtained by combining the subjective and objective weighting methods.

[0091] Subjective Weighting Using the AHP Method: The AHP method was used to subjectively weight the set indicators. Based on the principal, assistant, adjuvant, and guide relationships and dosages of the herbs in the muscle-relaxing and detoxifying formula, the indicators were hierarchically divided in the following order: gastrodin > paeoniflorin > astilbin > extract yield. A priority judgment matrix was then constructed, as shown in Table 8. SPSSAU was used to standardize and positively process the data. The calculated subjective weight coefficients W1 for gastrodin, paeoniflorin, astilbin, and extract yield were 0.42217, 0.26636, 0.18963, and 0.12184, respectively, with a consistency ratio of 0.026 (<0.10), indicating that the calculated indicator weight coefficients are reliable and effective.

[0092] Table 8. AHP Hierarchical Analysis Matrix

[0093]

[0094] Objective weighting using entropy method: The entropy method was used to objectively weight the set indicators. SPPSAU was used to standardize and positively process the data. The objective weighting coefficients W2 of gastrodin, paeoniflorin, astilbene, and extract yield were calculated by entropy method to be 0.4319, 0.1883, 0.1612, and 0.2168, respectively. The results are shown in Table 9.

[0095] Table 9. Entropy values ​​and weighting coefficients of each indicator

[0096]

[0097] Determination of comprehensive weighting coefficients and calculation of comprehensive score: The weighting coefficients of each indicator determined by the AHP method and the entropy weight method are W1 and W2, respectively, and the comprehensive weighting coefficient (W) is calculated. 综合 ): The comprehensive weights of gastrodin, paeoniflorin, astilbene, and extract yield were 0.6299, 0.1733, 0.1056, and 0.0913, respectively. The W values ​​for each indicator were... 综合 Substitute the values ​​into the comprehensive score for the test, and the comprehensive score is calculated as follows: (0.6299 × gastrodin content / maximum gastrodin content) + (0.1733 × paeoniflorin content / maximum paeoniflorin content) + (0.1056 × astilbin content / maximum astilbin content) + (0.0913 × extract yield / maximum extract yield) × 100.

[0098] 5. Single-factor experiment

[0099] This experiment used extraction times (1, 2, 3 times), soaking time (0, 30, 60, 90 min), water volume (8, 10, 12, 14 times), and extraction time (30, 60, 90, 120 min) as factors to conduct a single-factor experiment. The content of each component, the yield of extract, and its comprehensive score (M) were measured and calculated. The results are shown in Table 10.

[0100] Table 10 Results of the Single-Factor Experiment

[0101]

[0102]

[0103] As shown in Table 10, the soaking time P = 0.16 > 0.1, indicating no significant difference. Therefore, soaking time of 0 min was selected. For the other three single factors, a three-factor, three-level response surface model was designed with the optimal factor level (two extractions, 12 times water addition, and 90 min decoction time) as the center, and the results were repeated twice.

[0104] 6. Box-Behnken Response Surface Methodology

[0105] Based on the single-factor experiments, the number of extractions (A), the water addition ratio (B), and the decoction time (C) were selected as influencing factors. Each factor had three levels, and a three-factor, three-level experiment was established using Design-Expert 13.0 software. The factor levels are shown in Table 11. The experimental design and results are shown in Table 12. The optimal combination of factor levels was optimized using the overall score (M) as the evaluation index.

[0106] Table 11 Experimental Design Level and Factors

[0107]

[0108] Table 12 Box-Behnken Test Protocol and Results

[0109]

[0110]

[0111] Model fitting and analysis of variance: The data in Table 12 were fitted using Design-Expert 13.0 software, with A (number of extractions), B (cooking time), and C (water addition ratio) as independent variables, and Y as the comprehensive score. The results are as follows:

[0112] Y=80.81+11.88A+6.00B+5.45C+0.8057AB+0.5907AC-0.1817BC-0.1932A 2

[0113] -7.99B 2 -2.21, the results of the regression model variance analysis are shown in Table 13.

[0114] Table 13 Results of ANOVA for the Regression Model

[0115]

[0116] Note: *P<0.05, significant difference; **P<0.01, highly significant difference; ***P<0.001, extremely significant difference.

[0117] Table 13 shows that the model P < 0.0001, the lack-of-fit term value is 0.1906, indicating that the model has extremely significant differences, the lack-of-fit term is not significant, and the correlation coefficient R of the comprehensive score regression equation is... 2 =0.9844, and the coefficient of variation is 0.8274. The difference between the two is less than 0.2, indicating that the model is realistic and reliable, and can be used to analyze the degree of closeness between the experimental results of extraction process optimization and the predicted values ​​of the optimal extraction process. R adj=0.9644, indicating that 96.41% of the comprehensive score of the extraction process of the Shujin Jiedu formula was affected by the number of extractions, decoction time, and amount of water added. Factors A, B, and C showed highly significant differences in the score. The order of influence of each factor on the extraction process of the Shujin Jiedu formula was B > C > A. In the binomial formula, B... 2 The effect was extremely significant, and the F value was high, so the influence of simmering time should be fully considered. However, the interaction between the three factors was not significant.

[0118] Box-Behnken response surface analysis of the regression model: Response surface diagrams of the interactions of various factors were plotted using Design-Expert 13.0 software. (See...) Figure 2 .

[0119] Based on the model fitting results, the optimal extraction process was determined to be 2.939 extractions, with a water volume of 13.649 times, and an extraction time of 101.137 minutes. Considering industrial production, the optimal extraction process was determined to be 2 extractions, with water volume of 14 times for the first extraction and 12 times for the second extraction, each extraction lasting 100 minutes.

[0120] 7. Extraction process verification test

[0121] Three batches of samples were prepared according to the above-mentioned optimal extraction process, and verification tests were conducted. The results are shown in Table 14.

[0122] Table 14 Results of the Optimal Process Verification Test for the Muscle-Soothing and Detoxifying Formula

[0123]

[0124] As shown in Table 14, the average comprehensive score (M) is 96.321, which is close to the predicted value (96.623) (relative error < 3.0%), indicating that the extraction process is stable and reliable and the model has good predictive ability.

[0125] IV. Molding Process

[0126] Evaluation indicators were set as follows: Based on the general rules for granulation of granules in Part IV of the 2020 edition of the Chinese Pharmacopoeia and related references, the properties of soft material, ease of granulation, granulation rate, solubility, angle of repose, 48-hour moisture absorption rate, and granule appearance were used as evaluation indicators for the formulation process.

[0127] Formation rate (%): Tested according to the double sieving method of Part IV of the 2020 edition of the Chinese Pharmacopoeia. Particles that can pass through sieve No. 1 (10 mesh) but cannot pass through sieve No. 5 (80 mesh) are qualified particles. Accurately weigh the mass of the particles as M1, place them in the specified sieve, keep them horizontal and sieve them back and forth, tap them lightly for 3 minutes while sieving, collect the qualified particles that pass through sieve No. 1 but not sieve No. 5, accurately weigh them (M2), and calculate their formation rate (%) using formula (4).

[0128] Molding rate (%) = M2 / M1·100% Formula (4)

[0129] Solubility: According to the method under the General Rules for Preparations in Part IV of the 2020 edition of the Chinese Pharmacopoeia, take 10g of the test sample, add 200ml of hot water, stir for 5 minutes, and observe immediately. Soluble particles should be completely dissolved or slightly turbid.

[0130] Angle of repose (°): According to the reference "Optimization of the preparation process of compound five-flower granules", the fixed funnel method is adopted. Take a watch dish with a radius (R) of 2.675cm (placed flat on the table), fix the funnel on the top, and make its lower end face the center of the watch dish. Qualified granules (which can pass through No. 1 sieve but not No. 5 sieve) are slowly added from the top of the funnel and allowed to flow down naturally. When the edge of the cone formed by the granules at the bottom of the funnel coincides with the edge of the watch dish, the height (H) is measured and α is calculated. The angle of repose (°) is calculated by formula (5).

[0131] Angle of repose (°) = α = atctan(H / R) Formula (5)

[0132] 48h moisture absorption rate (%): According to the method in the literature "Preparation process and quality control of Erdongtang granules based on QbD", a glass desiccator containing a supersaturated sodium chloride solution was placed in a constant temperature incubator at 25℃ for 24h to achieve a relative humidity (RH) balance of 75%. The desiccator was then placed in a flat weighing bottle for 48h, weighed (m1), and an appropriate amount of granules (2g) was accurately weighed (m2). The granules were then spread evenly in the flat weighing bottle with a thickness not exceeding 5mm. The bottle was placed in the desiccator (with the cap of the flat weighing bottle open) for 48h, the cap was closed, and the bottle was weighed (m3). The moisture absorption rate was calculated using formula (6).

[0133] 48h moisture absorption rate (%) = [(m3-m1) / m2]×100% Formula (6)

[0134] 1. Preparation of dry extract powder

[0135] Weigh out all fourteen herbs in the formula. Add the raw dragon bone and other herbs to 14 times the amount of water in the formula and decoct for 30 minutes. Then add the gastrodia elata and other ten herbs and decoct for 110 minutes. Filter to obtain the filtrate. Add the dregs to 12 times the amount of water in the formula and decoct for 95 minutes. Add the uncaria rhizome and decoct for 15 minutes. Filter and combine the filtrates. Concentrate the filtrate under reduced pressure, dry under reduced pressure at 70℃ and 0.06–0.08 MPa, pulverize, and pass through a 60-mesh sieve to obtain the dry extract powder.

[0136] 2. Single-factor screening for formulation formation

[0137] Screening of excipients: An appropriate amount of dry extract powder from section "2.1" was used as excipients with dextrin:lactose ratios of 1:0, 0:1, 1:1, 1:2, and 2:1, respectively, and a drug-to-excipient ratio of 1:1. Granulation was performed using 80% ethanol concentration via extrusion granulation, followed by drying at 60℃. Granulation was then carried out to obtain the final product. The excipients were optimized based on the properties of the soft material, ease of granulation, and molding rate. The results are shown in Table 15.

[0138] Table 15 Results of the Investigation on Types of Auxiliary Materials

[0139]

[0140] Table 15 shows that the optimal ratio of dextrin to lactose is 1:1.

[0141] Excipient-drug ratio: Take an appropriate amount of the dry extract powder from section "2.1", use dextrin and lactose in a 1:1 ratio as excipients, and ethanol (volume fraction) as a wetting agent with a volume fraction of 80%. The excipient-drug ratios are 1:0.5, 1:1, and 1:1.5, respectively. Granulate by extrusion and dry at 60℃. Granulate to obtain the final product. The optimal excipient-drug ratio was selected based on granule formation rate, solubility, angle of repose, 48-hour moisture absorption rate, and granule appearance. The results are shown in Table 16.

[0142] Table 16 Results of the Drug-Excipient Ratio Study

[0143]

[0144] As shown in Table 16, the optimal drug-to-excipient ratio is 1:1.

[0145] Ethanol volume fraction as wetting agent: An appropriate amount of dry extract powder from section "2.1" was used as an excipient in a 1:1 ratio of dextrin and lactose, with a drug-to-excipient ratio of 1:1. Granulation was performed with ethanol volume fractions of 70%, 80%, and 90%, respectively. The granulation method was used, followed by drying at 60℃. Granulation was then completed to obtain the final product. The optimal ethanol volume fraction was selected based on granule formation rate, solubility, angle of repose, 48-hour moisture absorption rate, and granule appearance. The results are shown in Table 17.

[0146] Table 17 Results of the investigation on ethanol volume fraction

[0147]

[0148] As shown in Table 17, the optimal wetting agent is 80% ethanol by volume.

[0149] 3. Granulation process validation

[0150] Take an appropriate amount of the dry extract powder from section "2.1" and verify the granulation process according to the optimal single-factor level obtained above. That is, use dextrin to lactose in a 1:1 ratio as excipients, a pharmaceutical-excipient ratio of 1:1, and 80% ethanol by volume for granulation. Perform granulation in triplicate using the extrusion method. The granule formation rate, solubility, angle of repose, 48-hour moisture absorption rate, and granule appearance are used as evaluation indicators. The results are shown in Table 18.

[0151] Table 18 Validation Results of Granulation Process

[0152]

[0153] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. The preparation process of muscle-relaxing and detoxifying granules, characterized in that, Includes the following steps: Step 1: Take the herbs to be decocted first, add water and boil; Step 2: Add the usual decoction herbs, decoct, then filter and reserve the filtrate. Step 3: Take the dregs from Step 2, add water and decoct, then add Uncaria rhynchophylla, decoct again, then filter and keep the filtrate for later use; Step 4: Mix the medicinal liquids from Step 2 and Step 3, concentrate under reduced pressure to a thick paste, dry under reduced pressure, pulverize, and pass through a 60-mesh sieve to obtain medicinal powder; Step 5: Take another excipient, mix it well, then add the powder from Step 4, mix evenly, and then granulate.

2. The preparation process of the muscle-relaxing and detoxifying granules according to claim 1, characterized in that, The first decoction step includes: 280g-320g of raw dragon bone, 280g-320g of raw oyster shell, and 280g-320g of mother-of-pearl; the amount of water added is 14 times the total amount of the prescription, and the decoction time is 100 minutes.

3. The preparation process of the muscle-relaxing and detoxifying granules according to claim 1, characterized in that, The conventional decoction herbs added in step two include: Gastrodia elata 110g-130g, Rehmannia glutinosa 140g-160g, Paeonia lactiflora 280g-320g, Buthus martensii 50g-70g, Bombyx mori 90g-110g, Spatholobus suberectus 280g-320g, Magnolia officinalis 110g-130g, Smilax glabra 280g-320g, Lycopodium clavatum 140g-160g, and Glycyrrhiza uralensis 20g-40g; the decoction time is 100 minutes.

4. The preparation process of the muscle-relaxing and detoxifying granules according to claim 1, characterized in that, In step three, the amount of Uncaria rhynchophylla added is 140g to 160g; the amount of water added is 12 times the total amount of the prescription; the decoction time of the dregs is 85 minutes; after adding Uncaria rhynchophylla, continue to decoct for 15 minutes.

5. The preparation process of the muscle-relaxing and detoxifying granules according to claim 1, characterized in that, The reduced pressure drying conditions in step four are 70℃ and 0.06~0.08MPa.

6. The preparation process of the muscle-relaxing and detoxifying granules according to claim 1, characterized in that, In step five, the excipients include dextrin and lactose, with a mass ratio of dextrin to lactose of 1:1 and a mass ratio of drug powder to excipients of 1:

1. 80% ethanol is used as a lubricant during granulation, and the granules are dried at 60°C after granulation.

7. The preparation process of the muscle-relaxing and detoxifying granules according to claim 1, characterized in that, The granules obtained in step five must meet the following requirements: molding rate ≥ 93%, moisture absorption rate ≤ 10% after 48 hours, and melting time < 5 minutes.

8. The preparation process of the muscle-relaxing and detoxifying granules according to claim 1, characterized in that, The method also includes quality control steps: thin-layer chromatography is used to simultaneously identify Gastrodia elata and Paeonia lactiflora, with dichloromethane-ethyl acetate-methanol-formic acid as the developing solvent in a mass ratio of 7.3:0.9:1.8:0.

01. The mixture is sprayed with 5% vanillin-sulfuric acid solution and heated at 105℃ for color development. High-performance liquid chromatography is used to determine the content of gastrodin (≥30 mg / g) and paeoniflorin (≥5 mg / g).

9. The preparation process of the muscle-relaxing and detoxifying granules according to claim 8, characterized in that, The high-performance liquid chromatography (HPLC) conditions were as follows: using an Inert Sustain AQ-C18 column, gradient elution with acetonitrile-0.1% formic acid solution as the mobile phase, detection wavelength of 251 nm, column temperature of 25 °C, and injection volume of 50 μl.

10. The preparation process of the muscle-relaxing and detoxifying granules according to claim 1, characterized in that, The dosages of the pre-decocted herbs, the conventionally decocted herbs, and Uncaria rhynchophylla are as follows: 300g of raw dragon bone, 300g of raw oyster shell, 300g of mother-of-pearl, 120g of Gastrodia elata, 150g of Uncaria rhynchophylla, 150g of Rehmannia glutinosa, 300g of Paeonia lactiflora, 60g of scorpion, 100g of Bombyx mori, 300g of Spatholobus suberectus, 120g of Magnolia officinalis, 300g of Smilax glabra, 150g of Lycopodium clavatum, and 30g of Glycyrrhiza uralensis.

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