Preparation method and application of a traditional Chinese medicine composition for treating joint disease of damp-heat blocking arthralgia syndrome

Abstract

The application belongs to the technical field of traditional Chinese medicine preparation, and discloses a preparation method and application of a traditional Chinese medicine composition for treating joint disease of damp-heat resisting arthralgia syndrome, wherein the preparation method adopts pulse temperature control pre-extraction at 65 DEG C-75 DEG C, high-low temperature alternating period fluctuation extraction, online determination of extraction end point with a spectrum change rate of less than or equal to 0.02, low-temperature and reduced-pressure concentration at 55 DEG C-60 DEG C, dual quality control of components and anti-inflammatory activity, and low-temperature drying and granulation at 60 DEG C-70 DEG C; the dry extract prepared has polydatin greater than or equal to 2.8 mg / g and paeoniflorin greater than or equal to 1.5 mg / g, and is suitable for joint disease of damp-heat resisting arthralgia syndrome. The process can simultaneously improve the dissolution of heat-resistant components, protect heat-sensitive active substances, has small batch differences, and the dual quality control system guarantees the stable anti-inflammatory therapeutic effect of the finished product.

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese medicine pharmaceutical technology, specifically relating to a method for preparing a traditional Chinese medicine composition for treating joint diseases caused by damp-heat obstruction and its application in the preparation of anti-inflammatory, analgesic, and immunomodulatory drugs. Background Technology

[0002] Damp-heat obstruction syndrome is an important syndrome type in Traditional Chinese Medicine (TCM) arthralgia, clinically manifested as redness, swelling, heat, and pain in the joints, difficulty in flexion and extension, fever without fever, thirst without desire to drink, red tongue with yellow and greasy coating, and a slippery and rapid pulse. Modern medicine conditions such as rheumatoid arthritis, gouty arthritis, and acute exacerbations of osteoarthritis largely fall under this syndrome category. Epidemiological statistics indicate that there are over 50 million patients with rheumatic and autoimmune diseases in my country, of whom approximately 35% to 45% suffer from damp-heat obstruction syndrome, severely impacting their quality of life.

[0003] Existing traditional Chinese medicine preparations for treating damp-heat obstruction syndrome have the following shortcomings: Defect 1: Traditional decoction process parameters are crude, resulting in low dissolution rates of active ingredients. Most existing compound Chinese medicine preparations use a generic process of "decocting twice with water, 1-2 hours each time," without precise temperature control tailored to the differences in the physicochemical properties of the compound herbs. Heat-sensitive phenolic glycosides such as polydipsia glycoside and paeoniflorin are prone to hydrolysis and oxidation during high-temperature, long-term decoction, leading to low levels of effective ingredients in the finished product. Polydipsia glycoside is typically below 2.0 mg / g, indicating insufficient anti-inflammatory activity.

[0004] Defect 2: The extraction process lacks online monitoring, and endpoint determination relies on experience. Existing processes mostly use fixed-time extraction, such as decocting for 1 hour, which cannot reflect the dynamic changes in component dissolution caused by batch-to-batch and origin differences in medicinal materials. Insufficient extraction results in active ingredients remaining in the residue, while over-extraction leads to the degradation of heat-sensitive components, resulting in large batch-to-batch variations in product quality (RSD>15%).

[0005] Deficiency 3: Lack of quality control indicators oriented towards anti-inflammatory activity. Existing quality standards only control the content of indicator components such as polydipsia glycoside and paeoniflorin, without establishing a correlation between chemical components and pharmacodynamic activity. Some batches, although meeting the content standards, have insufficient anti-inflammatory activity, resulting in unstable clinical efficacy.

[0006] Defect 4: The single extraction process cannot simultaneously accommodate the differentiated dissolution characteristics of multiple active ingredients. This compound contains multiple active ingredients, including phenolic glycosides, flavonoids, triterpenoid saponins, and polysaccharides. Among them, phenolic glycosides include polysaccharide, paeoniflorin, and salvianolic acid; flavonoids include astragaloside and icariin; triterpenoid saponins include coixenoside and senna saponin; and polysaccharides include astragalus polysaccharide and achyranthes polysaccharide. The optimal dissolution temperature, time, and pH conditions for each type of component differ significantly, which is difficult to accommodate using traditional single decoction processes. Summary of the Invention

[0007] The present invention aims to solve the technical problems in the prior art that the extensive extraction process parameters of traditional Chinese medicine compound preparations lead to the degradation of heat-sensitive active ingredients, the lack of online monitoring leads to inaccurate end-point judgment, and the lack of anti-inflammatory activity-oriented quality control leads to unstable therapeutic effects.

[0008] To solve the above technical problems, one aspect of the present invention provides a preparation method of a traditional Chinese medicine composition for treating arthropathy caused by damp-heat obstruction syndrome, comprising the following steps: Mixing the prescription medicinal materials and water at a solid-liquid ratio of 1:8 to 1:12, controlling the pre-extraction temperature to 65°C to 75°C, and keeping the temperature for 8 min to 15 min to complete low-temperature temperature-controlled pre-extraction; After completing the pre-extraction, raise the temperature of the high-temperature section to 85°C to 95°C, keep the temperature in the high-temperature section for 20 min to 30 min, then lower the temperature of the low-temperature section to 70°C to 80°C, and keep the temperature in the low-temperature section for 15 min to 25 min, thus completing one temperature-rising and falling cycle; repeat 2 to 4 cycles to obtain an extract; An ultraviolet-visible spectrophotometry is used to online monitor the change rate of absorbance of the extract at a characteristic wavelength; the absorbance change rate is calculated based on continuous sampling data according to a preset sampling interval, and the extraction end point is determined when the spectral change rate is ≤ 0.02; Separating the extract from the medicinal residues by filtration, and concentrating the extract under reduced pressure to a clear paste with a relative density of 1.15 to 1.25; An LPS-induced NO release inhibition model in RAW264.7 cells is used to detect the in vitro anti-inflammatory activity of the clear paste, and it is qualified when the half-maximal inhibitory concentration IC50 is ≤ 50 μg / mL; meanwhile, HPLC is used to determine the contents of polydatin and paeoniflorin, and it is qualified when polydatin content is ≥ 2.8 mg / g and paeoniflorin content is ≥ 1.5 mg / g; Drying the clear paste to obtain a dry extract, pulverizing it, adding pharmaceutically acceptable excipients, and preparing granules, tablets, capsules or oral liquid.

[0009] Further, the prescription medicinal materials comprise Astragalus membranaceus, Flemingia philippinensis, Coix chinensis, Polygonum cuspidatum, Ramulus Mori, Achyranthes bidentata, Radix Paeoniae Rubra, Salvia miltiorrhiza and Epimedium brevicornu.

[0010] Further, the prescription medicinal materials in parts by weight are: 25 to 35 parts of Astragalus membranaceus, 15 to 25 parts of Flemingia philippinensis, 15 to 25 parts of Coix chinensis, 8 to 15 parts of Polygonum cuspidatum, 8 to 15 parts of Ramulus Mori, 8 to 12 parts of Achyranthes bidentata, 8 to 12 parts of Radix Paeoniae Rubra, 5 to 10 parts of Salvia miltiorrhiza, and 5 to 10 parts of Epimedium brevicornu.

[0011] Further, the pre-extraction temperature is 70°C, the heat preservation time is 12 min, and the solid-liquid ratio is 1:10.

[0012] Furthermore, the high-temperature section has a temperature of 90°C and a holding time of 25 minutes, the low-temperature section has a temperature of 75°C and a holding time of 20 minutes, and the cycle is repeated 3 times.

[0013] Furthermore, the characteristic wavelength is 340 nm, and the sampling interval is 3 min.

[0014] Furthermore, the temperature for the vacuum concentration is 55℃~60℃, and the vacuum degree is -0.08MPa~-0.06MPa.

[0015] Furthermore, the drying is vacuum drying, with a drying temperature of 60℃~70℃ and a vacuum degree of -0.08MPa~-0.06MPa.

[0016] On the other hand, the present invention provides applications of traditional Chinese medicine compositions, including: The in vitro anti-inflammatory IC50 of the traditional Chinese medicine composition is ≤50μg / mL.

[0017] The aforementioned traditional Chinese medicine composition is used to treat joint diseases caused by damp-heat obstruction.

[0018] The traditional Chinese medicine composition is used for anti-inflammatory, analgesic, and immune-modulating purposes.

[0019] Compared with the prior art, the present invention has the following beneficial effects: Furthermore, by fixing the pre-extraction temperature to a fixed parameter range of 65℃~75℃, the degree of pectin softening and melting is stably controlled within the moderate hydration range, cell wall pores are open while starch is not completely gelatinized, and turbidity is stabilized at 0.15NTU~0.25NTU. The dissolution rate of heat-sensitive phenolic glycosides such as polydipsia glycoside and paeoniflorin is increased by 25%~35%, and the measured dissolution rate of polydipsia glycoside can reach 82.3%, effectively avoiding the component degradation problem caused by high temperature above 78℃; Furthermore, by fixing the temperature range of the periodic fluctuation extraction to a fixed parameter range of 85℃~95℃ (high temperature range) and 70~80℃ (low temperature range), triterpenoid saponins and polysaccharides are fully dissolved in the high temperature range, while phenolic glycosides are effectively protected in the low temperature range. This results in a 40%–55% increase in the total saponin content of the final dry extract compared to the traditional decoction process. The measured dissolution rate of coix seed saponins can reach 78.5%, and the retention rate of icariin can reach 92.5%. Furthermore, by fixing the spectral change rate threshold to a fixed parameter of ≤0.02, the accuracy of extraction endpoint determination is stably controlled at over 95%, with a measured accuracy of 96.8%. This avoids the problems of insufficient or over-extraction caused by fixed-time extraction, and reduces the batch-to-batch RSD from 15%–20% in traditional processes to 3%–5%. Furthermore, by combining the in vitro anti-inflammatory activity index IC50≤50μg / mL with the chemical indicators glutinin≥2.8mg / g and paeoniflorin≥1.5mg / g for quality control, a dual quality correlation between chemical components and pharmacodynamic activity was established to ensure the stable and reliable clinical efficacy of each batch of products. Furthermore, the parameter systems of each step are coupled with each other. The pre-extraction process provides a stable substrate with low turbidity and low colloids for the periodic fluctuation extraction process. The online monitoring process can accurately control the extraction endpoint and avoid over-extraction. The dual quality control system can ensure the quality of the finished product after concentration and drying. Finally, the comprehensive quality indicators of the dry extract are achieved as follows: 3.2mg / g~4.5mg / g of polygalactoside, 1.8mg / g~2.6mg / g of paeoniflorin, 85mg / g~120mg / g of total saponins, and 35~50μg / mL of in vitro anti-inflammatory IC50. Attached Figure Description

[0020] Figure 1 This is a flowchart illustrating the preparation process of an embodiment of the invention; Figure 2 This is a flowchart illustrating the steps of low-temperature temperature-controlled pre-extraction in an embodiment of the invention. Figure 3 This is a flowchart illustrating the steps for determining the extraction endpoint in an embodiment of the present invention. Figure 4 This is a flowchart illustrating the steps involved in preparing granules, tablets, capsules, or oral liquids according to an embodiment of the present invention. Detailed Implementation

[0021] To make the objectives and advantages of the present invention clearer, the present invention will be further described below with reference to embodiments; it should be understood that the specific embodiments described herein are merely for explaining the present invention and are not intended to limit the present invention.

[0022] Preferred embodiments of the present invention will now be described with reference to the accompanying drawings. Those skilled in the art should understand that these embodiments are merely illustrative of the technical principles of the present invention and are not intended to limit the scope of protection of the present invention.

[0023] Please see Figure 1 As shown, it is a flowchart of the preparation process in this embodiment. The present invention includes: Step S1: Mix the medicinal materials and water at a material-to-liquid ratio of 1:8 to 1:12, pre-extract at a temperature of 65℃ to 75℃, and keep warm for 8 to 15 minutes to complete the low-temperature controlled pre-extraction.

[0024] Specifically, the medicinal materials in the formula include Astragalus membranaceus, Smilax glabra, Coix lacryma-jobi, Polygonum cuspidatum, Morus alba twigs, Achyranthes bidentata, Paeonia lactiflora, Salvia miltiorrhiza, and Epimedium.

[0025] Specifically, the weight proportions of the medicinal materials in the formula are as follows: Astragalus membranaceus 25-35 parts, Smilax glabra 15-25 parts, Coix lacryma-jobi 15-25 parts, Polygonum cuspidatum 8-15 parts, Morus alba twigs 8-15 parts, Achyranthes bidentata 8-12 parts, Paeonia lactiflora 8-12 parts, Salvia miltiorrhiza 5-10 parts, and Epimedium brevicornu 5-10 parts.

[0026] In this embodiment, the pre-extraction temperature is 70°C, the holding time is 12 min, and the material-to-liquid ratio is 1:10.

[0027] Optionally, the pre-extraction can be changed from constant temperature pre-extraction to gradient temperature mode, which involves holding at 65℃ for 5 minutes, 70℃ for 5 minutes, and 75℃ for 5 minutes in sequence, with a total holding time of 15 minutes. This method is suitable for formulations with more than 70% of dense root and rhizome medicinal materials, and can more fully soften the deep cell walls of the medicinal materials. Optionally, the pre-extraction can be pulsed within a temperature range of 65℃ to 75℃ with a range of ±3℃, maintaining a constant reference temperature of 70℃ and completing a temperature change operation every 3 minutes. The mass transfer efficiency can be enhanced by the convection effect generated by the temperature difference, which is suitable for formulations of medicinal materials with a high degree of cell wall lignification.

[0028] Please see Figure 2 The diagram shown illustrates the steps of temperature-controlled pre-extraction at low temperatures.

[0029] In this embodiment, step S1 includes the following specific steps: Step S11: Weigh the following medicinal materials: Astragalus membranaceus 30 parts by weight, Smilax glabra 20 parts by weight, Coix lacryma-jobi 20 parts by weight, Polygonum cuspidatum 12 parts by weight, Morus alba twigs 12 parts by weight, Achyranthes bidentata 10 parts by weight, Paeonia lactiflora 10 parts by weight, Salvia miltiorrhiza 8 parts by weight, and Epimedium brevicornu 8 parts by weight, and grind them into coarse powder of 20-40 mesh. Step S12: Add purified water at a ratio of 1:8 to 1:12 (g:mL) of total weight of medicinal materials to water volume, and stir and soak for 15 to 20 minutes; Step S13: Heat to 65℃~75℃, control the heating rate to 2℃ / min~4℃ / min, and hold at the constant temperature for 8min~15min; Step S14: During the heat preservation period, stir intermittently at a stirring speed of 40 r / min to 60 r / min. Stir for 5 minutes, then stop for 10 minutes. Repeat this process. After step S15, once the heat preservation is complete, keep the temperature of the pre-extraction solution from decreasing and proceed directly to step S2, periodic fluctuation extraction.

[0030] The function of this step is to pre-extract at a low temperature of 65℃~75℃, which moderately disrupts the pectin layer of the medicinal material cell wall based on the softening critical temperature of pectin at around 60℃, opens up the diffusion channels of active ingredients, and preferentially dissolves small molecule phenolic glycosides such as polysaccharide, paeoniflorin, and salvianolic acid. At the same time, it avoids the complete gelatinization of starch above 78℃ to form a high-viscosity colloid, thus providing a stable extraction substrate with low turbidity and low colloid content for subsequent steps.

[0031] The intercellular spaces of medicinal herbs are filled with pectin and hemicellulose. The critical temperature for pectin to melt and soften is approximately 60℃. Below 65℃, the pectin structure is relatively rigid, making it difficult for water molecules to penetrate the cell tissue, and only the active ingredients on the surface of the medicinal herb can dissolve. Above 75℃, starch granules break down in large numbers, forming a high-viscosity colloid that encapsulates the active ingredients and hinders their diffusion. The 65℃~75℃ range achieves a perfect balance of multiple effects: firstly, pectin moderately softens and breaks down cell walls, with a softening index stable between 0.50 and 0.80; secondly, starch does not undergo complete gelatinization, with a gelatinization degree below 20%; thirdly, heat-sensitive phenolic glycosides show no significant degradation, and icariin retention is above 95%; and fourthly, the endogenous cellulase and pectinase in the medicinal herb are at their peak activity, with an optimal temperature of 60℃~72℃, which can assist in the release of bound active ingredients.

[0032] Table 1: Effect of pre-extraction temperature on dissolution rate of active ingredients and system properties

[0033] Table 1 Note: The pectin softening index was calculated by measuring the change in shear modulus of the medicinal material particles in the extract using a rheometer; the starch gelatinization degree was determined by differential scanning calorimetry (DSC); the turbidity was determined by a turbidimeter (HACH2100Q); the dissolution rate = content of components in the pre-extract / total amount of components in the medicinal material × 100%.

[0034] As shown in Table 1, to further demonstrate the rationality and superiority of the pre-extraction parameter range defined in this invention, non-optimal process conditions at the critical boundary of the parameter range were selected for comparative verification. Under the premise of a unified basic extraction process, the influence of boundary parameters on the extraction system, dissolution of active ingredients, and product quality was examined. The low-temperature boundary process condition was a pre-extraction temperature of 60℃ and a holding time of 12 min. This temperature was lower than the lower limit of the process defined in this invention. The pectin of the medicinal material was not sufficiently softened, and the pectin softening index was only 0.28, which did not meet the reasonable process standard. The cell wall permeability of the medicinal material was poor, and the dissolution of active ingredients was hindered. The dissolution rate of polygalactoside was only 45.2%, and the dissolution rate of paeoniflorin was only 38.5%. The enrichment effect of active ingredients in the pre-extraction stage was poor, which greatly increased the process load of the subsequent cyclical extraction process. The polygalactoside content in the final dry extract was only 2.1 mg / g, which could not meet the quality standard of the finished product. The high-temperature boundary process conditions are a pre-extraction temperature of 78°C and a holding time of 12 minutes. This temperature is higher than the upper limit of the process specified in this invention. A large amount of starch gelatinizes in the system, with a starch gelatinization degree of 45.2%, which is far beyond the controllable process standard. The turbidity of the pre-extraction liquid is 0.85 NTU and the viscosity is 3.85 mPa·s. The high-viscosity colloidal system will encapsulate the active ingredients and inhibit the diffusion and dissolution of the ingredients. At the same time, the high temperature will cause the degradation of heat-sensitive components, and the retention rate of icariin will drop to 82.5%. In addition, the high turbidity of the extraction system will seriously interfere with the subsequent online spectral monitoring process, resulting in a decrease in the accuracy of the extraction endpoint determination and a significant reduction in the overall process stability.

[0035] Step S2: After completing the pre-extraction, raise the temperature of the high-temperature section to 85℃~95℃ and keep it at the high-temperature section for 20min~30min. Lower the temperature of the low-temperature section to 70℃~80℃ and keep it at the low-temperature section for 15min~25min. Complete one heating and cooling cycle. Repeat the cycle 2~4 times to obtain the extract.

[0036] In this embodiment, the high-temperature section has a temperature of 90°C and a holding time of 25 minutes, the low-temperature section has a temperature of 75°C and a holding time of 20 minutes, and the cycle is repeated 3 times.

[0037] Optionally, the first cycle uses a high-temperature section of 95℃ for 30 minutes and a low-temperature section of 80℃ for 20 minutes. Subsequent cycles gradually reduce the temperature of the high-temperature section from 95℃ to 90℃ to 85℃. This is suitable for formulations with a high saponin content where the total proportion of coix seed and frankincense is greater than 40%. Optionally, a pulse heating zone is set between the high-temperature and low-temperature zones. The temperature is first maintained at 90°C and then instantly raised to 100°C and held for 30 seconds before being lowered to 75°C. The steam explosion effect generated by the instantaneous high temperature enhances cell wall disruption, which is suitable for medicinal materials with a high degree of cell wall lignification.

[0038] The core mechanism of periodic fluctuation extraction lies in three points. First, the high-temperature range of 85℃ to 95℃ matches the optimal dissolution temperature of triterpenoid saponins and polysaccharides. The solubility of coixol increases significantly above 90℃, from 0.8 mg / mL at 70℃ to 2.5 mg / mL at 90℃, while astragalus polysaccharides can be fully dissolved at 95℃. Second, the low-temperature range of 70℃ to 80℃ protects heat-sensitive components. Polygonin undergoes phenolic hydroxyl oxidation when heated for a long time above 80℃, icariin's glycosidic bonds are easily broken above 85℃, and paeoniflorin lactone rings are easily hydrolyzed above 80℃; periodic cooling buffers heat-sensitive components in the low-temperature range, avoiding cumulative degradation caused by continuous high temperatures. Third, temperature gradient convection improves mass transfer efficiency. During heating, liquid expansion generates upward convection, and during cooling, liquid contraction generates downward convection. Temperature gradient-driven convection improves mass transfer efficiency by 30% to 40% compared to simple stirring.

[0039] Table 2: Effect of periodic fluctuation extraction parameters on the dissolution rate of active ingredients

[0040] Table 2 Note: Dissolution rate is the percentage of cumulative dissolution rate to the total amount of medicinal materials; Retention rate = content of components after extraction / content after pre-extraction × 100%; Traditional constant temperature decoction for 60 minutes is a common process parameter in the industry, representing the current level of technology.

[0041] As shown in Table 2, the extraction effect of active ingredients is excellent under the three sets of parameters of the periodic fluctuation extraction process of this invention. Specifically, when using the median parameter, the dissolution rate of coixol was 78.5%, the dissolution rate of astragalus polysaccharides was 82.3%, and the retention rate of icariin was 92.5%, achieving both efficient dissolution of heat-resistant components and stable retention of heat-sensitive components, resulting in the best overall extraction effect. When using the lower limit parameter, the retention rate of icariin increased to 95.8%, making it more suitable for formulations with a high proportion of heat-sensitive components. When using the upper limit parameter, the dissolution rate of coixol increased to 82.5%, and the dissolution rate of astragalus polysaccharides increased to 85.6%, making it more suitable for formulations with a high proportion of saponins and polysaccharides, demonstrating wide adaptability of the parameter range. Traditional constant-temperature decoction processes yield only 65.2% dissolution rate of coix seed saponins, 70.5% dissolution rate of astragalus polysaccharides, and only 68.5% retention rate of icariin, indicating significant shortcomings in the extraction efficiency of both types of components. High-temperature cyclic extraction without a low-temperature stage results in only 72.5% retention rate of icariin, with significant degradation of heat-sensitive components. These results demonstrate that the high-low temperature cyclical extraction process of this invention can simultaneously achieve efficient dissolution of heat-resistant components such as saponins and polysaccharides, as well as effective protection of heat-sensitive components such as phenolic glycosides and flavonoids, through alternating temperature cycles, exhibiting significant technical advantages compared to traditional processes.

[0042] Step S3: The absorbance change rate of the extract at a characteristic wavelength is monitored online using ultraviolet-visible spectrophotometry. The absorbance change rate is calculated based on continuous sampling data according to a preset sampling interval. The extraction endpoint is determined when the spectral change rate is ≤0.02.

[0043] Specifically, the characteristic wavelength is 340 nm, and the sampling interval is 3 min.

[0044] Optionally, multi-wavelength fusion judgment can be used to simultaneously monitor the SR at three wavelengths: 340nm, 230nm, and 270nm. The endpoint is determined when the SR at all three wavelengths is ≤0.02. This method is suitable for products with high quality control requirements for multiple components. Optionally, the solids change rate can be jointly determined by using a refractometer to monitor the solids content of the extract online. The endpoint is determined when the solids change rate is ≤0.5% / min and SR≤0.02, thereby improving the accuracy of the determination.

[0045] Please see Figure 3 As shown, it is a flowchart of the steps for determining the extraction endpoint.

[0046] In this embodiment, step S3 includes the following specific steps: Step S31, Set online monitoring parameters: characteristic wavelength This corresponds to the maximum absorption of polygalactoside; This corresponds to the maximum absorption of paeoniflorin; This corresponds to the characteristic absorption of total polyphenols; Step S32: Take 1 mL to 2 mL of sample from the sampling port of the extraction vessel every 2 to 5 minutes, dilute it 10 times, and then measure the absorbance at each wavelength. ; Step S33, calculate the spectral change rate SR: ,in The absorbance at the current moment. The absorbance at the previous moment; Step S34: When the SR values ​​of three consecutive measurements are all ≤0.02, the extraction endpoint is determined. Step S35: Record the total extraction time required to reach the extraction endpoint as batch quality control data.

[0047] This step aims to monitor the real-time absorbance changes of the extract online, dynamically determine the dissolution equilibrium of active ingredients, accurately control the extraction endpoint, and avoid two types of defects caused by fixed-time extraction: insufficient extraction due to residual active ingredients and degradation of heat-sensitive components caused by continuous high temperatures. During the extraction stage, active ingredients inside the medicinal material continuously dissolve into the liquid phase, the concentration of components in the system gradually increases, and the absorbance increases synchronously. After dissolution reaches equilibrium, the concentration of components in the liquid phase tends to stabilize, and the change in absorbance approaches zero. This invention uses the spectral change rate (SR) to characterize the real-time dissolution rate of the components. An SR greater than 0.02 indicates that the dissolution process is still proceeding rapidly, and the extraction is not yet sufficient; an SR less than or equal to 0.02 indicates that dissolution has basically reached equilibrium, and further extending the extraction time will not provide significant benefits and will even exacerbate the degradation of heat-sensitive components. If a fixed-time extraction mode is used, it cannot adapt to batch differences in medicinal materials. The dissolution rate of medicinal materials from different origins, harvest periods, and storage times can vary by 30% to 50%, ultimately resulting in quality fluctuations where some batches are under-extracted and some batches are over-extracted.

[0048] Table 3: Impact of Online Monitoring Parameters on Endpoint Determination

[0049] Table 3 Note: Dissolution rate is the percentage of cumulative dissolution rate to the total amount of medicinal materials; batch-to-batch variation RSD is the relative standard deviation of the polygalactosidase content in 10 batches of products; fixed extraction time is 120 min; experience judgment is determined by the operator based on experience by judging the color and odor of the extract to determine the endpoint.

[0050] As shown in Table 3, the median group with an SR threshold of 0.02 as the judgment standard has the best overall quality control effect. Under the conditions of a sampling interval of 3 min and a monitoring wavelength of 340 nm, the extraction time up to the judgment endpoint of 138 min is 92.5% for polygalactoside, 89.8% for paeoniflorin, and 90.5% for icariin. The batch-to-batch RSD is only 3.8%, which takes into account both the dissolution efficiency of active ingredients and the stability of heat-sensitive components. It is the preferred parameter suitable for most production scenarios. When the SR threshold is adjusted to 0.03, the endpoint determination time is shortened to 125 minutes, and the extraction efficiency is improved. However, the dissolution balance judgment standard is relaxed, and the dissolution rate of polygalactoside drops to 88.2% and the dissolution rate of paeoniflorin drops to 85.5%. This is only suitable for situations where production efficiency is prioritized and the upper limit of component dissolution is not high. When the SR threshold is tightened to 0.01, although the dissolution rates of polygalactoside and paeoniflorin are slightly improved, the retention rate of heat-sensitive icariin decreases significantly, and the production cycle is extended. Therefore, it is only suitable for a few scenarios with special requirements for the content of phenolic glycosides. Compared with the online dynamic monitoring scheme, both traditional judgment methods have obvious defects. Extraction at a fixed duration of 120 minutes, without considering batch-to-batch dissolution differences, resulted in a dissolution rate of only 78.5% for polygalactosidin, with a batch-to-batch RSD as high as 18.5%. Relying on operator experience based on vision and smell led to significant human error, with endpoint determination time fluctuating between 30 and 60 minutes, and a batch-to-batch RSD reaching 25.2%. The large fluctuations in polygalactosidin dissolution rate made it difficult to guarantee product quality stability. These data clearly demonstrate that using an online dynamic endpoint determination with an SR threshold of 0.02 can effectively eliminate quality fluctuations caused by batch-to-batch and human operation, and simultaneously balance the extraction effects of multiple active ingredients.

[0051] Step S4: Filter and separate the extract from the residue, and concentrate the extract under reduced pressure to a clear paste with a relative density of 1.15 to 1.25.

[0052] Specifically, the vacuum concentration temperature is 55℃~60℃, and the vacuum degree is -0.08MPa~-0.06MPa.

[0053] Optionally, a rotary thin-film evaporator can be used, with an evaporation temperature of 45℃~55℃ and a shorter concentration time (30min~40min), which is suitable for products with extremely high requirements for the protection of heat-sensitive components; Optionally, the extract can be pre-concentrated to 1 / 3 of its volume using a nanofiltration membrane, and then concentrated to the target density under reduced pressure, which can reduce energy consumption by 30% to 40% and is suitable for large-scale production.

[0054] In this embodiment, step S4 includes the following specific steps: Step S41: After extraction, filter the extract with a 200-mesh sieve while it is still hot to separate the extract from the residue. Step S42: Wash the dregs twice with 80℃ hot water, each time using water equal to the weight of the dregs, and combine the washing liquid and the extract. Step S43: Concentrate the combined liquid under reduced pressure, with the vacuum degree controlled at -0.08 to -0.06 MPa and the temperature maintained at 55°C to 60°C, until a clear paste with a relative density of 1.15 to 1.25 is obtained at 60°C. In step S44, the extract is filtered through a 120-mesh sieve to remove suspended particles, resulting in a clear concentrate.

[0055] Step S4 involves separating the residue from the extract. A low-temperature, reduced-pressure process is used to concentrate the extract to the target density, protecting heat-sensitive active ingredients such as polydipsia glycoside, paeoniflorin, and icariin throughout the process and avoiding the degradation problems associated with conventional high-temperature concentration. Reduced-pressure concentration lowers the boiling point of water by reducing the vacuum level. When the vacuum level is maintained at -0.07 MPa, the boiling point of water can be lowered to around 62°C. This allows the concentration temperature to be stably controlled at 55°C to 60°C, far lower than the high-temperature environment of 95°C to 100°C required for open-air concentration. Prolonged heating above 70°C accelerates the oxidation of the phenolic hydroxyl groups of polydipsia glycoside, paeoniflorin lactone rings are prone to hydrolysis above 80°C, and icariin glycosidic bonds are easily broken above 85°C. The low-temperature range of 55°C to 60°C in this invention can stably control the heat-sensitive components within safe heating conditions. Step S4 limits the concentration temperature to 55℃~60℃ and the vacuum degree to -0.08MPa~-0.06MPa. Under this parameter range, the retention rate of polygalactoside can be stably maintained at 93% to 97%, with a measured average of 95.5%, and the retention rate of paeoniflorin can be stably maintained at 92% to 96%, with a measured average of 94.2%. In contrast, the open-air high-temperature concentration process results in severe loss of heat-sensitive components, with a polygalactoside retention rate of only 62.5%. This fully demonstrates the protective advantage of the low-temperature vacuum concentration parameter range of this invention for active ingredients.

[0056] Table 4: Effect of Concentration Parameters on Retention Rate of Active Ingredients

[0057] Table 4 Note: Retention rate = Component content in concentrated extract / Component content in extract before concentration × 100%; Open-air concentration is carried out by open-air heating concentration (atmospheric pressure, 95℃); Reduced pressure concentration at 70℃ is too high.

[0058] It can be seen from Table 4 that the present invention defines the vacuum concentration parameter interval of a concentration temperature of 55°C to 60°C and a vacuum degree of -0.08MPa to -0.06MPa, which can maximize the retention of heat-sensitive active ingredients while taking concentration efficiency into account. The median condition of the interval is a concentration temperature of 58°C and a vacuum degree of -0.07MPa. After concentration, the retention rates of polydatin in the clear paste, paeoniflorin and icariin are 95.5%, 94.2% and 97.8% respectively. The comprehensive retention effect of the components is balanced, which meets the production requirements of most products; the low-temperature and high-vacuum condition at the lower limit of the interval is 55°C and -0.08MPa, with a lower heating temperature. The retention rate of polydatin can reach 97.2% and the retention rate of paeoniflorin can reach 96.5%, which is more suitable for products with strict standards on the content of heat-sensitive components; the high-temperature and low-vacuum condition at the upper limit of the interval is 60°C and -0.06MPa, which can greatly shorten the concentration time, is preferentially suitable for working conditions pursuing production efficiency, and the retention level of active ingredients is still in the qualified range. Comparing the two groups of control processes, the damaging effect of high temperature on heat-sensitive components can be clearly seen. Open concentration at 95°C under atmospheric pressure lacks low-temperature vacuum protection, and the retention rates of polydatin, paeoniflorin and icariin drop significantly under long-term high temperature environment, which are only 62.5%, 58.2% and 55.8% respectively; even if a reduced pressure system is used but the temperature is increased to 70°C, the retention rate of polydatin is only 85.2%, which is far lower than the retention level in the low temperature range of the present invention. The above data verify that the reduced pressure concentration scheme of controlling the concentration temperature within 60°C and matching with the corresponding vacuum degree is a key condition for reducing the degradation of heat-sensitive components and stabilizing the finished product indicators.

[0059] In step S5, the LPS-induced RAW264.7 cell NO release inhibition model is used to detect the in vitro anti-inflammatory activity of the clear paste, and IC50 ≤ 50μg / mL is regarded as qualified; meanwhile, HPLC is used to determine the contents of polydatin and paeoniflorin, and polydatin ≥ 2.8mg / g and paeoniflorin ≥ 1.5mg / g are regarded as qualified.

[0060] In this example, during the detection in step S5, the clear paste is temporarily stored at 4°C, and enters the drying process of step S6 within 24 hours after passing the detection, so as to avoid the degradation of active ingredients during the storage of the clear paste Optionally, on the basis of polydatin and paeoniflorin, adding icariin ≥ 1.0mg / g and salvianolic acid B ≥ 0.8mg / g as quality control indicators is applicable to scenarios with higher requirements for product activity; Optionally, in addition to in vitro anti-inflammatory activity, carrageenan-induced rat paw edema model is used to verify in vivo anti-inflammatory activity, and inhibition rate ≥ 50% is regarded as qualified, which is suitable for new drug declaration or quality control of high-end products.

[0061] In this embodiment, the specific steps of step S5 include: In step S51, an appropriate amount of the clear paste obtained in step S4 is taken, dried under reduced pressure to obtain dry extract, crushed and passed through an 80-mesh sieve; Step S52: The contents of polydipsia glycoside and paeoniflorin in the dry extract were determined by HPLC: the chromatographic conditions were C18 column, mobile phase elution of acetonitrile to 0.1% phosphoric acid water gradient, detection wavelength of 340 nm corresponding to polydipsia glycoside and 230 nm corresponding to paeoniflorin. Step S53: The in vitro anti-inflammatory activity was detected using an LPS-induced NO release inhibition model in RAW264.7 cells. The dry extract was prepared into a series of concentrations of 10, 25, 50, and 100 μg / mL and added to the LPS-stimulated RAW264.7 cell culture system. After culturing for 24 h, the NO content in the culture medium was measured, the inhibition rate at different concentrations was calculated, and the half-maximal inhibitory concentration (IC50) was calculated by nonlinear fitting. Step S54: The judgment criteria are ≥2.8mg / g for polygalactoside, ≥1.5mg / g for paeoniflorin, and ≤50μg / mL for IC50. If all three criteria are met, the batch is released; if any one criterion is not met, the batch must be reworked or scrapped.

[0062] Step S5 involves simultaneous testing of both chemical and biological activity indicators to construct a composite quality control system linking components and activities. This ensures that the chemical components of each batch of products meet standards and that the anti-inflammatory activity remains stable, thereby stabilizing clinical efficacy. The efficacy of traditional Chinese medicine compound formulas is produced synergistically by multiple active ingredients. Relying solely on the content of a single ingredient cannot fully reflect the overall efficacy. Polygonum cuspidatum glycoside and paeoniflorin are the core anti-inflammatory substances in this compound formula. Icariin, salvianolic acid, and astragalus polysaccharides also possess synergistic anti-inflammatory effects. If process defects cause the loss of synergistic active ingredients, even if the content of polygonum cuspidatum glycoside and paeoniflorin meets the standards, the overall anti-inflammatory activity will still be deficient. Under the corresponding control process, polygonum cuspidatum glycoside (3.2 mg / g) and paeoniflorin (2.0 mg / g) both meet the content limits, but the IC50 reaches 55.2 μg / mL, indicating that the activity indicator is unqualified. This step combines in vitro anti-inflammatory IC50 of ≤50μg / mL with chemical indicators such as polygaloside content of ≤2.8mg / g and paeoniflorin content of ≤1.5mg / g for joint quality control. By using in vitro IC50 anti-inflammatory detection, the synergistic effect of various active ingredients in the compound is comprehensively characterized, completing the upgrade from simple ingredient content quality control to activity-oriented quality control, effectively avoiding the shortcomings of traditional quality control models where ingredient content meets the standards but overall efficacy is insufficient.

[0063] Table 5: Batch Quality Control Test Results

[0064] It can be known from Table 5 that the present invention adopts a dual quality control standard combining chemical indicators and in vitro anti-inflammatory activity IC50, which can completely distinguish the quality of products from different batches and effectively avoid judgment deviations caused by single-content quality control. All indicators of the three groups of samples within the qualified range meet the standard requirements: the median batch contains 3.5 mg / g of polydatin, 2.2 mg / g of paeoniflorin, 98.5 mg / g of total saponins, and an IC50 of 42.5 μg / mL. All indicators are balanced and up to standard, which is a qualified batch for conventional production; the upper limit batch has higher contents of various active ingredients, with an IC50 as low as 38.5 μg / mL, showing the optimal anti-inflammatory activity; the indicators of the lower limit batch are close to the qualified limit, containing 3.2 mg / g of polydatin, 1.9 mg / g of paeoniflorin, and an IC50 of 48.2 μg / mL, which still meets the release standard. The two groups of unqualified batches directly reflect the advantages of the dual quality control system: for the batch with unqualified chemical indicators, the contents of polydatin and paeoniflorin do not meet the limited standard, the content of total saponins is low, and the IC50 is as high as 68.5 μg / mL. Both the components and the activity are not up to standard, and it can be directly determined to be reworked; for the batch with unqualified activity indicators, the contents of polydatin and paeoniflorin both meet the chemical standard, but the IC50 reaches 55.2 μg / mL, which exceeds the activity limit. Relying only on single-content detection would misjudge this batch as a qualified product. The root cause is that excessive high-temperature duration during the extraction stage causes degradation of synergistic anti-inflammatory components, leading to a decrease in overall anti-inflammatory efficacy. The above detection results fully show that the combined quality control mode combining component content and in vitro anti-inflammatory activity can accurately identify unqualified batches with qualified components but insufficient efficacy, ensuring stable clinical anti-inflammatory effect of the finished product.

[0065] Step S6: drying the clear ointment to obtain dry extract, pulverizing the dry extract, adding pharmaceutically acceptable excipients, and preparing into granules, tablets, capsules or oral liquids.

[0066] Specifically, the drying is vacuum drying, the drying temperature is 60°C to 70°C, and the vacuum degree is -0.08MPa to -0.06MPa.

[0067] Optionally, the clear ointment is spray-dried, with an inlet air temperature of 180°C and an outlet air temperature of 80°C, and the material stays in the drying tower for about 30s. The obtained dry powder can be directly packaged or granulated. Spray drying has a fast speed and is suitable for large-scale production, but the outlet air temperature of 80°C is relatively high, and the retention rate of heat-sensitive components is slightly lower than that of vacuum drying; Optionally, the clear ointment is pre-frozen and then subjected to vacuum freeze-drying, with a temperature range of -40°C to 30°C, a vacuum degree of less than 10Pa, and a drying duration controlled at 48h to 72h. The freeze-dried product has the optimal retention effect of active ingredients, but the production cost is relatively high, which is suitable for the preparation of high-end products.

[0068] Please refer to Figure 4 , which is a flow chart of the steps for preparing granules, tablets, capsules or oral liquids.

[0069] In this embodiment, step S6 includes the following specific steps: Step S61: Place the clear paste obtained in step S4 into a vacuum drying oven, set the temperature to 60℃~70℃, the vacuum degree to -0.08~-0.06MPa, and dry for 8h~12h until the moisture content is ≤5.0%; Step S62: The dried extract is pulverized and passed through an 80-mesh sieve to obtain dry extract powder; Step S63: Mix dry extract powder: microcrystalline cellulose: lactose in a ratio of 5:2:3, add an appropriate amount of 70% ethanol to granulate, pass through a 16-mesh sieve, and dry at 60℃ until the moisture content is ≤3.0%; Step S64: Granulate, dispense, and prepare into granules; or compress into tablets, or fill into capsules.

[0070] The function of this step is to dry the extract into a dry extract for subsequent processing into a pharmaceutical dosage form suitable for clinical use. Simultaneously, it utilizes low-temperature drying conditions to protect the heat-sensitive active ingredients. This step limits the vacuum drying temperature to the range of 60℃ to 70℃, which is significantly lower than the high-temperature environment of 80℃ to 100℃ achieved by hot air drying. Furthermore, the low oxygen content within the vacuum system reduces the oxidative degradation of active ingredients. By reducing system pressure to achieve low-temperature evaporation of moisture, the retention rates of polygalactoside and paeoniflorin can be stably maintained above 95%. Actual measurements show a polygalactoside retention rate of 96.5% and a paeoniflorin retention rate of 95.8%. In contrast, hot air drying causes significant loss of active ingredients due to high temperatures; under hot air drying conditions, the polygalactoside retention rate is only 68.5%. Therefore, the 60℃ to 70℃ vacuum drying range can maintain the stability of heat-sensitive components such as polygalactoside, paeoniflorin, and icariin, significantly improving the final retention levels of various active ingredients.

[0071] Table 6: Effect of Drying Method on Retention Rate of Active Ingredients

[0072] Table 6 Note: Retention rate = Content of ingredients after drying / Content of ingredients in the paste before drying × 100%.

[0073] Table 6 shows that the three drying methods differ significantly in terms of heating conditions, processing time, retention of active ingredients, and formulation performance. Vacuum drying, using a low-temperature vacuum environment of 65℃, took 10 hours, achieving the best retention rates for heat-sensitive components: 96.5% for polygalactoside, 95.8% for paeoniflorin, and 98.2% for icariin. The resulting dry extract had a moisture content of 3.5%, and the granulation time was 3.5 minutes, resulting in balanced overall quality and making it the preferred drying process for this invention. Spray drying, with an inlet air temperature of 180℃ and an outlet air temperature of 80℃, completed drying in just 30 seconds, demonstrating high production efficiency. The dry powder had a moisture content of 4.2%, and the granule dissolving speed was [not specified]. The fastest drying time is 2.8 minutes, but the outlet air temperature is relatively high, and the retention rate of the three types of active ingredients is slightly lower than that of vacuum drying, making it suitable for large-scale industrial production. Hot air drying maintains a high temperature of 90℃ throughout the process and the drying time reaches 24 hours. The prolonged high-temperature environment severely damages the heat-sensitive active ingredients. The retention rates of polygalactoside, paeoniflorin, and icariin are only 68.5%, 62.5%, and 55.8%, respectively. At the same time, the moisture content of the dry extract is too high, reaching 5.5%, and the particle dissolution time is extended to 5.2 minutes, resulting in more impurities and by-products and poorer product quality. The above data fully demonstrate that a low-temperature vacuum environment can significantly reduce the oxidation and decomposition of polygalactoside, paeoniflorin, and icariin, and has irreplaceable advantages in protecting the active substances and stabilizing the properties of the formulation.

[0074] Specifically, the traditional Chinese medicine composition prepared in this embodiment contains ≥2.8 mg / g of polygalactoside, ≥1.5 mg / g of paeoniflorin, and ≤50 μg / mL of in vitro anti-inflammatory IC50.

[0075] Specifically, this embodiment describes the application of the traditional Chinese medicine composition prepared in the preparation of a drug for treating joint diseases caused by damp-heat obstruction.

[0076] Specifically, this embodiment describes the application of the traditional Chinese medicine composition prepared in the preparation of anti-inflammatory, analgesic, and immunomodulatory drugs.

[0077] Example 1

[0078] Formula: Astragalus membranaceus 300g, Smilax glabra 200g, Coix lacryma-jobi 200g, Polygonum cuspidatum 120g, Morus alba twig 120g, Achyranthes bidentata 100g, Paeonia lactiflora 100g, Salvia miltiorrhiza 80g, Epimedium brevicornu 80g.

[0079] Preparation process: S1: Grind to 30 mesh, material-to-liquid ratio 1:10, pre-extract at 70℃ for 12 min; S2: Extraction with cyclic fluctuations at 90℃ for 25 min + 75℃ for 20 min, repeated 3 times; S3: Online monitoring of absorbance at 340nm, SR≤0.02 to determine the endpoint, endpoint time 138min; S4: Filtered through a 200-mesh filter and concentrated under reduced pressure at 58°C to a relative density of 1.20; S5: polydatin in dry extract 3.5 mg / g, paeoniflorin 2.2 mg / g, IC50 42.5 μg / mL, all three items are qualified; S6: vacuum drying at 65°C for 10 h, granulating to obtain granules, 5 g per bag.

[0080] Quality indicators: yield of dry extract 18.5%, polydatin 3.5 mg / g, paeoniflorin 2.2 mg / g, total saponins 98.5 mg / g, icariin 1.8 mg / g, in vitro anti-inflammatory IC50 42.5 μg / mL, inter-batch difference RSD 3.8%.

[0081] Example 2 (production region in southern China)

[0082] The formulation is the same as in Example 1, and the medicinal materials are produced in Guizhou (Astragali Radix), Yunnan (Flemingiae Radix) and Guangxi (Coicis Semen).

[0083] Preparation process parameters: S1 pre-extraction temperature 65°C, time 15 min (the texture of medicinal materials is relatively dense); S2 periodic fluctuation 85°C×30 min + 70°C×25 min, 2 cycles (the proportion of heat-sensitive components is slightly higher); S3 end point time 125 min, and the remaining steps are the same as in Example 1.

[0084] Quality indicators: polydatin 3.8 mg / g, paeoniflorin 2.5 mg / g, total saponins 105.2 mg / g, IC50 38.5 μg / mL. It proves that the method of the present invention has good adaptability to medicinal materials from different producing regions.

[0085] Example 3 (formulation with high saponin content)

[0086] Formulation adjustment: Astragali Radix 350 g, Coicis Semen 250 g, Flemingiae Radix 250 g (increasing the proportion of saponin-containing medicinal materials).

[0087] Preparation process parameters: S1 pre-extraction temperature 75°C, time 8 min; S2 periodic fluctuation 95°C×20 min + 80°C×15 min, 4 cycles, and the remaining steps are the same as in Example 1.

[0088] Quality indicators: polydatin 3.2 mg / g, paeoniflorin 1.9 mg / g, total saponins 125.8 mg / g, IC50 48.2 μg / mL. It is suitable for application scenarios with high requirements for total saponin content.

[0089] Example 4: In vivo anti-inflammatory activity verification (carrageenan-induced rat paw edema model) 4.1 Experimental method Sixty male SPF-grade SD rats, weighing 180–220 g, were randomly divided into six groups of 10 rats each: (1) Blank control group (0.5% CMC-Na solution); (2) Positive control group (indomethacin 10 mg / kg); (3) Low-dose group of Example 1 (raw drug 5 g / kg); (4) Medium-dose group of Example 1 (raw drug 10 g / kg); (5) High-dose group of Example 1 (raw drug 20 g / kg); (6) Comparative Example 1 (traditional decoction process, raw drug 10 g / kg). Each group was administered the drug by gavage once a day for 7 consecutive days. One hour after the last administration, 0.1 mL of 1% carrageenan was injected subcutaneously into the right hind paw of the rats to induce inflammation. The volume of the right hind paw was measured before inflammation and at 1, 2, 3, 4, and 6 hours after inflammation, and the paw swelling rate and swelling inhibition rate were calculated.

[0090] Please refer to Table 7, which shows the data for in vivo anti-inflammatory activity validation. Table 7: Data on in vivo anti-inflammatory activity validation

[0091] Note: Compared with the blank control group, *P<0.05, **P<0.01; Swelling inhibition rate = (swelling rate of blank group - swelling rate of drug-treated group) / swelling rate of blank group × 100%.

[0092] 4.3 Results Analysis In Example 1, the medium-dose group (10 g / kg of raw herb) showed a foot swelling rate of 35.2% and a swelling inhibition rate of 51.3% 4 hours after inflammation induction, which was significantly different from the blank control group (P<0.01). The high-dose group (20 g / kg of raw herb) in Example 1 achieved a swelling inhibition rate of 60.2%, which was not significantly different from the positive control indomethacin (65.1%) (P>0.05). The comparative example 1 group (traditional decoction process, 10 g / kg of raw herb) showed a swelling inhibition rate of only 22.8%, significantly lower than the medium-dose group of this invention (P<0.05), and a 55.6% decrease in inhibition rate compared to the Example 1 group (medium dose, same amount of raw herb). These experimental results demonstrate that the traditional Chinese medicine composition prepared by the method of this invention has significant in vivo anti-inflammatory activity, and its effect is significantly better than that of the traditional decoction process.

[0093] Example 5: Verification of analgesic activity (acetic acid writhing test) 5.1 Experimental Methods Sixty SPF-grade KM mice, half male and half female, weighing 18-22g, were randomly divided into 6 groups of 10 mice each: (1) blank control group (0.5% CMC-Na solution); (2) positive control group (aspirin 200mg / kg); (3) low-dose group of Example 1 (raw drug 5g / kg); (4) medium-dose group of Example 1 (raw drug 10g / kg); (5) high-dose group of Example 1 (raw drug 20g / kg); (6) comparative example 1 group (traditional decoction process, raw drug 10g / kg). Each group was administered the drug by gavage for 5 consecutive days, once a day. One hour after the last administration, 0.2mL of 0.6% acetic acid solution was injected intraperitoneally per mouse. The number of writhing reactions within 5-20 minutes after injection was recorded (abdominal concavity, trunk and hind leg extension were defined as one writhing reaction), and the writhing inhibition rate was calculated.

[0094] See Table 8 for the data on analgesic activity validation.

[0095] Table 8: Data on Analgesic Activity Validation

[0096] Note: Compared with the blank control group, *P<0.05, **P<0.01; writhing inhibition rate = (number of writhing in the blank group - number of writhing in the drug-treated group) / number of writhing in the blank group × 100%.

[0097] 5.3 Results Analysis In Example 1, the medium-dose group (10 g / kg of raw herb) had an average of 15.2 writhing episodes and an inhibition rate of 53.2%, which was significantly different from the blank control group (P<0.01). The high-dose group in Example 1 (20 g / kg of raw herb) showed a writhing inhibition rate of 67.7%, which was not significantly different from the positive control aspirin (73.8%) (P>0.05). Comparative Example 1 (traditional decoction process, 10 g / kg of raw herb) showed a writhing inhibition rate of only 18.5%, a decrease of 65.2% compared to the medium-dose group in Example 1. These experimental results demonstrate that the traditional Chinese medicine composition prepared by the method of this invention has significant analgesic activity.

[0098] Example 6: Validation of the efficacy of adjuvant-induced arthritis rat model 6.1 Experimental Methods Seventy male SPF-grade SD rats, weighing 160-200g, were randomly divided into seven groups of 10 rats each after one week of acclimatization: (1) normal control group; (2) model control group (complete Freund's adjuvant + 0.5% CMC-Na); (3) positive control group (methotrexate 0.5mg / kg, twice a week); (4) low-dose group of Example 1 (raw drug 5g / kg); (5) medium-dose group of Example 1 (raw drug 10g / kg); (6) high-dose group of Example 1 (raw drug 20g / kg); (7) comparative example 1 (traditional decoction process, raw drug 10g / kg).

[0099] Except for the normal control group, rats in each group were injected intradermally with 0.1 mL of complete Freund's adjuvant in the right hind paw to induce inflammation. The drug was administered by gavage starting on day 8 post-immunization, once daily for 21 consecutive days. Bilateral paw volume was measured weekly, and an arthritis index score (0–4 points) was calculated. Serum levels of TNF-α, IL-6, and IL-1β were measured at the end of the experiment.

[0100] Please refer to Table 9, which is a data table for verifying the efficacy of adjuvant-induced arthritis in rat models.

[0101] Table 9: Data on efficacy validation of adjuvant-induced arthritis rat model

[0102] Note: Compared with the normal control group, #P<0.05, ##P<0.01; compared with the model control group, *P<0.05, **P<0.01.

[0103] 6.3 Results Analysis After injection of complete Freund's adjuvant, rats in the model control group showed obvious primary paw edema (82.5%) and secondary paw edema (65.8%), with an arthritis index of 3.6. Serum TNF-α, IL-6, and IL-1β levels were significantly elevated (P<0.01 compared with the normal control group), indicating successful model establishment.

[0104] In Example 1, the primary foot edema rate (32.5%), secondary foot edema rate (22.8%), and arthritis index (1.5) in the medium-dose group (10 g / kg of raw drug) were significantly different from those in the model control group (P < 0.01). Serum levels of inflammatory factors TNF-α (145.5 pg / mL), IL-6 (92.5 pg / mL), and IL-1β (68.5 pg / mL) were significantly reduced (P < 0.01). The high-dose group (20 g / kg of raw drug) in Example 1 showed even better results, with no significant differences in any indicators compared to the positive control group (methotrexate 0.5 mg / kg) (P > 0.05).

[0105] In Comparative Example 1 (traditional decoction process, same raw herb dosage 10g / kg), the rates of primary foot edema (65.8%), secondary foot edema (52.5%), and arthritis index (3.2) were significantly higher than those in the dosage group of Example 1 (P<0.05), and serum inflammatory factor levels were also significantly higher. This confirms that the process parameters of the present invention have a significantly better protective effect on the anti-inflammatory activity of the traditional Chinese medicine compound than the traditional decoction process.

[0106] This experiment also verified the advantages of the process parameter coupling of the method of the present invention: the whole-link low temperature protection system of S1 pre-extraction → S2 periodic fluctuation extraction → S3 online monitoring → S4 low temperature concentration → S6 vacuum drying ensures that the retention rate of thermosensitive anti-inflammatory active ingredients such as polysaccharide, paeoniflorin, and icariin is ≥95%, so that the total saponin content (98.5mg / g) and IC50 (42.5μg / mL) reach the optimal level, which ultimately manifests as a significant therapeutic effect on adjuvant arthritis rats.

[0107] Comparative Example 1 (Traditional Boiling Process)

[0108] The formulation is the same as in Example 1.

[0109] Preparation process: Boil twice with water, 1 hour each time, combine the decoctions, concentrate in an open container to a relative density of 1.20, dry with hot air (90℃×24h), and pulverize into granules.

[0110] Quality indicators: Polygonin 2.1 mg / g (<2.8 mg / g, unqualified), Paeoniflorin 1.3 mg / g (<1.5 mg / g, unqualified), Total saponins 68.5 mg / g, IC50 72.5 μg / mL (50 μg / mL, unqualified). All three indicators failed to meet the standards, proving that traditional decoction processes cannot meet the quality control requirements of this compound formula.

[0111] Comparative Example 2 (without S1 pre-extraction)

[0112] The formulation is the same as in Example 1.

[0113] Preparation process: Direct S2 periodic fluctuation extraction (90℃×25min+75℃×20min×3 times), without S1 pre-extraction step.

[0114] Quality indicators: Polygonin 2.8 mg / g (just met the standard), paeoniflorin 1.6 mg / g (just met the standard), but the pre-extraction solution had high turbidity (0.45 NTU), the S3 online monitoring spectrum was interfered with, the endpoint was determined earlier (110 min), and the IC50 reached 58.5 μg / mL (50 μg / mL is unacceptable). This proves that S1 pre-extraction plays a key role in reducing system turbidity and ensuring the accuracy of online monitoring.

[0115] Comparative Example 3 (No S3 online monitoring, fixed time extraction)

[0116] The formula is the same as that in Example 1.

[0117] Preparation process: Steps S1 and S2 are the same as those in Example 1, except that the extraction time is fixed at 120 min in S2, and there is no online monitoring for end point determination.

[0118] Quality indicators: polydatin content is 2.5 mg / g (product is unqualified if content < 2.8 mg / g), paeoniflorin content is 1.4 mg / g (product is unqualified if content < 1.5 mg / g), and the inter-batch difference RSD is 22.5% (much higher than 3.8% of Example 1). This proves that end point determination via online monitoring plays a key role in stabilizing product quality.

[0119] Comparative Example 4 (without S5 activity quality control, only chemical quality control)

[0120] The formula is the same as that in Example 1.

[0121] Preparation process: Steps S1 to S4 are the same as those in Example 1, except that only contents of polydatin and paeoniflorin are detected in S5, and in vitro anti-inflammatory activity is not detected.

[0122] Results: In a certain batch, the polydatin content is 3.2 mg / g and the paeoniflorin content is 2.0 mg / g, both meeting the standards, and the batch is determined as qualified and released according to traditional quality control standards. However, the actual IC50 reaches 55.2 μg / mL (product is unqualified if IC50 > 50 μg / mL), resulting in unstable clinical efficacy. This proves the necessity of activity-oriented quality control.

[0123] Comparative Example 5 (high-temperature concentration and drying)

[0124] The formula is the same as that in Example 1.

[0125] Preparation process: Open concentration is performed in S4 (95°C, 90 min), and hot-air drying is performed in S6 (90°C, 24 h).

[0126] Quality indicators: polydatin content is 1.8 mg / g (product is unqualified if content < 2.8 mg / g), paeoniflorin content is 0.9 mg / g (product is unqualified if content < 1.5 mg / g), icariin content is 0.5 mg / g (severely degraded). High-temperature concentration and drying cause massive loss of heat-sensitive components, which proves the necessity of low-temperature concentration and drying.

[0127] Comparative Example 6 (constant-temperature decoction vs. periodic fluctuation extraction)

[0128] The formula is the same as that in Example 1.

[0129] Preparation process: Step S1 is the same as that in Example 1; constant-temperature heating at 95°C for 60 min is adopted in S2 (instead of periodic fluctuation extraction); steps S3, S4 to S6 are the same as those in Example 1.

[0130] Quality indicators: Polygonin 2.2 mg / g, Paeoniflorin 1.2 mg / g, Total saponins 72.5 mg / g, Icariin 0.8 mg / g, IC50 68.5 μg / mL. Compared with periodic fluctuation extraction, constant temperature decoction showed lower retention of heat-sensitive components (icariin 0.8 mg / g vs 1.8 mg / g, a decrease of 55.6%), lower saponin dissolution rate (72.5 mg / g vs 98.5 mg / g, a decrease of 26.4%), and insufficient anti-inflammatory activity (IC50 68.5 μg / mL vs 42.5 μg / mL, a decrease of 38.0%). This demonstrates the necessity of periodic fluctuation extraction to balance the dissolution of heat-resistant and heat-sensitive components.

[0131] Please refer to Table 10, which is a comprehensive data table of overall beneficial effects. Table 10: Overall Beneficial Effects Summary Table

[0132] Table 10 Note: Dry extract yield = dry extract weight / total weight of medicinal materials × 100%; batch-to-batch variation RSD is the relative standard deviation of polygalactosidin content in 10 batches of products; the smaller the IC50 value, the stronger the anti-inflammatory activity.

[0133] Analysis of the data in Table 10 shows that, compared with the traditional decoction method in Comparative Example 1, Example 1 showed a 66.7% increase in polygalactoside content (3.5 mg / g and 2.1 mg / g, respectively), a 69.2% increase in paeoniflorin content (2.2 mg / g and 1.3 mg / g, respectively), a 43.8% increase in total saponin content (98.5 mg / g and 68.5 mg / g, respectively), a 41.4% increase in in vitro anti-inflammatory activity, and IC50 values ​​of 42.5 μg / mL and 72.5 μg / mL, respectively. This demonstrates that the method of the present invention is significantly superior to the traditional decoction process. Compared with the constant-temperature decoction method in Comparative Example 6, Example 1 showed a 125% increase in icariin content. The percentages were 1.8 mg / g and 0.8 mg / g, respectively, demonstrating that periodic fluctuation extraction has a significant effect on protecting heat-sensitive components. The total saponin content increased by 35.9%, with values ​​of 98.5 mg / g and 72.5 mg / g, respectively, demonstrating that periodic fluctuation extraction promotes the dissolution of heat-resistant components. The batch-to-batch RSD of Example 1 was 3.8%, significantly lower than 22.5% of Comparative Example 1 and 18.5% of Comparative Example 6, demonstrating that the method of the present invention has excellent process stability. Examples 2 and 3 verified the adaptability of the method to medicinal materials from different origins and different formulation ratios. The three key indicators of polygalactoside, paeoniflorin, and IC50 were all qualified.

[0134] Explanation of the overall beneficial effects

[0135] I. Synergistic Anti-inflammatory Mechanism of Multiple Active Ingredients This compound achieves comprehensive and efficient extraction of multiple active ingredients, including phenolic glycosides, flavonoids, saponins, and polysaccharides, through parameterized control of S1 pre-extraction and S2 periodic fluctuation extraction. (1) Phenolic glycosides such as glutinin, paeoniflorin, and salvianolic acid inhibit the NF-κB pathway and reduce the expression of inflammatory factors such as TNF-α, IL-6, and IL-1β, with IC50 values ​​of 12.5, 15.8, and 18.2 μg / mL, respectively, which are the core material basis for their in vitro anti-inflammatory activity. (2) Flavonoids such as icariin and astragaloside can regulate immune cell function and inhibit the release of inflammatory mediators, and synergistically enhance the anti-inflammatory effect with phenolic glycosides (combination drug index CI<0.8, synergistic effect). (3) Triterpenoid saponins such as coix seed saponin, smilax saponin, and mulberry branch saponin exert their anti-inflammatory and analgesic effects by inhibiting the expression of COX-2 and iNOS and reducing the production of PGE2 and NO. (4) Astragalus polysaccharide and Achyranthes bidentata polysaccharide enhance the body’s anti-inflammatory and immune capabilities and prolong the duration of anti-inflammatory effects by regulating the Th1 / Th2 balance.

[0136] II. Principle of Cooperative Coupling of Process Parameters The parameter systems of each step in this invention are coupled with each other to form a self-consistent design logic: (1) The pre-extraction temperature of S1 (65℃~75℃) is matched with the temperature gradient of S2 periodic fluctuation extraction: S1 is above the critical temperature for pectin softening (60℃) and below the critical temperature for starch gelatinization (78℃), which opens up the diffusion channels of components without producing high-viscosity colloids, providing a low-turbidity base for S2; the high temperature range of S2 (85℃~95℃) matches the optimal dissolution temperature of saponins and polysaccharides, and the low temperature range (70~80℃) protects heat-sensitive components. (2) Matching S3 online monitoring with S2 periodic fluctuations: After each heating and cooling cycle in the periodic fluctuation extraction, SR shows a phased downward trend. After the third cycle, SR stabilizes at ≤0.02, accurately determining the extraction endpoint; (3) The composition protection logic of S4 low-temperature concentration and S6 low-temperature drying is consistent with that of S1 to S2: the temperature throughout the process is ≤70℃, and the retention rate of heat-sensitive components is >90% throughout the process; (4) S5 dual quality control and process parameters: the content of glutinin reflects the pre-extraction effect of S1, the content of icariin reflects the protection effect of S2 periodic fluctuation, and IC50 reflects the overall synergistic activity of components.

[0137] III. Principle of Self-Consistency of Parameter System The parameter ranges for each step of this invention have been systematically optimized to form a mutually matching and logically self-consistent parameter system: (1) S1 pre-extraction temperature 65℃~75℃: below 65℃, pectin softening is insufficient, above 75℃, starch gelatinization is excessive; (2) The number of S2 cycle fluctuations is 2 to 4 times: less than 2 times means that the saponins are not fully dissolved, and more than 4 times means that the degradation of heat-sensitive components increases (the retention rate of icariin after 4 cycles is 88.2% vs. 92.5% after 3 cycles). (3) S3 SR threshold 0.02: if it is higher than 0.02, the extraction is insufficient (the dissolution rate of citronella glycoside is 88.2% when SR=0.03), and if it is lower than 0.01, the extraction is excessive (the retention rate of icariin is 85.2% when SR=0.01). (4) S4~S6 temperature ≤70℃: If the temperature is higher than 70℃, the heat-sensitive components will degrade during the concentration and drying stages (70℃ concentration of polygalactoside retention rate is 85.2% vs 58℃ 95.5%).

[0138] The boundary values ​​of the above parameter ranges were determined through a large number of experiments, and the parameters were precisely matched. Any parameter exceeding the range will lead to a decrease in the quality of the final product.

[0139] The technical solution of the present invention has been described above with reference to the preferred embodiments shown in the accompanying drawings. However, it will be readily understood by those skilled in the art that the scope of protection of the present invention is obviously not limited to these specific embodiments. Without departing from the principles of the present invention, those skilled in the art can make equivalent changes or substitutions to the relevant technical features, and the technical solutions after these changes or substitutions will all fall within the scope of protection of the present invention.

Claims

1. A method for preparing a traditional Chinese medicine composition for treating joint diseases caused by damp-heat obstruction, characterized in that, comprising the following steps: mixing the prescription medicinal materials with water at a solid-liquid ratio of 1:8 to 1:12, controlling the pre-extraction temperature at 65°C to 75°C and keeping the temperature for 8 min to 15 min to complete temperature-controlled pre-extraction at low temperature; after completing the pre-extraction, raising the temperature of the high-temperature section to 85°C to 95°C, keeping the temperature in the high-temperature section for 20 min to 30 min, lowering the temperature of the low-temperature section to 70°C to 80°C, keeping the temperature in the low-temperature section for 15 min to 25 min to complete one temperature rise-fall cycle, repeating the cycle for 2 to 4 times to obtain an extract; adopting ultraviolet-visible spectrophotometry to monitor the change rate of absorbance of the extract at the characteristic wavelength online, calculating the absorbance change rate based on continuous sampling data according to a preset sampling interval, and determining the extraction end point when the spectral change rate is ≤0.02; filtering and separating the extract from medicinal residues, and concentrating the extract under reduced pressure to obtain a clear paste with a relative density of 1.15 to 1.25; adopting an LPS-induced RAW264.7 cell NO release inhibition model to detect the in vitro anti-inflammatory activity of the clear paste, and half maximal inhibitory concentration (IC50) ≤50 μg / mL is qualified; meanwhile, adopting HPLC to determine the contents of polydatin and paeoniflorin, and polydatin ≥2.8 mg / g and paeoniflorin ≥1.5 mg / g are qualified; drying the clear paste to obtain a dry extract, pulverizing the dry extract, adding pharmaceutically acceptable excipients, and preparing into granules, tablets, capsules or oral liquids.

2. The method for preparing the traditional Chinese medicine composition for treating joint diseases caused by damp-heat obstruction according to claim 1, characterized in that, the prescription medicinal materials comprise Astragali Radix, Flemingiae Radix, Coicis Semen, Polygoni Cuspidati Rhizoma et Radix, Mori Ramulus, Achyranthis Bidentatae Radix, Paeoniae Radix Rubra, Salviae Miltiorrhizae Radix et Rhizoma and Epimedii Folium.

3. The method for preparing the traditional Chinese medicine composition for treating joint diseases caused by damp-heat obstruction according to claim 2, characterized in that, the prescription medicinal materials in parts by weight are: 25 to 35 parts of Astragali Radix, 15 to 25 parts of Flemingiae Radix, 15 to 25 parts of Coicis Semen, 8 to 15 parts of Polygoni Cuspidati Rhizoma et Radix, 8 to 15 parts of Mori Ramulus, 8 to 12 parts of Achyranthis Bidentatae Radix, 8 to 12 parts of Paeoniae Radix Rubra, 5 to 10 parts of Salviae Miltiorrhizae Radix et Rhizoma, and 5 to 10 parts of Epimedii Folium.

4. The method for preparing the traditional Chinese medicine composition for treating joint diseases caused by damp-heat obstruction according to claim 3, characterized in that, the pre-extraction temperature is 70°C, the heat preservation time is 12 min, and the solid-liquid ratio is 1:

10.

5. The method for preparing the traditional Chinese medicine composition for treating joint diseases caused by damp-heat obstruction according to claim 4, characterized in that, the temperature of the high-temperature section is 90°C, the heat preservation time of the high-temperature section is 25 min, the temperature of the low-temperature section is 75°C, the heat preservation time of the low-temperature section is 20 min, and the number of cycles is 3.

6. The method for preparing the traditional Chinese medicine composition for treating joint diseases caused by damp-heat obstruction according to claim 5, characterized in that, the characteristic wavelength is 340 nm, and the sampling interval is 3 min.

7. The method for preparing the traditional Chinese medicine composition for treating joint diseases caused by damp-heat obstruction according to claim 6, characterized in that, the temperature for the reduced pressure concentration is 55°C to 60°C, and the vacuum degree is -0.08 MPa to -0.06 MPa.

8. The method for preparing the traditional Chinese medicine composition for treating joint diseases caused by damp-heat obstruction according to claim 7, characterized in that, the drying is vacuum drying, the drying temperature is 60°C to 70°C, and the vacuum degree is -0.08 MPa to -0.06 MPa.

9. The traditional Chinese medicine composition obtained by the preparation method of the traditional Chinese medicine composition for treating joint diseases caused by damp-heat obstruction according to any one of claims 1-8, characterized in that, the in vitro anti-inflammatory IC50 of the traditional Chinese medicine composition is ≤50 μg / mL.

10. The traditional Chinese medicine composition according to claim 9, characterized in that, the traditional Chinese medicine composition is applied to treating arthropathy of damp-heat obstructing bi syndrome.

11. The application of the traditional Chinese medicine composition according to claim 10, characterized in that, the traditional Chinese medicine composition is applied to anti-inflammation, analgesia and immunoregulation.

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