Pharmaceutical compositions, methods of making and using the same

Abstract

The application provides a pharmaceutical composition, a preparation method and application thereof, and the pharmaceutical composition comprises a first component and a second component, the first component is a soft capsule, and the second component is a hard capsule or a tablet; the content of the soft capsule comprises total flavonoids of pueraria lobata and epimedium, an oily base, a surfactant and a suspending agent, and the second component comprises a panax notoginseng polysaccharide extract, a filler and a lubricant. The total flavonoids of pueraria lobata and epimedium are more easily absorbed in the oily base, so that the dissolution of the total flavonoids of pueraria lobata and epimedium is effectively improved, and the problem of low bioavailability of traditional water-soluble preparations is overcome; the panax notoginseng polysaccharide extract is placed in the hard capsule or the tablet, deformation caused by contact with the oily base is avoided, and therefore the activity and bioavailability of the panax notoginseng polysaccharide extract are ensured.

Description

Technical Field

[0001] This application relates to the field of traditional Chinese medicine preparation technology, and in particular to pharmaceutical compositions, their preparation methods and applications. Background Technology

[0002] Depression is characterized by a significant and persistent depressed mood and is a major type of mood disorder. It primarily manifests as a significant and persistent low mood, characterized by depression and pessimism. Mild cases present with melancholy, lack of pleasure, and decreased interest; severe cases involve unbearable suffering, despair, a feeling that time drags on endlessly, and a sense that life is worse than death. Typical patients exhibit a diurnal variation in their depressive mood, with symptoms being worse in the morning and milder at night. Clinically, this is seen as reduced speech, significantly slower speech rate, a lower voice, difficulty answering questions, and in severe cases, impaired communication. In recent years, increasing research data indicates that with the gradual increase in social life pressures, the incidence and treatment rate of depression in my country are on the rise.

[0003] Currently, most antidepressants are chemical drugs, and there are many varieties. However, Western medicines are often highly addictive, have significant toxic side effects, and are prone to withdrawal symptoms or even a series of reactions, leading to serious damage to liver and kidney function, forming a vicious cycle. Furthermore, improper use during treatment may not only fail to treat depression but may also exacerbate the condition or lead to other mental illnesses.

[0004] Traditional Chinese medicine (TCM) preparations can effectively improve the aforementioned problems in the treatment of depression. Currently, TCM preparations used to treat depression include commercially available Shen Ge Bushen Capsules (hard capsules), whose bioavailability has room for improvement, which could enhance their therapeutic effects. Summary of the Invention

[0005] Therefore, it is necessary to provide a pharmaceutical composition with effectively improved bioavailability, its preparation method, and its application.

[0006] A first aspect of this application provides a pharmaceutical composition comprising a first component and a second component, wherein the first component is a soft capsule and the second component is a hard capsule or tablet; the contents of the soft capsule include total flavonoids from Pueraria lobata and Epimedium, an oily matrix, a surfactant, and a suspending agent, and the second component includes a polysaccharide extract of Codonopsis pilosula, a filler, and a lubricant.

[0007] In some embodiments, the oily matrix accounts for 20%-90% of the mass of the contents of the soft capsule.

[0008] In some embodiments, the surfactant in the contents of the soft capsule accounts for 0.2%-0.6% by mass.

[0009] In some embodiments, the suspending agent constitutes 0.2%-0.6% of the mass of the contents of the soft capsule.

[0010] In some embodiments, the suspending agent constitutes 0.3%-0.6% of the mass of the contents of the soft capsule.

[0011] In some embodiments, the oily matrix includes one or more of vegetable oils, medium-chain triglycerides, and polyethylene glycol.

[0012] In some embodiments, the vegetable oil includes one or more of soybean oil and peanut oil.

[0013] In some embodiments, the surfactant includes Tween 80.

[0014] In some embodiments, the suspending agent includes one or more of xanthan gum and beeswax.

[0015] In some embodiments, the capsule shell material of the soft capsule includes gelatin, glycerin, and water.

[0016] In some embodiments, the mass ratio of the gelatin to the glycerin is 1:(0.1-0.5).

[0017] In some embodiments, the mass ratio of the gelatin to the water is 1:(0.8-1.2).

[0018] In some embodiments, the total polysaccharide content in the Codonopsis pilosula polysaccharide extract is 50%-90% by mass.

[0019] In some embodiments, the second component comprises, by weight parts: 150-250 parts of Codonopsis pilosula polysaccharide extract, 40-60 parts of filler, and 1-3 parts of lubricant.

[0020] In some embodiments, the filler includes one or more of starch and microcrystalline cellulose.

[0021] In some embodiments, the lubricant includes one or more of magnesium stearate, talc, and silica.

[0022] In some embodiments, the dose of the first component is 90 mg-110 mg based on the total flavonoids of Pueraria lobata and Epimedium; and the dose of the second component is 200 mg-230 mg based on the polysaccharide extract of Codonopsis pilosula.

[0023] In some embodiments, the ratio of the number of the first component to the number of the second component is 1:2.

[0024] A second aspect of this application provides a method for preparing a pharmaceutical composition, comprising the following steps:

[0025] The contents were prepared by mixing total flavonoids from Pueraria lobata and Epimedium, an oily matrix, a surfactant, and a suspending agent.

[0026] A gel solution is prepared using the material for preparing soft capsules, and the gel solution and the contents are pressed together to prepare soft capsules, which serve as the first component.

[0027] The polysaccharide extract of Codonopsis pilosula, fillers, and lubricants are mixed to prepare hard capsules or tablets as a second component.

[0028] In some embodiments, the method for preparing the total flavonoids from Pueraria lobata and Epimedium includes:

[0029] The medicinal materials of kudzu root and epimedium were refluxed and extracted to prepare an alcohol extract;

[0030] The alcohol extract is concentrated to prepare a concentrated solution;

[0031] The concentrate was passed through a resin column;

[0032] The resin column was washed with water to remove impurities and with alcohol to elute flavonoids, and the eluent was collected.

[0033] The eluent was concentrated under reduced pressure and dried to prepare the total flavonoids from Pueraria lobata and Epimedium.

[0034] In some embodiments, the preparation method of the total extract of Codonopsis pilosula polysaccharides includes:

[0035] The medicinal material of Codonopsis pilosula was soaked and extracted to prepare an aqueous extract;

[0036] The aqueous extract was concentrated under reduced pressure to prepare a concentrated extract of Codonopsis pilosula.

[0037] Mix the concentrated extract of Codonopsis pilosula with ethanol, stir, let stand and collect the precipitate, add water to dissolve it, add ethanol again, and collect the precipitate again.

[0038] The precipitate was washed and dried to prepare the Codonopsis pilosula polysaccharide extract.

[0039] A third aspect of this application provides the use of at least one of the pharmaceutical compositions of the first aspect of this application and the pharmaceutical compositions prepared by the preparation method of the second aspect of this application in the preparation of a medicament for treating depression.

[0040] In the aforementioned pharmaceutical composition, the total flavonoids of Epimedium puerarin are more easily absorbed in an oily matrix, thereby effectively improving the solubility of the total flavonoids of Epimedium puerarin and overcoming the problem of low bioavailability in traditional water-soluble preparations. The Codonopsis pilosula polysaccharide extract is placed in hard capsules or tablets, avoiding potential deformation due to contact with the oily matrix, thus ensuring the activity and bioavailability of the Codonopsis pilosula polysaccharide extract. In summary, the bioavailability of the pharmaceutical composition of this application is significantly improved. Attached Figure Description

[0041] To more clearly illustrate the technical solutions in the embodiments of this application and to more completely understand this application and its beneficial effects, the drawings used in the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0042] Figure 1 The pharmacokinetic curves are for Example 1 and Comparative Example 1.

[0043] Figure 2 The change in body weight of mice during the experiment (n=8).

[0044] Figure 3 The experiment investigated the effects on the sucrose preference index and the number of times mice remained still (n=8).

[0045] Figure 4 The experiment aimed to investigate the effects on the cumulative immobility time and sleep duration in mice (n=8).

[0046] Figure 5 The experiment aimed to investigate the effect of open field test on the movement behavior of mice (n=8).

[0047] Figure 6 The experiment aimed to investigate the effects on relevant neurotransmitters in mouse brain tissue (n=8).

[0048] Figure 7 H&E pathological sections of mouse hippocampus (n=8) with a scale bar of 100 µm.

[0049] Figure 8 Nissl stained sections of mouse hippocampus (n=8) Scale bar 100 µm. Detailed Implementation

[0050] To facilitate understanding of this application, a more complete description of the application will be provided below with reference to relevant embodiments. Preferred embodiments of the application are given below. However, the application can be implemented in many different forms and is not limited to the embodiments described herein. Rather, these embodiments are provided to provide a thorough and complete understanding of the disclosure of this application.

[0051] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the application.

[0052] As used herein, the terms "and / or," "or / and," and "and / or" encompass any one of two or more of the related listed items, as well as any and all combinations of the related listed items. These arbitrary and all combinations include any two related listed items, any more related listed items, or a combination of all related listed items. It should be noted that when at least three items are connected using at least two conjunctions selected from "and / or," "or / and," and "and / or," it should be understood that, in this application, the technical solution undoubtedly includes solutions connected by "logical AND," and also undoubtedly includes solutions connected by "logical OR."

[0053] In this application, the technical features described in an open-ended manner include both closed technical solutions consisting of the listed features and open technical solutions that include the listed features.

[0054] In this application, numerical ranges are referred to as continuous unless otherwise specified, and include the minimum and maximum values ​​of the range, as well as every value between the minimum and maximum values. Furthermore, when the range refers to integers, it includes every integer between the minimum and maximum values ​​of the range. Additionally, when multiple ranges are provided to describe a feature or characteristic, the ranges may be merged. In other words, unless otherwise specified, all ranges disclosed herein should be understood to include any and all subranges to which they are incorporated.

[0055] This document only specifically discloses some numerical ranges. However, any lower limit can be combined with any upper limit to form an unspecified range; and any lower limit can be combined with other lower limits to form an unspecified range, just as any upper limit can be combined with any other upper limit to form an unspecified range. Furthermore, each individually disclosed point or single value can itself serve as a lower or upper limit and be combined with any other point or single value or with other lower or upper limits to form an unspecified range.

[0056] Unless otherwise specified, the temperature parameters in this application may be either constant temperature processing or processing within a certain temperature range. The constant temperature processing allows temperature fluctuations within the precision range controlled by the instrument, such as ±5°C, ±4°C, ±3°C, ±2°C, or ±1°C.

[0057] In this document, the term "suitable" as used in phrases such as "suitable combination," "suitable method," and "any suitable method" refers to the ability to implement the technical solution of this application, solve the technical problem of this application, and achieve the expected technical effect of this application.

[0058] In this application, terms such as "further," "even more," and "particularly" are used for descriptive purposes and to indicate differences in content, but should not be construed as limiting the scope of protection of this application.

[0059] In this application, "optionally," "optionally," and "optional" mean that something is optional, that is, it means that it is selected from either "with" or "without." If there are multiple "optional" entries in a technical solution, unless otherwise specified, and there are no contradictions or mutual constraints, each "optional" entry shall be independent.

[0060] In the description of the application, "multiple" means at least two, such as two, three, etc., unless otherwise explicitly specified.

[0061] Unless otherwise specified, all embodiments and optional embodiments of this application can be combined to form new technical solutions. Unless otherwise specified, all technical features and optional technical features of this application can be combined to form new technical solutions.

[0062] Unless otherwise specified, all steps in this application may be performed sequentially or randomly, but sequentially is preferred.

[0063] Commercially available Shen Ge Bu Shen Capsules (hard capsules) contain total flavonoids from Pueraria lobata and Epimedium, and polysaccharide extract from Codonopsis pilosula. The total flavonoids from Pueraria lobata and Epimedium are obtained through extraction from Pueraria lobata and Epimedium. Puerarin and total flavonoids from Epimedium are the main active ingredients. Puerarin is almost insoluble in water and slightly soluble in ethanol, resulting in low dissolution and poor bioavailability in commercially available Shen Ge Bu Shen Capsules. The main components of total Epimedium flavonoids (such as icariin A, icariin B, icariin C, and icariin glycosides) have poor water solubility and an unsatisfactory oil-water partition coefficient, leading to poor absorption. Codonopsis pilosula polysaccharides have high bioavailability but poor lipid solubility and are insoluble in oil. If formulated with flavonoid components, this may affect the manufacturing process and formulation stability, leading to drug separation or clogging of equipment pipelines. Therefore, commercially available Ginseng and Kudzu Kidney-Nourishing Capsules (hard capsules) have problems such as low bioavailability of some ingredients and short shelf life, which urgently need to be addressed.

[0064] In view of the above problems, one or more embodiments of this application provide a pharmaceutical composition comprising a first component and a second component, wherein the first component is a soft capsule and the second component is a hard capsule or tablet; the contents of the soft capsule include total flavonoids from Pueraria lobata and Epimedium, an oily matrix, a surfactant and a suspending agent, and the second component includes a polysaccharide extract of Codonopsis pilosula, a filler and a lubricant.

[0065] It should be noted that the terms "first component," "second component," etc., mentioned in the context are for descriptive purposes only and should not be construed as indicating or implying relative importance or quantity, nor should they be interpreted as implicitly specifying the importance or quantity of the indicated technical features. Moreover, "first," "second," etc., serve only as a non-exhaustive enumeration and should be understood as not constituting a closed limitation on quantity.

[0066] Understandably, in the pharmaceutical composition of this application, the total flavonoids of Epimedium puerarin are more easily absorbed in an oily matrix, thereby effectively improving the solubility of the total flavonoids of Epimedium puerarin and overcoming the problem of low bioavailability in traditional water-soluble preparations; the polysaccharide extract of Codonopsis pilosula is placed in hard capsules or tablets, avoiding deformation that may occur due to contact with the oily matrix, thus ensuring the activity and bioavailability of the Codonopsis pilosula polysaccharide extract. In summary, the bioavailability of the pharmaceutical composition of this application is significantly improved.

[0067] In some embodiments, the oily matrix in the contents of the soft capsules comprises 20%-90% by mass; for example, it can be, but is not limited to, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or any range between two of the above values. When the amount of oily matrix is ​​within the above range, it can disperse and dissolve the total flavonoids of Epimedium puerariae.

[0068] As one possible implementation, the surfactant content in the soft capsules comprises 0.2%-0.6% by mass; for example, it can be, but is not limited to, 0.2%, 0.25%, 0.3%, 0.35%, 0.4%, 0.45%, 0.5%, 0.55%, 0.6%, or any range between two of the above mass percentages. When the amount of surfactant is within the above range, the suspension is stable and there is no obvious stratification.

[0069] In some embodiments, the mass percentage of the suspending agent in the contents of the soft capsule is 0.2%-0.6%; for example, it can be, but is not limited to, 0.2%, 0.25%, 0.3%, 0.35%, 0.4%, 0.45%, 0.5%, 0.55%, 0.6%, or any range between two of the above mass percentages. When the amount of suspending agent is within the above range, it is beneficial to form a homogeneous suspension with suitable viscosity. Optionally, the mass percentage of the suspending agent in the contents of the soft capsule is 0.3%-0.6%.

[0070] In some embodiments, the oily matrix includes one or more of vegetable oils, medium-chain triglycerides, and polyethylene glycol; thereby facilitating the dissolution and absorption of total flavonoids from Pueraria lobata.

[0071] In some alternative implementations, the vegetable oil includes one or more of soybean oil and peanut oil.

[0072] As one possible implementation, the surfactant includes Tween 80.

[0073] In some embodiments, the suspending agent includes one or more of xanthan gum and beeswax.

[0074] In some embodiments, the capsule shell material of the soft capsule includes gelatin, glycerin, and water.

[0075] As one possible implementation, the mass ratio of gelatin to glycerin is 1:(0.1-0.5); for example, it can be, but is not limited to, 1:0.1, 1:0.2, 1:0.3, 1:0.4, 1:0.5 or any range between the above two mass ratios.

[0076] In some embodiments, the mass ratio of gelatin to water is 1:(0.8-1.2); for example, it can be, but is not limited to, 1:0.8, 1:0.9, 1:1.0, 1:1.1, 1:1.2 or any range between two of the above mass ratios.

[0077] As one possible implementation, the total polysaccharide content in the Codonopsis pilosula polysaccharide extract is 50%-90% by mass; for example, it can be, but is not limited to, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90% or any range between the above two mass percentages.

[0078] In some embodiments, the second component comprises, by weight parts: 150-250 parts of Codonopsis pilosula polysaccharide extract, 40-50 parts of filler, and 1-2 parts of lubricant.

[0079] As an example, the mass fractions of the Codonopsis pilosula polysaccharide extract contained in the second component may be, but are not limited to, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, 250 parts or any range between two of the above mass fractions.

[0080] The filler contained in the second component may be, but is not limited to, 40, 42, 44, 46, 48, 50, 52, 54, 56, 58, 60 parts by mass, or any range between any two of the above.

[0081] The mass fraction of the lubricant contained in the second component can be, but is not limited to, 1 part, 2 parts, 3 parts, or any range between any two of the above mass fractions.

[0082] It should be noted that when the amounts of Codonopsis pilosula polysaccharide extract, filler, and lubricant are within the above-mentioned ranges, it is beneficial to improve the flowability of Codonopsis pilosula polysaccharide extract during the preparation process and the stability of Codonopsis pilosula polysaccharide extract in the obtained product.

[0083] In some embodiments, the filler includes one or more of starch and microcrystalline cellulose.

[0084] As one possible implementation, the lubricant includes one or more of magnesium stearate, talc, and silica.

[0085] In some embodiments, the dosage of the first component, calculated based on the total flavonoids of Epimedium puerarin, is 90 mg to 110 mg; for example, it can be, but is not limited to, 90 mg, 100 mg, 110 mg, or any range between the two of the above dosages.

[0086] As one possible implementation, the dosage of the second component, based on the Codonopsis pilosula polysaccharide extract, is 200mg-230mg; for example, it can be, but is not limited to, 200mg, 210mg, 220mg, 230mg or any range between the two of the above dosages.

[0087] In some implementations, the ratio of the first component to the second component is approximately 1:2; thus, it has excellent therapeutic effects on depression.

[0088] One or more embodiments of this application provide a method for preparing a pharmaceutical composition, which can be used to prepare the above-mentioned pharmaceutical composition. The preparation method includes the following steps: mixing total flavonoids from Pueraria lobata and Epimedium, an oily matrix, a surfactant, and a suspending agent to prepare a content; preparing a gel using materials for preparing soft capsules, and pressing the gel and the content together to form soft capsules, as the first component; mixing Codonopsis pilosula polysaccharide extract, a filler, and a lubricant to prepare hard capsules or tablets, as the second component.

[0089] It should be noted that the pharmaceutical compositions prepared by the preparation method of this application have the same or similar technical solutions and beneficial effects as the pharmaceutical compositions described above, and will not be repeated here.

[0090] In some embodiments, the method for preparing total flavonoids from kudzu and epimedium includes: reflux extraction of kudzu root and epimedium to prepare an alcohol extract; concentration of the alcohol extract to prepare a concentrate; passing the concentrate through a resin column; washing the resin column with water to remove impurities and washing with alcohol to elute flavonoids, collecting the eluent; concentrating the eluent under reduced pressure and drying it to prepare total flavonoids from kudzu and epimedium.

[0091] As an example, the preparation method of total flavonoids from Pueraria lobata and Epimedium includes: placing Pueraria lobata slices and Epimedium slices in an extraction tank, refluxing at 80℃-90℃ for three times, adding 10-14 times the weight of the slices of 80% ethanol aqueous solution each time, for 1-2 hours each time, and filtering the extract through a 40-mesh sieve. Combine the ethanol extracts of Pueraria lobata and Epimedium after the three extractions, concentrate at 60℃-70℃, and filter for later use. Let the concentrated Pueraria lobata and Epimedium extract stand at 10℃ for 40 hours, take the supernatant and pass it through an AB-8 macroporous adsorption resin column at a flow rate of approximately 0.5 BV / h-1.5 BV / h, first washing away impurities with 4 times the weight of the resin of water at a flow rate of 1.5-3 BV / h, and discarding the water. The flavonoids were then desorbed using an 80% ethanol aqueous solution (8 times the resin weight). The eluent was collected, and the vacuum was controlled at -0.05 to -0.09 MPa and the temperature at 60 to 80°C. The ethanol eluent was concentrated under reduced pressure to a relative density of approximately 1.08 to 1.15 (65 to 75°C), and then placed in a vacuum drying oven at a vacuum of -0.06 to -0.09 MPa and a temperature below 60°C for approximately 26 hours. The dried extract was pulverized and passed through a 100-mesh sieve to obtain total flavonoids from Epimedium puerariae.

[0092] In some embodiments, a gelatin solution is prepared using materials for preparing soft capsules. The gelatin solution and contents are then combined for granulation. The steps for preparing soft capsules include: weighing glycerin and water and adding them to a gelatin mixing tank, stirring until completely mixed. The temperature of the gelatin mixing tank is set to 70℃-80℃. After the tank reaches the set temperature, gelatin is poured in, and stirring is started until no visible particles are visible on the surface of the gelatin solution. A vacuum pump is turned on, and the vacuum degree is controlled at -0.04 MPa to -0.08 MPa until the gelatin solution becomes clear. After vacuuming, the solution is filtered through a 100-mesh filter and stored in an insulated storage tank. The temperature of the insulated storage tank is set to 45℃-60℃ until no visible bubbles are visible inside the gelatin solution. A chiller is turned on to lower the temperature by 14℃ to 24℃ and the temperature of the capsule box is raised by 35℃ to 55℃. When the temperature reaches the set value, the gelatin is fed in and granulated. The thickness of the capsule shell is 0.60mm to 0.80mm, and the content is controlled at 0.4g / capsule to 0.5g / capsule.

[0093] As one possible implementation, a gelatinous solution is prepared using the material for preparing soft capsules. The gelatinous solution and contents are then compressed into capsules. The steps for preparing soft capsules also include: placing the sample in a dryer with an oil-absorbing cloth and shaping it until the capsules are no longer sticky; removing defective capsules such as irregularly shaped capsules, deflated capsules, air-filled capsules, leaking capsules, large capsules, and small capsules (compared to the normal filling weight); and placing the capsule drying rack in a drying room at a temperature of 18℃~26℃, a humidity of 35%RH~65%RH, and a moisture content controlled at 6%~10%.

[0094] In some embodiments, the method for preparing the total extract of Codonopsis pilosula polysaccharides includes: soaking and extracting Codonopsis pilosula medicinal material to prepare an aqueous extract; concentrating the aqueous extract under reduced pressure to prepare a Codonopsis pilosula concentrate; mixing the Codonopsis pilosula concentrate with ethanol, stirring, allowing it to stand to collect the precipitate, adding water to dissolve it, adding ethanol again, and collecting the precipitate again; washing and drying the precipitate to prepare the Codonopsis pilosula polysaccharide extract.

[0095] As an example, the preparation method of Codonopsis pilosula polysaccharide extract includes: taking Codonopsis pilosula slices, adding 8-10 times the total mass of water to an extraction tank, and extracting three times at 90℃-100℃ for 2-4 hours each time. The extract is then filtered through a 40-mesh sieve. The Codonopsis pilosula aqueous extracts after the three extractions are combined and concentrated under reduced pressure at a temperature of 60℃-70℃. The concentrated extract is then set aside. The Codonopsis pilosula concentrate is placed in an alcohol precipitation tank, and the stirring device is turned on. Ethanol is added while stirring to achieve an alcohol content of 75%-85%. The mixture is stirred evenly, allowed to stand, and the precipitate is collected. Water is added to dissolve the precipitate, and ethanol is added again. The precipitate is collected again, and an appropriate amount of ethanol is added to wash the precipitate. The supernatant is discarded, and the precipitate is placed in a vacuum drying oven at a vacuum of -0.06~-0.09 MPa and a temperature below 50℃ for approximately 26 hours. The dry extract is pulverized and passed through a 60-mesh sieve to obtain the Codonopsis pilosula polysaccharide extract.

[0096] In some embodiments, the extract of Codonopsis pilosula polysaccharide, filler, and lubricant are mixed and then tablets are prepared by direct compression of the powder. Optionally, the tablet hardness is 20N-40N.

[0097] One or more embodiments of this application provide the use of at least one of the above-described pharmaceutical compositions and pharmaceutical compositions prepared by the above-described preparation methods in the preparation of a medicament for treating depression.

[0098] The technical solutions of this application will be described in detail below with reference to specific embodiments. It should be understood that these embodiments are only for illustrating this application and are not intended to limit the scope of this application. For experimental methods in the following embodiments where specific conditions are not specified, please refer to the guidelines given in this application first, or follow experimental manuals or conventional conditions in the field, or follow the conditions recommended by the manufacturer, or refer to experimental methods known in the field.

[0099] In the specific embodiments described below, the measurement parameters involving raw material components may have slight deviations within the weighing accuracy range unless otherwise specified. Temperature and time parameters are subject to acceptable deviations due to instrument testing accuracy or operational precision.

[0100] It should be noted that the preparation method of total flavonoids from Pueraria lobata and Epimedium used in the following examples and comparative examples is as follows: Pueraria lobata slices and Epimedium slices are placed in an extraction tank and extracted three times by reflux at 90°C. Each time, 12 times the weight of the slices of 80% ethanol aqueous solution is added, and the extraction time is 1 hour each time. The extract is filtered through a 40-mesh sieve. The ethanol extracts of Pueraria lobata and Epimedium after the three extractions are combined, concentrated at 60°C, and filtered for later use. The concentrated Pueraria lobata and Epimedium extract is allowed to stand at 10°C for 40 hours. The supernatant is then passed through an AB-8 macroporous adsorption resin column at a flow rate of approximately 1 BV / h. Impurities are first washed away with 4 times the weight of the resin of water at a flow rate of 2 BV / h, and the water is discarded. The flavonoids were then desorbed using an 80% ethanol aqueous solution (8 times the resin weight). The eluent was collected, and the vacuum was controlled at -0.07 MPa and the temperature at 70°C. The ethanol eluent was concentrated under reduced pressure to a relative density of approximately 1.10 (70°C), and then placed in a vacuum drying oven at a vacuum of -0.08 MPa and a temperature below 60°C for approximately 26 hours. The dried extract was pulverized and passed through a 100-mesh sieve to obtain total flavonoids from Epimedium puerariae.

[0101] The preparation method of the Codonopsis pilosula polysaccharide extract used in the following examples and comparative examples includes: taking Codonopsis pilosula slices, adding 8 times the total mass of water to an extraction tank, and extracting three times at 90°C for 3 hours each time. The extract is then filtered through a 40-mesh sieve. The Codonopsis pilosula aqueous extracts after the three extractions are combined and concentrated under reduced pressure at 65°C for later use. The concentrated Codonopsis pilosula extract is placed in an alcohol precipitation tank, and the stirring device is turned on. Ethanol is added while stirring to make the alcohol content reach 80%. The mixture is stirred evenly, allowed to stand, and the precipitate is collected. Water is added to dissolve the precipitate, and then ethanol is added again. The precipitate is collected again, and an appropriate amount of ethanol is added to wash the precipitate. The supernatant is discarded, and the precipitate is placed in a vacuum drying oven. The vacuum degree is set to -0.08 MPa, and the temperature is below 50°C for drying for about 26 hours. The dry extract is pulverized and passed through a 60-mesh sieve to obtain the Codonopsis pilosula polysaccharide extract.

[0102] I. Preparation of Pharmaceutical Compositions

[0103] Example 1

[0104] 1. Preparation of soft capsules

[0105] Step S1: Take 200g of total flavonoid powder from Pueraria lobata and Epimedium, add it together with 4g of Tween 80 and 4g of xanthan gum into 800g of soybean oil, homogenize it and use it as a filler.

[0106] Step S2: Weigh glycerin and water and add them to the gelatinizing tank, stirring until completely mixed. Set the temperature of the gelatinizing tank to 75℃. Once the tank reaches the set temperature, pour in the gelatin and start stirring until no visible particles are visible on the surface of the solution. Turn on the vacuum pump and control the vacuum at -0.06 MPa until the solution becomes clear. After vacuuming, filter through a 100-mesh filter into a heat-insulated storage tank. Set the temperature of the heat-insulated storage tank to 50℃ until no visible bubbles are visible inside the solution. Turn on the chiller to lower the temperature by 14℃ and raise the temperature of the gelatin box by 35℃. Once the temperature reaches the set value, begin feeding the gelatin and performing pelleting. The thickness of the gelatin shell is 0.70 mm, and the content is controlled so that each pellet contains 90 mg of total flavonoids from Pueraria lobata and Epimedium. Place the sample in a dryer with an oil-absorbing cloth and shape it until the capsules are no longer sticky; pick out the defective capsules such as irregularly shaped capsules, shriveled capsules, bubble capsules, leaking capsules, large capsules, and small capsules (compared with the normal filling amount); place the capsule drying rack in the capsule drying room at a temperature of 25℃, humidity of 50%RH, and moisture content controlled at 8%.

[0107] 2. Tablet preparation

[0108] Step S3: Take 200g of Codonopsis pilosula polysaccharide powder, mix it with 50g of microcrystalline cellulose and 2g of magnesium stearate, and directly compress the powder into tablets using a tableting machine to make tablets containing 210mg of Codonopsis pilosula polysaccharide extract per tablet. The tablet hardness is 30N.

[0109] 3. Preparation of pharmaceutical compositions

[0110] Step S4: Fill one soft capsule and one tablet into a 00# capsule shell to obtain the Ginseng and Kudzu Kidney-Nourishing Soft Capsule Composition.

[0111] Example 2

[0112] The preparation method of Example 2 is similar to that of Example 1, except that the amount of xanthan gum used in step S1 of the preparation of soft capsules in Example 2 is 2g; all other aspects are the same.

[0113] Example 3

[0114] The preparation method of Example 3 is similar to that of Example 1, except that the amount of xanthan gum used in step S1 of the preparation of soft capsules in Example 3 is 3g; all other aspects are the same.

[0115] Example 4

[0116] The preparation method of Example 4 is similar to that of Example 1, except that the amount of xanthan gum used in step S1 of the preparation of soft capsules in Example 4 is 5g; all other aspects are the same.

[0117] Example 5

[0118] The preparation method of Example 5 is similar to that of Example 1, except that the amount of xanthan gum used in step S1 of the preparation of soft capsules in Example 5 is 6g; all other aspects are the same.

[0119] Example 6

[0120] The preparation method of Example 6 is similar to that of Example 1, except that beeswax is used instead of xanthan gum in step S1 when preparing soft capsules in Example 6; all other steps are the same.

[0121] The soft capsules in Examples 1-6 were tested for their suspension-enhancing effect, and the properties of the suspension were observed. The test results are shown in Table 1.

[0122] Table 1

[0123]

[0124] As can be seen from the comparison of the results of Examples 1-6 in Table 1, using xanthan gum and beeswax as suspending agents, and controlling the mass ratio of the suspending agents in the soft capsule contents to be 0.2%-0.6%, has a better suspending effect, which is beneficial to improving the stability of the soft capsule.

[0125] Example 7

[0126] The preparation method of Example 7 is similar to that of Example 1, except that the amount of Tween 80 used in step S1 of the preparation of soft capsules in Example 7 is 2g; all other aspects are the same.

[0127] Example 8

[0128] The preparation method of Example 8 is similar to that of Example 1, except that the amount of Tween 80 used in step S1 of the preparation of soft capsules in Example 8 is 3g; all other aspects are the same.

[0129] Example 9

[0130] The preparation method of Example 9 is similar to that of Example 1, except that the amount of Tween 80 used in step S1 of the preparation of soft capsules in Example 9 is 6g; all other aspects are the same.

[0131] The soft capsules from Examples 1 and 7-9 were subjected to suspension-enhancing effect tests, and the properties of the suspension were observed. The test results are shown in Table 2.

[0132] Table 2

[0133]

[0134] As can be seen from the comparison of the results of Examples 1 and 7-9 in Table 2, when the mass percentage of surfactant Tween 80 in the soft capsule contents is 0.2%-0.6%, it is beneficial to the stability of the suspension.

[0135] Example 10

[0136] The preparation method of Example 10 is similar to that of Example 1, except that the amount of microcrystalline cellulose used in step S3 of the tablet preparation in Example 10 is 40g; all other aspects are the same.

[0137] Example 11

[0138] The preparation method of Example 11 is similar to that of Example 1, except that the amount of microcrystalline cellulose used in step S3 of the tablet preparation in Example 11 is 60g; all other aspects are the same.

[0139] Example 12

[0140] The preparation method of Example 12 is similar to that of Example 1, except that starch is used instead of microcrystalline cellulose in step S3 when preparing tablets in Example 12; all other steps are the same.

[0141] Example 13

[0142] The preparation method of Example 13 is similar to that of Example 1, except that the amount of magnesium stearate used in step S3 of the tablet preparation in Example 13 is 1g; all other aspects are the same.

[0143] Example 14

[0144] The preparation method of Example 14 is similar to that of Example 1, except that the amount of magnesium stearate used in step S3 of the tablet preparation in Example 14 is 3g; all other aspects are the same.

[0145] The tablets from Examples 1 and 10-14 were subjected to disintegration time test, friability test, and hardness test, respectively, using the following methods:

[0146] Disintegration time limit: According to the Disintegration Time Limit Test Method of Part IV of the Pharmacopoeia of the People's Republic of China, 0921.

[0147] Friability: According to the 0923 Tablet Friability Test Method in Part IV of the Pharmacopoeia of the People's Republic of China.

[0148] Tablet hardness: Take 10 tablets of this product and measure them using a hardness tester.

[0149] The test results of Examples 1 and 10-14 are shown in Table 3.

[0150] Table 3

[0151]

[0152] As can be seen from the comparison of the results of Examples 1 and 10-12 in Table 3, using at least one of starch and microcrystalline cellulose as filler, and controlling the amount of filler within a specific range, that is, controlling the mass ratio of Codonopsis pilosula polysaccharide extract to filler to be (150-250):(40-60), the prepared tablets have suitable brittleness, hardness and disintegration properties.

[0153] As can be seen from the comparison of the results of Examples 1 and 13-14 in Table 3, when the amount of lubricant is controlled within a specific range, that is, when the mass ratio of Codonopsis pilosula polysaccharide extract to lubricant is controlled at (150-250):(1-3), the resulting tablets have suitable brittleness, hardness and disintegration properties.

[0154] Comparative Example 1

[0155] Take 210g of Codonopsis pilosula polysaccharide extract, 90g of total flavonoids from Pueraria lobata and Epimedium, and 1g of silicon dioxide. Pass the silicon dioxide through a 60-mesh sieve and place it in a two-dimensional motion mixer to obtain a traditional Chinese medicine composition. Fill the traditional Chinese medicine mixture into hard capsule shells to obtain Ginseng and Pueraria lobata Kidney-Tonifying Hard Capsules.

[0156] II. Pharmacokinetic and Pharmacodynamic Studies

[0157] 1. Pharmacokinetic studies

[0158] 1.1 Experimental Reagents and Materials

[0159] Acetonitrile (chromatographic grade, Fisher Scientific); methanol and ethanol (analytical grade, Tianjin Zhiyuan Chemical Reagent Co., Ltd.); puerarin (batch number: 110752-201313) and icariin (batch number: 110737-202017) were purchased from the National Institutes for Food and Drug Control; Codonopsis pilosula (batch number: 230901), Pueraria lobata (batch number: 250701), and Epimedium (batch number: 231201) were purchased from Hebei Shuangning Pharmaceutical Co., Ltd.

[0160] 1.2 Instruments and Equipment

[0161] LT1000B electronic balance, Changshu Tianliang Instrument Co., Ltd.; BS323S electronic balance, Beijing Sartorius Scientific Instruments Co., Ltd.; JXFSTPRP-CL fully automatic sample cryogenic grinder, Shanghai Jingxin Industrial Development Co., Ltd.; H1650R refrigerated high-speed centrifuge, Shanghai Luxiangyi Centrifuge Instrument Co., Ltd.; Leica; SC082132 ultrapure water system, Sichuan Wote Experimental Instruments and Equipment. Acquity UPLC® HSS T3 (100 × 2.1 mm, 1.8 μm), ACQUITY ultra-high performance liquid chromatography-Xevo G2-XS high resolution time-of-flight mass spectrometer (Waters), MS205DU analytical balance (Mettler-Toledo Instruments (Shanghai) Co., Ltd.).

[0162] 1.3 Chromatographic conditions

[0163] Chromatographic column: Acquity UPLC® HSS T3 (100 × 2.1 mm, 1.8 μm); column temperature: 45 ℃; flow rate: 0.3 mL / min; injection volume: 1 μL; mobile phase: A is 0.1% formic acid aqueous solution, B is acetonitrile; gradient elution: 0~7 min, 90~80% A; 7~14 min, 80~65% A; 14~21 min, 65~50% A; 21~28 min, 50~35% A; 28~35 min, 35~10% A; 35~37 min, 10% A; 37~37.1 min, 10~90% A; 37.1~40 min, 90% A.

[0164] 1.4 Mass Spectrometry Conditions

[0165] Ion source temperature: 100 ℃; desolvation gas temperature: 550 ℃; desolvation gas flow rate: 600 L / h; cone gas flow rate: 50 L / h; capillary voltage: 3 kV under positive ion conditions, 2 kV under negative ion conditions; cone voltage: 40 eV; operating mode: MSE mode; scanning mode: sensitivity mode; scanning range: m / z 50~1200; scanning time: 0.5 s; collision energy: low collision energy is off, high collision energy is 20~40 eV.

[0166] 1.5 Laboratory Animals

[0167] ICR mice, male, provided by Beijing Huafukang Biotechnology Co., Ltd., production license number: NO.110322251100607823. The animals were kept in the animal center of Xinjiang Uygur Autonomous Region Institute of Materia Medica, and the use license number: SYXJ (Xin) 2022 - 0002. Humidity was 30 - 50%, and the temperature was controlled at 20 - 26 °C. There was a 12h:12h light - dark cycle; the cage environment was kept clean and dry; the animals were fed with food and water daily; the animals were allowed to move freely. This experiment has been approved by the Animal Ethics Review Committee of Xinjiang Uygur Autonomous Region Institute of Materia Medica, and the ethical approval number: XJIMM - 20250103.

[0168] 1.6 Grouping and drug administration

[0169] After 7 days of adaptive feeding, the mice were randomly divided into 17 groups, with 5 mice in each group. Each group represented a sampling time point, including a blank control group (administered at 0 h) and sampling groups of Shenge Bushen Soft Capsule Composition of Example 1 and Shenge Bushen Hard Capsule of Comparative Example 1 at 0.5, 1, 2, 4, 6, 8, 12, and 24 h after administration. The drug was administered by gavage at a dose of 800 mg / kg and a volume of 20 ml / kg.

[0170] 1.7 Biological sample processing

[0171] For mouse plasma (300 μL / mouse), add 1.2 mL of methanol, vortex for 30 s, then sonicate (800 W, 50 Hz) for 30 min, centrifuge at 12000 r / min (4 °C) for 10 min, take the supernatant and dry it in a metal bath at 40 °C to obtain plasma samples for standby. Add 60 μL of 50% methanol solution to the plasma sample, vortex for 3 min, then sonicate at 800 W, 50 Hz for 10 min, take the supernatant into a centrifuge tube, place the centrifuge tube in a centrifuge, centrifuge at 13000 r / min (4 °C) for 10 min, and quantitatively absorb the supernatant (add 45 μL of methanol to each plasma sample) and put it into a clean sample bottle for detection.

[0172] 1.8 Experimental results of pharmacokinetic parameters

[0173] Blood was collected from the orbital venous plexus of mice at different times after administration (anticoagulated with 1% heparin), centrifuged at 12000 r / min for 10 min to separate plasma, and stored frozen for standby. The pharmacokinetic parameters were fitted using 3P87 software (Chinese Pharmacological Society). The results were obtained by Figure 1As shown in Table 4, after oral administration, the plasma concentrations of puerarin in mice peaked at 0.95 h, while those of icariin peaked at 2.25 h. Subsequently, the plasma concentrations of both components decreased. The plasma concentration of puerarin reached its lowest value at 24 h after administration; the plasma concentration of icariin reached its lowest value at 6 h after administration, then rebounded and increased until 12 h after administration, before reaching its lowest value at 24 h after administration.

[0174] Drug metabolism parameters showed that, comparing the AUC0-24 values ​​of puerarin, the soft capsule composition had a value of 512.5 ng / ml, while the hard capsule composition had a value of 432.3 ng / ml, representing an increase of nearly 1.18 times; comparing the AUC0-24 values ​​of icariin, the soft capsule composition had a value of 1.83 ng / ml, while the hard capsule composition had a value of 1.52 ng / ml, representing an increase of nearly 1.20 times.

[0175] Table 4 Pharmacokinetic curves (n=5)

[0176]

[0177] Depend on Figure 1 As shown in Table 4, in mouse models, the bioavailability of puerarin and icariin in the drug combination formulation of this application is approximately 1.2 times higher than that of the original dosage form (Comparative Example 1 hard capsule).

[0178] 2. Pharmacodynamic studies

[0179] 2.1 Test substance

[0180] Example 1: The composition of Shenge Bushen soft capsule and Comparative Example 1: Shenge Bushen hard capsule.

[0181] Venlafaxine Hydrochloride Extended-Release Capsules, Specification: 75mg (calculated as venlafaxine), Dosage and Administration: Oral administration, 1 capsule once a day; Indications: For the treatment of depression (including depression with anxiety) and generalized anxiety disorder, Batch No.: 8177036, Pfizer Pharmaceuticals Co., Ltd.

[0182] 2.2 Reagents

[0183] Sodium pentobarbital, batch number: P3761, Sigma, USA; 4-Chloro-DL-phenylalanine (PCPA), batch number: A12IS208585, Yuanye Bio-Technology Co., Ltd., Shanghai; 4% Paraformaldehyde Fixative, batch number: DF0135, Biosharp; Endogenous Peroxidase Scavenger, batch number: SI25-01, Seven Innovation Reagent Co., Ltd.; Goat Serum, batch number: SL038, Solarbio Science & Technology Co., Ltd., Beijing; Hematoxylin Staining, batch number: BL702B, Biosharp; 5-Hydroxytryptamine (5-HT) ELISA Kit, batch numbers: WU06FH667168, WU0766689981, Gamma-Aminobutyric Acid (GABA) Colorimetric Test Kit, batch number: WU114TPP8459, Glutamic Acid (GLU) Colorimetric Test Kit, batch numbers: WU142J663774, WU13PN840804, all purchased from Wuhan Elabscience Biotechnology Co., Ltd.

[0184] 2.3 Instrument and Equipment

[0185] LT1000B Electronic Balance, Changshu Tianliang Instrument Co., Ltd.; BS323S Electronic Balance, Sartorius Scientific Instruments Co., Ltd., Beijing; Open Field Test Analysis System for Mice and Rats, Xinruan Information Technology Co., Ltd., Shanghai; Animal Behavior Video Analysis System, ZS Type Tail Suspension Hardware, Zhongshi Ditech Science & Technology Development Co., Ltd., Beijing; JXFSTPRP-CL Automatic Sample Freezing Grinder, Jingxin Industrial Development Co., Ltd., Shanghai; H1650R Refrigerated High-Speed Centrifuge, Shanghai L湘仪 Centrifuge Instrument Co., Ltd.; Cytation3 Multifunctional Microplate Reader, BioTek Instruments, Inc., USA; LEICA RM2245 Microtome, Leica; DM4000 Optical Microscope, Leica; SC082132 Ultra-Pure Water Instrument, Sichuan Wote Experimental Instrument Equipment.

[0186] 2.4 Experimental Animals

[0187] ICR mice, male, provided by Huafukang Bio-Technology Co., Ltd., Beijing, production license number: NO.110322251100607823, the animals were housed in the Animal Center of Xinjiang Uygur Autonomous Region Institute of Materia Medica, use license number: SYXJ (Xin) 2022-0002; humidity 30-50%, temperature control 20-26 °C. 12h:12h light-dark alternating illumination; keep the cage environment clean and dry; add feed and water to the animals daily; keep the animals free to move. This experiment has been approved by the Animal Ethics Review Committee of Xinjiang Uygur Autonomous Region Institute of Materia Medica, ethical approval number: XJIMM-20250103.

[0188] 2.5 Experimental Grouping and Drug Administration

[0189] (1) Establishment of the depression model (CUMS): ① Fasting for 24 h; ② Water restriction for 24 h; ③ Ice water bath at 4℃ for 2 min; ④ Inclined cage at 45° for 24 h; ⑤ Noise for 4 h; ⑥ Moist bedding for 24 h; ⑦ Restraint; ⑧ Running for 5 min. Two types of stimuli were used each day, and the same type of stimulus could not be used consecutively.

[0190] (2) Drug grouping: chemical positive drug group (venlafaxine hydrochloride), soft capsule group (ginseng and kudzu kidney-tonifying soft capsule composition), hard capsule group (ginseng and kudzu kidney-tonifying hard capsule).

[0191] (3) Grouping and administration: Forty male ICR mice, weighing 18-20g, were used in this study. They were housed in a specific pathogen-free (SPF) experimental animal center at a room temperature of (25±2)℃, relative humidity of (40%-50%), and a light / dark cycle of (12h / 12h). After 7 days of acclimatization, the mice were randomly divided into 5 groups (n=8): control group, CUMS group, positive control group, soft capsule group, and hard capsule group. Mice in the CUMS group, positive control group, soft capsule group, and hard capsule group underwent CUMS stimulation for 7 consecutive days, while the control group was fed normally. On the 8th day of the experiment, mice in the positive control group (venlafaxine hydrochloride, 11mg / kg), soft capsule group (400mg / kg), and hard capsule group (400mg / kg) were administered the drug by gavage. At the same time, the control group and CUMS group were given the same volume of distilled water. The administration was continued by gavage until the end of the experiment. During the entire administration period, except for the control group, all other groups continued to undergo CUMS stimulation. The mice were treated after the open field test on the 21st day.

[0192] 2.5 Behavioral testing of mice

[0193] (1) Sucrose Preference Test (SPT)

[0194] SPT was conducted from day 15 to day 18. On day 15, a 1% sucrose solution was placed on each side of the mouse cage. On day 16, a 1% sucrose solution and pure water were placed on each side of the mouse cage, with the water bottle positions rotated periodically. After the mice were fasted and deprived of water for 24 hours, they were housed individually on day 17, with a 1% sucrose solution and water placed on each side of the cage, and the weight of each level was recorded. After 24 hours, the weight of the water bottle was recorded again, and the sugar preference index was calculated.

[0195] Sugar water preference index = Sugar water consumption / (Sugar water + Pure water consumption) * 100%.

[0196] (2) Mouse tail suspension test (TST)

[0197] On day 18, a mouse tail suspension device was used for testing. The mouse tail was fixed to the suspension bar, so that the mouse was suspended head upside down. The camera lens was adjusted so that it was horizontally aligned with the mouse's body position. After 2 minutes of adaptation, the video recording was started, and the number of times the mouse did not move within 4 minutes was observed.

[0198] (3) Forced swimming test (FST)

[0199] FST was performed on day 19. The mice were placed in a forced swimming tub (12 cm in diameter) filled with water at a depth of 30 cm and a temperature of 23-25°C. The cumulative time the mice remained still in the water for 5 minutes was recorded.

[0200] (4) Sodium pentobarbital test (PST)

[0201] PST was performed on day 20. Mice were fasted for 12 hours but allowed free access to water. Each mouse was injected intraperitoneally with sodium pentobarbital (45 mg / kg), and the duration of sleep in each group was observed and recorded. The formula for calculating sleep duration is: Sleep duration = Wake time - Sleep onset time.

[0202] (5) Open field test

[0203] The open field test was conducted on day 21. During the test, the laboratory was kept quiet, the room temperature was constant, and the lighting was uniform. Mice were placed in the center of the open field box, and the total distance traveled, the time spent in the central area, and the number of times the mice entered the central area were observed and recorded within 5 minutes. Between the two experiments, mouse feces and urine were removed, and odors were eliminated by spraying with 75% alcohol.

[0204] 2.6 Pathological Examination

[0205] Brain tissue was collected from each group of mice, stained with hematoxylin and eosin (HE) and Nissl stain, sectioned, photographed under a light microscope, and subjected to pathological examination. Based on the histopathological changes in the brain, the degree of brain injury was graded using the following main observation indicators: disordered and loose arrangement of neurons, irregular cell morphology, pyknosis and deep staining of nuclei, ruptured and reduced Nissl bodies, neuronal loss, and vacuolar degeneration.

[0206] "—" indicates that the neurons and Nissl bodies have normal structural morphology.

[0207] "+" indicates a small number of neuronal and Nissl body structures showing pathological changes (lesion area <25%).

[0208] "++" indicates pathological changes in some neurons and Nissl bodies (lesion range >25% & <50%).

[0209] "++" indicates pathological changes in most neurons and Nissl bodies (lesion extent >50%).

[0210] 2.7 Neurotransmitter Detection

[0211] The brain was dissected, rinsed with saline, dried with filter paper, and weighed. A portion of the brain tissue was then homogenized in PBS. Homogenization was performed according to the kit instructions, and the levels of 5-HT, GLU, and GABA were measured.

[0212] 2.8 Data Statistics

[0213] Measurement data adopted Description. Data comparison: Analysis of variance was used for comparisons among multiple groups; nonparametric tests were used for non-normally distributed continuous data (significance level α = 0.05). T-tests were used for analysis of two independent samples (significance level α = 0.05). All data were processed using SPSS 22.0 statistical software.

[0214] 2.9 Experimental Results

[0215] 2.9.1 Effects on behavioral activity and body weight

[0216] Mice were weighed once before the experiment and every three days during the experiment. At the end of the final open field experiment, mice were weighed and euthanized. No abnormal behavior was observed in the control group mice. Mice in the CUMS group exhibited irritability, disordered fur, and huddling behavior. The mean weight of mice in the CUMS group was lower than that of the control group (p < 0.01). Compared with the CUMS group, the weight of mice in the positive control group, soft capsule group, and hard capsule group all increased (p < 0.05), and the weight of mice in the soft capsule group was higher than that in the hard capsule group (p < 0.05). Results are shown in […]. Figure 2 Note: **P<0.01, compared with the CUMS group, #p<0.05, compared with the hard capsule group, p<0.05.

[0217] 2.9.2 Effects on SPT sucrose preference index and number of suspension tail rests

[0218] The saccharide preference test showed that, compared with the control group, the saccharide preference index of mice in the CUMS group was significantly lower (P<0.01). After drug intervention, compared with the CUMS group, the saccharide preference index of mice in the positive control group, soft capsule group, and hard capsule group showed a significant rebound, with statistically significant differences (P<0.05 or P<0.01). The saccharide preference index of mice in the soft capsule group was higher than that in the hard capsule group (P<0.05), suggesting that the soft capsules were more effective than the hard capsules in improving saccharide preference. The tail suspension test showed that, compared with the control group, the number of times mice remained still was significantly increased, with statistical significance (P<0.01). Compared with CUMS, the number of times mice remained still was significantly reduced in the positive control group, soft capsule group, and hard capsule group (P<0.05 or P<0.01). At the same time, the number of times mice remained still was significantly reduced in the soft capsule group compared with the hard capsule group (P<0.05), suggesting that the soft capsules were more effective than the hard capsules in improving the number of times mice remained still. The results are shown in the table below. Figure 3 Note: Compared with the control group, **P<0.01; compared with the CUMS group, #P<0.05, ##P<0.01; compared with the hard capsule group, ·P<0.05.

[0219] 2.9.3 Effects on FST immobility cumulative time and PST sleep duration

[0220] To verify whether soft capsules and hard capsules could improve sleep in depressed mice, the results showed that, compared with the control group, the cumulative immobility time of mice in the CUMS group was prolonged, which was statistically significant (P<0.01). Compared with CUMS, the cumulative immobility time of mice in the positive control group, soft capsule group, and hard capsule group was significantly reduced, which was statistically significant (P<0.05 or P<0.01); the soft capsules were significantly better than the hard capsules in improving the cumulative immobility time of mice (P<0.05).

[0221] PST results showed that, compared with the control group, the sleep duration of mice in the CUMS group was significantly shortened (P<0.01). Compared with the CUMS group, the sleep duration of mice in the positive control group and the soft capsule group was significantly prolonged (P<0.01). The soft capsule was more effective than the hard capsule in improving sleep in mice, but no significant difference was observed. See the results below. Figure 4 Note: Compared with the control group, **P<0.01; compared with the CUMS group, #P<0.05, ##P<0.01; compared with the hard capsule group, ·P<0.05.

[0222] 2.9.4 Effects on changes in motion behavior during open field tests

[0223] The results showed that, compared with the control group, the total distance, center distance, and center entry frequency were all reduced in the CUMS group, which was statistically significant (P<0.05 or P<0.01). Compared with CUMS, the total distance was significantly increased in the positive control group, soft capsule group, and hard capsule group, which was statistically significant (P<0.05 or P<0.01); the center distance was increased in all three groups, which was statistically significant (P<0.01); and the center entry frequency was increased in all three groups, which was statistically significant (P<0.01). Soft capsules had a slightly better effect on the total distance than hard capsules, but no significant difference was observed. Soft capsules had a similar effect on center distance and center entry frequency as hard capsules. (See attached results). Figure 5 Note: Compared with the control group, **P<0.01; compared with the CUMS group, #P<0.05, ##P<0.01.

[0224] 2.9.5 Effects on related neurotransmitters in brain tissue

[0225] The results showed that, compared with the control group, the levels of 5-HT and GABA in the brain tissue of the CUMS group were significantly decreased, while the level of GLU was significantly increased (P<0.01). Compared with CUMS, the levels of 5-HT in the brain tissue of mice in the positive control group, soft capsule group, and hard capsule group were significantly increased (P<0.05 or P<0.01), and the levels of GABA in the brain tissue of mice in the positive control group and soft capsule group were significantly increased (P<0.05 or P<0.01). The levels of GLU in the brain tissue of mice in the positive control group, soft capsule group, and hard capsule group were significantly decreased (P<0.05 or P<0.01). Soft capsules were significantly better than hard capsules at increasing the levels of 5-HT and GABA in mice (P<0.05 or P<0.01), and soft capsules were better than hard capsules at reducing the level of GLU in mouse brain tissue, but no significant difference was observed. (See attached results). Figure 6 Note: Compared with the control group, **P<0.01; compared with the CUMS group, #P<0.05, ##P<0.01; compared with the hard capsule group, ·P<0.05, ··P<0.01.

[0226] 2.9.6 Brain tissue (H&E) pathology and Nissl staining analysis

[0227] Pathological results showed that: ① In the control group (n=8), neurons were structurally intact, neatly arranged, with clear nucleoli and basically normal Nissl bodies. ② In the CUMS group, most neurons showed disordered and loose arrangement. Some cells had irregular morphology, pyknosis, and deep staining; some Nissl bodies were broken and reduced; some showed vacuolar degeneration in the hippocampus; the severity of the lesion was graded (+, 8 / 8). ③ In the positive drug group (n=8): no abnormalities were observed in the brain tissue (-, 3 / 8); a small number of neurons showed disordered and loose arrangement; some cells had irregular morphology, pyknosis, and deep staining; some Nissl bodies were broken and reduced; some showed vacuolar degeneration in the hippocampus (+, 4 / 8). ④ In the hard capsule group (n=8): no abnormalities were observed in the brain tissue (-, 4 / 8); a small number of neurons showed disordered and loose arrangement; some cells had irregular morphology, pyknosis, and deep staining; some Nissl bodies were broken and reduced; some showed vacuolar degeneration in the hippocampus (+, 4 / 8). ⑤ Soft capsule group (8 animals): No abnormalities were observed in the brain tissue of the animals (-, 5 / 8); a small number of neurons showed disordered and loose arrangement; some cells had irregular morphology, pyknosis, and deep staining; some Nissl bodies were broken and reduced; some showed vacuolar degeneration in the hippocampus (+, 3 / 8). Results are as follows. Figure 7 , 8 As shown.

[0228] Depend on Figure 2-8 The results show that the Shenge Bushen soft capsule composition is more effective than the Shenge Bushen hard capsule in treating depression. The technical features of the above embodiments can be combined arbitrarily. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as the combination of these technical features does not contradict each other, it should be considered within the scope of this specification.

[0229] The embodiments described above are merely illustrative of several implementations of the present invention, and while the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the invention patent. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these all fall within the protection scope of the present invention. Therefore, the protection scope of this invention patent should be determined by the appended claims.

Claims

1. A pharmaceutical composition, characterized in that, It includes a first component and a second component, wherein the first component is a soft capsule and the second component is a hard capsule or tablet; the contents of the soft capsule include total flavonoids from Pueraria lobata and Epimedium, an oily matrix, a surfactant and a suspending agent, and the second component includes Codonopsis pilosula polysaccharide extract, a filler and a lubricant.

2. The pharmaceutical composition according to claim 1, characterized in that, The oily matrix in the contents of the soft capsule comprises 20%-90% by mass; and / or, The surfactant in the contents of the soft capsule accounts for 0.2%-0.6% by mass; and / or, The suspending agent in the contents of the soft capsule accounts for 0.2%-0.6% by mass.

3. The pharmaceutical composition according to claim 1, characterized in that, The oily matrix includes one or more of vegetable oils, medium-chain triglycerides, and polyethylene glycol; Optionally, the vegetable oil includes one or more of soybean oil and peanut oil; and / or, The surfactant includes Tween 80; and / or, The suspending agent includes one or more of xanthan gum and beeswax.

4. The pharmaceutical composition according to claim 1, characterized in that, The capsule shell material of the soft capsule includes gelatin, glycerin and water; Optionally, the mass ratio of the gelatin to the glycerin is 1:(0.1-0.5); Optionally, the mass ratio of the gelatin to the water is 1:(0.8-1.2).

5. The pharmaceutical composition according to claim 1, characterized in that, The total polysaccharide content of the Codonopsis pilosula polysaccharide extract is 50%-90% by mass; and / or, By weight, the second component comprises: 150-250 parts of Codonopsis pilosula polysaccharide extract, 40-60 parts of filler, and 1-3 parts of lubricant; and / or, The filler includes one or more of starch and microcrystalline cellulose; and / or, The lubricant includes one or more of magnesium stearate, talc, and silicon dioxide.

6. The pharmaceutical composition according to any one of claims 1 to 5, characterized in that, The dosage of the first component, calculated based on the total flavonoids of Pueraria lobata and Epimedium, is 90mg-110mg; the dosage of the second component, calculated based on the polysaccharide extract of Codonopsis pilosula, is 200mg-230mg.

7. The pharmaceutical composition according to claim 6, characterized in that, The ratio of the number of the first component to the number of the second component is 1:

2.

8. A method for preparing a pharmaceutical composition, characterized in that, Includes the following steps: The contents were prepared by mixing total flavonoids from Pueraria lobata and Epimedium, an oily matrix, a surfactant, and a suspending agent. A gel solution is prepared using the material for preparing soft capsules, and the gel solution and the contents are pressed together to prepare soft capsules, which serve as the first component. The polysaccharide extract of Codonopsis pilosula, fillers, and lubricants are mixed to prepare hard capsules or tablets as a second component.

9. The method for preparing the pharmaceutical composition according to claim 8, characterized in that, The method for preparing the total flavonoids from Pueraria lobata and Epimedium includes: The medicinal materials of kudzu root and epimedium were refluxed and extracted to prepare an alcohol extract; The alcohol extract is concentrated to prepare a concentrated solution; The concentrate was passed through a resin column; The resin column was washed with water to remove impurities and with alcohol to elute flavonoids, and the eluent was collected. The eluent was concentrated under reduced pressure and dried to prepare the total flavonoids from Pueraria lobata and Epimedium; and / or, The preparation method of the total extract of Codonopsis pilosula polysaccharides includes: The medicinal material of Codonopsis pilosula was soaked and extracted to prepare an aqueous extract; The aqueous extract was concentrated under reduced pressure to prepare a concentrated extract of Codonopsis pilosula. Mix the concentrated extract of Codonopsis pilosula with ethanol, stir, let stand and collect the precipitate, add water to dissolve it, add ethanol again, and collect the precipitate again. The precipitate was washed and dried to prepare the Codonopsis pilosula polysaccharide extract.

10. Use of at least one of the pharmaceutical compositions according to any one of claims 1 to 7 and the pharmaceutical compositions prepared by the preparation method of claim 8 or 9 in the preparation of a medicament for treating depression.

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