CN112957387B

CN112957387B - Traditional Chinese medicine composition for improving asthenopia, preparation method and application thereof

Info

Publication number: CN112957387B
Application number: CN202110467274.1A
Authority: CN
Inventors: 吴素香; 石森林; 任敏霞; 郑碧莹
Original assignee: Zhejiang Chinese Medicine University ZCMU
Current assignee: Zhejiang Chinese Medicine University ZCMU
Priority date: 2021-04-28
Filing date: 2021-04-28
Publication date: 2022-08-02
Anticipated expiration: 2041-04-28
Also published as: CN112957387A

Abstract

The invention discloses a traditional Chinese medicine composition for improving asthenopia, which comprises raw material medicines of chrysanthemum morifolium and safflower; the invention also discloses a preparation method of the crude drug extract, which comprises the steps of taking the crude drug, heating and refluxing the crude drug by using ethanol to obtain ethanol extract, removing the ethanol from the ethanol extract, then loading the ethanol extract on a macroporous adsorption resin column, removing impurities by using NaCl solution, then eluting by using ethanol, and drying the eluent to obtain the crude drug extract; the invention also discloses a preparation method of the effervescent granule of the traditional Chinese medicine composition, which comprises the steps of taking 5-25 parts of raw material medicine extract, 5-25 parts of diluent and 50-70 parts of effervescent disintegrant, uniformly mixing the raw material medicine extract and the diluent, dividing into two parts, uniformly mixing one part of the mixture with citric acid to prepare acid granules, uniformly mixing the other part of the mixture with sodium bicarbonate coated by PEG-6000 to prepare alkali granules, and uniformly mixing the acid granules and the alkali granules to obtain effervescent granules; the invention also discloses application of the traditional Chinese medicine composition as a medicine for improving visual fatigue, and the curative effect of improving visual fatigue is obvious.

Description

Traditional Chinese medicine composition for improving asthenopia, preparation method and application thereof

Technical Field

The invention relates to the field of traditional Chinese medicines, in particular to a traditional Chinese medicine composition for improving visual fatigue, a preparation method and application thereof.

Background

The asthenopia is a syndrome of interweaving organic factors and psychosocial factors of eyes or the whole body, and belongs to the category of psychosomatic medicine; the eye fatigue syndrome is a group of symptoms which are manifested by visual disturbance, eye discomfort and general symptoms after eyes are used, so that the visual operation cannot be normally carried out, and is also commonly called as eye fatigue syndrome clinically. In recent years, with the increase of social rhythm and the increase of pressure in all aspects of work, life, spirit and the like, the incidence rate of asthenopia tends to rise year by year in long-term work and life of computers, and asthenopia patients also tend to be younger. The asthenopia not only can seriously interfere the vision and the life quality of patients, but also can further cause eye diseases such as age-related macular degeneration, cataract, blindness and the like. Therefore, the active development and enhancement of research on the prevention and treatment of the disease are of great significance.

The causes of asthenopia mainly include ametropia, binocular disparity, binocular anisotropic motor dysfunction, parasympathetic excitation, vergence regulation abnormality, xerophthalmia, the influence of visual display terminal VDT, and the like. The clinical common treatment methods comprise ametropia correction, eye position correction, extraocular muscle training, auricular point pressure application, operation methods, treatment of chemical drugs and traditional Chinese medicines, life and working habit change and the like. The curative effect is more definite by using surgical treatment and prism correction, but the use face is narrow, so that the requirements of preventing and treating the asthenopia cannot be met. The chemical medicine is mainly used for treating local symptoms, and the traditional Chinese medicine is based on the whole and carries out treatment based on syndrome differentiation, so that the traditional Chinese medicine has characteristics and advantages in the aspect of treating asthenopia syndrome.

The visual fatigue is similar to the traditional Chinese medicine ophthalmology liver fatigue, the liver fatigue is firstly seen in the Qianjin Yao Fang Jiu Meng far away, the pathogenesis is thought to look out at the utmost and consume essence and blood in darkness. The relation between heart and visual fatigue is demonstrated in Huangdi's inner channel essence questions and' Yao's letter of examination, internal and external two barrier treatises', the eyes are thought not to be dazzling, the eyes will stay in the interior, the interior stays there, and the heart fire moves, so the eyes will feel painful. The theory of traditional Chinese medicine holds that the occurrence of asthenopia is the eye-orifice malnutrition caused by liver and kidney deficiency, spleen deficiency and qi weakness, heart and blood deficiency, liver depression and qi stagnation and the like, and is related to heart, liver and kidney, so that the traditional Chinese medicines for tonifying qi, soothing liver, strengthening spleen and tonifying kidney are commonly used for preventing and treating in clinical treatment.

The traditional Chinese medicine composition disclosed by the prior art has the problems of complex prescription, unobvious drug effect and the like.

Disclosure of Invention

An object of the present invention is to provide a Chinese medicinal composition for improving asthenopia, which overcomes the above-mentioned disadvantages of the prior art. The Chinese medicinal composition has simple prescription, and obvious curative effect of improving asthenopia.

The invention provides a technical scheme for solving a technical problem, and provides a traditional Chinese medicine composition for improving visual fatigue, wherein the traditional Chinese medicine composition comprises the following raw material medicines in parts by mass: 16-24 parts of Hangzhou white chrysanthemum and 8-12 parts of safflower.

Preferably, the traditional Chinese medicine composition comprises the following raw material medicines in parts by mass: 19-21 parts of Hangzhou white chrysanthemum and 9.5-10.5 parts of safflower.

Preferably, the formulation of the Chinese medicinal composition comprises effervescent granules.

Preferably, the effervescent granule comprises the following components in parts by weight: 5-25 parts of crude drug extract, 5-25 parts of diluent and 50-70 parts of effervescent disintegrant;

the diluent comprises: at least one of dextrin, lactose, and D-mannitol;

effervescent disintegrants include: citric acid, sodium bicarbonate coated with PEG-6000; in the sodium bicarbonate coated by PEG-6000, the mass ratio of the PEG-6000 to the sodium bicarbonate is (0.5-1) to 1; the mass ratio of the citric acid to the sodium bicarbonate is (1.0-1.7) to 1;

the raw material medicine extract is prepared by the following steps:

a1: taking 16-24 parts by weight of chrysanthemum morifolium and 8-12 parts by weight of safflower, heating and refluxing the chrysanthemum morifolium and the safflower by using ethanol to obtain an ethanol extract, and removing the ethanol from the ethanol extract to obtain an ethanol-free extract;

a2: loading macroporous adsorption resin into a column, then sampling alcohol-free extract on the macroporous adsorption resin column, removing impurities from the macroporous adsorption resin column by using NaCl solution, then eluting by using ethanol to obtain eluent, and drying the eluent to obtain a crude drug extract; the macroporous adsorbent resin comprises at least one of H1020, LS206, HPD600 and LSA-10.

Compared with the prior art, the traditional Chinese medicine composition for improving the visual fatigue has the following beneficial effects:

(1) the Chinese medicinal composition has simple prescription and is prepared from flos Chrysanthemi,Is prepared from safflower, and has obvious curative effect on improving asthenopia. Hangzhou white chrysanthemum is responsible for lung and liver channel, has functions of dispelling wind and clearing heat, calming liver and improving eyesight, and clearing heat and detoxicating, and can be used for treating conjunctival congestion, swelling and pain, blurred vision, etc. The effective material basis of the Hangzhou white chrysanthemum is mainly flavonoid which can reduce the occurrence of xerophthalmia, inhibit the apoptosis of lacrimal gland acinus and glandular cells, improve the basic secretion of lacrimal gland and maintain the stability of lacrimal film. Luteolin is one of indexes for controlling quality of flos Chrysanthemi, and has antipyretic, antiinflammatory, and capillary vasodilatation enhancing effects. Carthami flos is Compositae plant, and has effects of invigorating blood circulation, dredging channels, removing blood stasis, relieving pain, protecting cardiovascular system, resisting thrombi, resisting cerebral ischemia injury, protecting nerve, enhancing immunity, relieving inflammation and pain, resisting tumor, resisting bacteria, relieving fatigue, and protecting liver. The main medicinal component of safflower is safflower yellow which is a natural yellow extracted from petals of safflower and is a chalcone compound. The hydroxy safflower yellow A is the main component of safflower yellow, and the safflower yellow has scavenging effect on oxygen free radical, and can remarkably antagonize H ₂ O ₂ The induced oxidative damage and apoptosis can reduce the inflammatory reaction induced by oxygen free radicals, and the like, and can effectively prevent and treat the photochemical damage of the retina by removing excessive free radicals. The combination of the two has obvious curative effect on dry eyes, cataract and blurred eyes.

(2) Clinical experiment results show that the total effective rate of the traditional Chinese medicine composition for improving the clinical symptoms of the asthenopia is 95.8%, and the effect is obvious; the traditional Chinese medicine composition disclosed by the invention has the advantages that the photopic vision persistence percentage is improved by 19.34%, the effect is obvious, and the popularization and application values are realized.

(3) The traditional Chinese medicine composition is prepared into effervescent granules, the effervescent granules can be quickly dissolved in water and can be uniformly dispersed in water, and the effervescent granules have the characteristics of convenient taking, quick response, high bioavailability and the like, have good mouthfeel, have the medicinal experience like drinking beverages, are easily accepted by patients, are suitable for people of different ages, and are particularly suitable for the old, children and patients with difficulty in swallowing solid preparations.

(4) The diluent of the effervescent granule is generally selected to have good water solubility, and meanwhile, the effervescent granule is easy to react and lose the effervescence when being wetted, so the diluent of the effervescent granule also has strong moisture absorption resistance, comprising: at least one of dextrin, lactose, and D-mannitol; when the effervescent granules are granulated, the diluent D-mannitol has excellent granulation difficulty, clarity and effervescence time, and the raw material medicine extract of the effervescent granules is light in smell and bitter in taste, and the D-mannitol has the function of a flavoring agent, so the D-mannitol is preferably used as the diluent for preparing the effervescent granules.

Another object of the present invention is to provide a method for preparing an extract of a crude drug for improving asthenopia, which overcomes the above-mentioned disadvantages of the prior art. The crude drug extract prepared by the preparation method can obviously improve asthenopia.

The invention solves another technical problem by adopting a technical scheme that a preparation method of a raw material medicine extract for improving visual fatigue is provided, and comprises the following steps:

s1: taking 16-24 parts by weight of chrysanthemum morifolium and 8-12 parts by weight of safflower, heating and refluxing the chrysanthemum morifolium and the safflower by using ethanol to obtain an ethanol extract, and removing the ethanol from the ethanol extract to obtain an ethanol-free extract;

s2: loading macroporous adsorption resin into a column, then sampling alcohol-free extract on the macroporous adsorption resin column, removing impurities from the macroporous adsorption resin column by using NaCl solution, then eluting by using ethanol to obtain eluent, and drying the eluent to obtain a crude drug extract; the macroporous adsorbent resin comprises at least one of H1020, LS206, HPD600 and LSA-10.

Preferably, the preparation method of the crude drug extract for improving the asthenopia comprises the following steps:

s1: taking 19-21 parts by mass of chrysanthemum morifolium and 9.5-10.5 parts by mass of safflower, heating and refluxing for 1-4 times by using 10-25 times of 20-80% ethanol by mass, wherein the extraction time is 0.5-2 h each time, combining ethanol extract, concentrating under reduced pressure until no ethanol smell exists, carrying out centrifugal separation, taking supernatant, and obtaining the supernatant which is the ethanol-free extract;

s2: loading macroporous adsorption resin into a column by a wet method according to the diameter-height ratio of 1: 3-7, loading 0.5BV-2BV of alcohol-free extract on the macroporous adsorption resin column at the flow rate of 1BV/h-3BV/h, removing impurities from the macroporous adsorption resin column by 0.72BV-1.26BV of NaCl solution with the concentration of 0.05mol/L-0.2mol/L at the flow rate of 1BV/h-3BV/h, eluting by 1.8BV-2.7BV of 40-80% ethanol at the flow rate of 1BV/h-3BV/h, discarding the former 0.5BV-0.6BV, collecting the rest of effluent as eluent, and performing vacuum drying on the eluent to obtain a crude drug extract; the macroporous adsorption resin comprises LS206 macroporous adsorption resin;

in this patent, the volume of the resin column loaded with resin is called bed volume (bed volume), abbreviated as BV, which is the basic unit of the resin column, and it works with various amounts of material in BV units, for example, the flow rate of the solution through the resin column is 2BV/h to 4BV/h, i.e., the volume of solution passing through the resin bed per hour is 2 to 4 times the volume of the resin bed.

Compared with the prior art, the preparation method of the bulk drug extract for improving the asthenopia has the following beneficial effects:

(1) animal experiment results show that the crude drug extract prepared by the preparation method has a good relaxation effect on rat stomach smooth muscle, and has the effect of obviously increasing the auricle microcirculation blood flow of a mouse.

(2) Because ethanol is easy to elute substances adsorbed by macroporous adsorption resin, the ethanol extract needs to be concentrated under reduced pressure until no alcohol smell exists, and then can be loaded on a macroporous adsorption resin column, but partial substances are precipitated during the concentration under reduced pressure, so that the supernatant needs to be centrifuged for experiment.

(3) In the purification process, pure water or low-concentration ethanol solution is generally adopted for impurity removal at present, a NaCl solution is used for impurity removal in a new way, and the three advantages of low total flavone loss rate, more impurity-removed total cured substances and high impurity content in the impurity-removed cured substances in the impurity removal process are realized, so that the impurity removal process is improved.

(4) During the collection of the eluate, it was found that the initially effluent fraction had an excessive content of impurities, resulting in a lower overall purity of the effluent; after the segmented collection and investigation, the former 0.5BV-0.6BV effluent liquid is preferably discarded, so that the total flavone purity is effectively improved while the total flavone transfer rate is ensured.

(5) Aiming at the characteristics of ethanol-free extracts of chrysanthemum morifolium and safflower, by using the content of total flavonoids as an index and adopting a method combining static adsorption and dynamic adsorption, macroporous adsorption resin with excellent adsorption capacity and elution capacity on the total flavonoids is found, and the optimal macroporous adsorption resin impurity removal process with small leakage amount, high transfer rate, low loss rate and high impurity removal amount of the total flavonoids is realized by further observing factors such as sample loading amount, sample loading speed, diameter-height ratio, impurity removal solvent type and dosage and the like. The macroporous adsorption resin purification method provided by the invention does not need to adjust the pH value, has simple steps, is more environment-friendly and has good purification effect.

Another objective of the present invention is to provide a method for preparing a Chinese medicinal composition for relieving asthenopia. The effervescent granule prepared by the preparation method is more uniform and has good stability.

The invention solves a technical problem by adopting the technical scheme that a preparation method of a traditional Chinese medicine composition for improving visual fatigue is provided, the preparation of the traditional Chinese medicine composition comprises effervescent granules, and the preparation method of the effervescent granules comprises the following steps:

b1: heating and melting PEG-6000, adding sodium bicarbonate, stirring, cooling, and pulverizing into fine powder to obtain PEG-6000-coated sodium bicarbonate; the mass ratio of PEG-6000 to sodium bicarbonate is (0.5-1) to 1;

b2: taking 5-25 parts of raw material medicine extract, 5-25 parts of diluent and 50-70 parts of effervescent disintegrant according to parts by weight; effervescent disintegrants include: citric acid, sodium bicarbonate coated with PEG-6000; the mass ratio of the citric acid to the sodium bicarbonate is (1.0-1.7) to 1;

b3: uniformly mixing the raw material medicine extract and the diluent in the step B2, and dividing into two parts to obtain a mixture of the two parts of the raw material medicine extract and the diluent;

b4: mixing a part of the mixture of the crude drug extract and the diluent in the step B3 with citric acid, adding a wetting agent, granulating by a wet method, and drying to prepare acid granules; mixing the other part of the mixture of the crude drug extract and the diluent in the step B3 with sodium bicarbonate coated with PEG-6000, adding wetting agent, granulating by wet method, drying, and making into alkali granule;

b5: and D, uniformly mixing the acid granules and the alkali granules in the step B4, and finishing granules to obtain the effervescent granules.

Preferably, the preparation method of the effervescent granule comprises the following steps:

b1: heating and melting PEG-6000 to 61-70 deg.C, adding sodium bicarbonate, stirring, cooling, and pulverizing into fine powder to obtain PEG-6000-coated sodium bicarbonate; the mass ratio of PEG-6000 to sodium bicarbonate is (0.65-0.85) to 1;

b2: taking 10-20 parts of raw material medicine extract, 10-20 parts of diluent and 50-70 parts of effervescent disintegrant according to parts by weight; effervescent disintegrants include: citric acid, sodium bicarbonate coated with PEG-6000; the mass ratio of the citric acid to the sodium bicarbonate is (1.3-1.5) to 1;

b3: uniformly mixing the crude drug extract and the diluent in the step B2, and dividing into two equal parts to obtain a mixture of two crude drug extracts and the diluent;

b4: mixing one part of the crude drug extract obtained in step B3 with diluent, adding citric acid, mixing, adding wetting agent, wet granulating, drying at 40 deg.C, and making into acid granule; mixing the other part of the extract of the raw material medicine obtained in the step B3 with diluent, adding sodium bicarbonate coated with PEG-6000, mixing, adding wetting agent, wet granulating, drying at 40 deg.C, and making into alkali granule;

b5: and D, uniformly mixing the acid granules and the alkali granules in the step B4, and granulating to obtain the effervescent granules.

Preferably, the diluent is D-mannitol; the wetting agent is 75-95% ethanol.

Compared with the prior art, the preparation method of the traditional Chinese medicine composition for improving the visual fatigue has the following beneficial effects:

(1) the preparation method of the invention is scientific, reasonable and advanced, and the effervescent granules with uniform granules, good stability, safety and effectiveness are successfully prepared.

(2) Ethanol is used as a wetting agent, water is generally used as the wetting agent, and the ethanol is selected as the wetting agent due to the fact that the extract of the raw material medicine has high viscosity and is easy to absorb moisture. The concentration of the wetting agent ethanol is preferably 75-95%, because the smaller the concentration of the wetting agent, that is, the larger the water content, the lower the granule forming rate, and the moisture induces the hygroscopicity of the extract, so that the mixture becomes viscous and unfavorable for granulation. When 85% ethanol is selected as the wetting agent, the dissolving time is short and the forming rate is high.

(3) Because sodium bicarbonate is easily decomposed by heat at high temperature, and the effervescence effect is influenced, the PEG-6000 is heated and melted, and the sodium bicarbonate is added when the temperature is controlled to be 61-70 ℃, so that the decomposition of the sodium bicarbonate is reduced. Due to the characteristics of high viscosity and high moisture absorption of the crude drug extract, the quality of the crude drug extract in the granulation process is increased due to water absorption, even the crude drug extract absorbs water to form a block, in the granulation process, the crude drug extract and the diluent are uniformly mixed, the crude drug extract mixed with the diluent has high moisture absorption resistance and is more stable in the environment, and errors caused by moisture in the weighing process are avoided. In order to solve the problem, the mixed extract of the raw material medicines is divided into two parts, and the two parts of mixture are respectively mixed with citric acid and the sodium bicarbonate coated by the PEG-6000 to prepare acid particles and alkali particles by adopting an acid-alkali separation granulation method. The effervescent granules finally prepared by the invention have the advantages of uniform granules, good stability and good moisture absorption resistance.

Another objective of the present invention is to provide an application of the above traditional Chinese medicine composition, or the crude drug extract prepared by the above method, or the traditional Chinese medicine composition prepared by the above method as a medicine for improving asthenopia, wherein the medicine has a significant effect on improving asthenopia.

Drawings

Fig. 1 is a photograph of a Chinese medicinal effervescent granule prepared in example 8 of the present invention.

FIG. 2 is a diagram showing the ultraviolet absorption spectra of the rutin control solution and the test solution in comparative example 1.

Detailed Description

The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and the described embodiments are only some embodiments, but not all embodiments, of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments of the present invention without making any creative effort, shall fall within the protection scope of the present invention.

Example 1

A preparation method of a crude drug extract for improving asthenopia comprises the following steps:

s1: according to the mass parts, 8g of chrysanthemum morifolium and 4g of safflower are taken, heated and refluxed for 3 times by using 40 percent ethanol with the mass being 20 times that of the chrysanthemum morifolium and the safflower, the extraction time is 2 hours each time, the ethanol extract is combined, decompressed and concentrated until no alcohol smell exists, the ethanol extract is centrifugally separated in a centrifuge (25 ℃, 7000r/min and 30min), and the supernatant is taken as the alcohol-free extract; preparing a loading solution of 0.2g crude drug/mL by using the alcohol-free extracting solution, wherein the pH value is 4.90, and the weight of the crude drug is calculated by the weighed raw materials; because the ethanol is easy to elute substances adsorbed by the macroporous resin, the ethanol extract can be loaded on the macroporous adsorption resin column after being decompressed and concentrated until no alcohol smell exists, but partial substances are precipitated during decompression and concentration, and the supernatant liquid is taken for experiment after centrifugation. The ethanol is heated and refluxed to extract the total flavone, hydroxy safflower yellow A and luteolin with higher content than that obtained by ultrasonic extraction. The total flavone is rutin (C) ₂₇ H ₃₀ O ₁₆ ) And (6) counting.

S2: taking LS206 macroporous adsorption resin, performing wet column packing according to the diameter-height ratio of 1:5(3cm 15cm), then taking 1.3BV of sample loading liquid of 0.2g crude drug/mL to sample the macroporous adsorption resin column at the flow rate of 2BV/h, then removing impurities from the macroporous adsorption resin column at the flow rate of 2BV/h by using 0.9BV NaCl solution with the concentration of 0.1mol/L, then eluting the macroporous adsorption resin column at the flow rate of 2BV/h by using 2.5BV ethanol with the concentration of 60%, discarding the first 0.6BV effluent, collecting the rest effluent as eluent, and performing vacuum drying on the eluent to obtain a crude drug extract;

example 2

s1: taking 16g of chrysanthemum morifolium ramat and 12g of safflower according to parts by weight, heating and refluxing the chrysanthemum morifolium ramat and 12g of safflower by using 20% ethanol with the mass being 10 times that of the chrysanthemum morifolium ramat for 4 times, wherein the extraction time is 0.5h each time, merging ethanol extract, concentrating the ethanol extract under reduced pressure until no ethanol smell exists, carrying out centrifugal separation, taking supernatant, and obtaining the supernatant which is the ethanol-free extract;

s2: taking H1020 macroporous absorption resin, loading the H1020 macroporous absorption resin into a column by a wet method according to the diameter-height ratio of 1:3, then sampling 0.5BV of alcohol-free extract on the macroporous absorption resin column at the flow rate of 1BV/H, then removing impurities from the macroporous absorption resin column at the flow rate of 1BV/H by using 0.72BV of NaCl solution with the concentration of 0.05mol/L, then eluting the macroporous absorption resin column at the flow rate of 1BV/H by using 1.8BV of ethanol with the concentration of 40%, discarding the former 0.5BV effluent, collecting the rest effluent as eluent, and drying the eluent in vacuum to obtain the crude drug extract.

Example 3

s1: according to the mass parts, taking 24g of chrysanthemum morifolium and 8g of safflower, heating and refluxing the chrysanthemum morifolium and the safflower for 1 time by using 80% ethanol with the mass of 25 times, wherein the extraction time is 1.5h each time, combining ethanol extract, concentrating the ethanol extract under reduced pressure until no ethanol smell exists, carrying out centrifugal separation, taking supernatant, and obtaining the supernatant which is the ethanol-free extract;

s2: taking HPD600 macroporous adsorption resin, filling the macroporous adsorption resin into a column by a wet method according to the diameter-height ratio of 1:5, then sampling 1.3BV of alcohol-free extract on the macroporous adsorption resin column at the flow rate of 3BV/h, then removing impurities from the macroporous adsorption resin column at the flow rate of 3BV/h by using 1.26BV of NaCl solution with the concentration of 0.2mol/L, then eluting the mixture by using 2.7BV of ethanol with the concentration of 80% at the flow rate of 3BV/h, discarding the previous 0.57BV effluent, collecting the rest effluent as eluent, and drying the eluent in vacuum to obtain the crude drug extract.

Example 4

s1: according to the mass parts, 19g of chrysanthemum morifolium and 10.5g of safflower are taken, heated and refluxed for 1 time by using 80% ethanol with the mass of 25 times, the extraction time is 1.5h each time, the ethanol extract is combined, the ethanol extract is concentrated under reduced pressure until no alcohol smell exists, the centrifugation is carried out, the supernatant fluid is taken, and the supernatant fluid is the alcohol-free extract;

s2: mixing LSA-10 and LS206 macroporous adsorption resins, performing wet column packing according to the diameter-height ratio of 1:7, loading 2BV of alcohol-free extract on the macroporous adsorption resin column at the flow rate of 3BV/h, removing impurities from the macroporous adsorption resin column at the flow rate of 3BV/h by using 1.26BV of NaCl solution with the concentration of 0.2mol/L, eluting by using 2.7BV of ethanol with the concentration of 80% at the flow rate of 3BV/h, discarding the former 0.57BV of effluent, collecting the rest of effluent as eluent, and performing vacuum drying on the eluent to obtain the crude drug extract.

Example 5

s1: according to the mass parts, 21g of chrysanthemum morifolium and 9.5g of safflower are taken, heated and refluxed for 3 times by using 40% ethanol with the mass being 20 times that of the chrysanthemum morifolium and the safflower, the extraction time is 2 hours each time, the ethanol extract is combined, the ethanol extract is concentrated under reduced pressure until no ethanol smell exists, the centrifugal separation is carried out, the supernatant fluid is taken, and the supernatant fluid is the ethanol-free extract;

s2: taking LSA-10 macroporous adsorption resin, filling the column according to a wet method with the diameter-height ratio of 1:5, then sampling 1.3BV of alcohol-free extract on the macroporous adsorption resin column at the flow rate of 2BV/h, then removing impurities from the macroporous adsorption resin column at the flow rate of 2BV/h by using 0.9BV of NaCl solution with the concentration of 0.1mol/L, then eluting the column at the flow rate of 2BV/h by using 2.5BV of ethanol with the concentration of 60%, discarding the previous 0.6BV effluent, collecting the rest effluent as eluent, and drying the eluent in vacuum to obtain the crude drug extract.

Example 6

s1: according to the mass parts, taking 207g of chrysanthemum morifolium and 104g of safflower, heating and refluxing the chrysanthemum morifolium and the safflower for 3 times by using 40% ethanol with the mass being 20 times that of the chrysanthemum morifolium and the safflower, extracting for 2 hours each time, combining ethanol extract, concentrating the ethanol extract under reduced pressure until no ethanol smell exists, carrying out centrifugal separation, taking supernatant, and obtaining the supernatant which is the ethanol-free extract;

s2: mixing H1020, LS206, HPD600 and LSA-10 macroporous adsorption resins, performing wet column packing according to the diameter-height ratio of 1:5, loading 1.3BV of alcohol-free extract on the macroporous adsorption resin column at the flow rate of 2BV/H, removing impurities from the macroporous adsorption resin column at the flow rate of 2BV/H by using 0.9BV of NaCl solution with the concentration of 0.1mol/L, eluting by using 2.5BV of ethanol with the concentration of 60% at the flow rate of 2BV/H, discarding the former 0.6BV of effluent, collecting the rest of effluent as eluent, and performing vacuum drying on the eluent to obtain the crude drug extract.

Example 7

Pharmacodynamic study of the crude drug extract:

the anti-asthenopia drug effect of the crude drug extract is evaluated in isolated tissues and organs.

The raw material medicine extract: the crude drug extract was prepared as in example 1.

The alcohol extract of the raw material medicine: the alcohol-free extract prepared according to the example 1 is decompressed, concentrated and dried in vacuum to obtain the alcohol extract of the raw material medicine. Compared with the extract of the raw material medicine, the alcohol extract of the raw material medicine is not purified by macroporous absorption resin, and is not subjected to impurity removal by NaCl solution.

Clinical studies have shown that the formation of asthenopia is closely related to ocular factors, which influence the accommodative ability of the eyes. The ciliary muscle, during contraction, can alter the focusing force controlled by the lens, thereby focusing the eye on an object. Therefore, overuse of the eye can cause fatigue of the ciliary muscle, and as the function of the ciliary muscle is reduced, the accommodation ability of the eye is gradually reduced or even lost, thereby causing asthenopia. The ciliary muscle of rat is small and is easy to cause physical damage or breakage during separation process, because a plurality of mucous membranes are adhered to the surface of the ciliary muscle and are difficult to be stripped. The ciliary muscle is composed of smooth muscle fibers, the structure and the movement law of the ciliary muscle are similar to those of the stomach smooth muscle, the stomach smooth muscle is large and good in elasticity, and tissues are not easy to damage during separation, so that the stomach smooth muscle of a rat is selected as an experimental object.

(I) the relaxation effect of the extract of the raw material medicine on the smooth muscle of the stomach of a rat

1. Preparation of rat stomach smooth muscle specimen

Cutting off the head of an SD rat, killing the SD rat, immediately cutting the stomach, soaking the stomach in Krebs liquid, cutting the longitudinal muscle of the stomach body, fixing the longitudinal muscle on a glass slide soaked with the Krebs liquid, removing a mucous layer, cutting a gastric muscle strip specimen with the length of about 2.0cm and the width of about 0.3cm by using a blade, and knotting two ends of the muscle strip by using No. 1 suture lines. Placing in a constant temperature water bath (Krebs liquid 15ml, introducing oxygen) at 37 deg.C, fixing one end on an L-shaped ventilation rod, connecting the other end with a tension transducer, and recording the tension change with MPA2000 biological signal quantitative recording and analyzing system.

Preparation of Krebs solution

Krebs solution contains 8.0g NaCl, 0.2g KCl and MgCl ₂ 0.1g、NaH ₂ PO ₄ 0.05g、NaHCO ₃ 1.0g、CaCl ₂ 0.2g and glucose 1.0 g. Dissolving the above components with small amount of distilled water, respectively, and mixing (except CaCl) ₂ Solution and glucose) and adding CaCl ₂ Adding the solution into the mixed solution, dropwise adding while stirring, otherwise generating calcium salt precipitate, adding glucose when in use, and supplementing distilled water to a constant volume of 1000 mL.

3. Method of administration and myotonometry

The administration dose is expressed as crude drug amount/animal body weight (mg/kg). The administration dose of the bulk drug alcohol extract group is 1080mg/kg, and the administration doses of the bulk drug extract in the low, medium and high dose groups are 540mg/kg, 1080mg/kg and 2160mg/kg respectively. The administration dosage of the positive control Qijudihuang oral liquid group is 1.8 mL/kg. All drugs were dissolved or diluted in Krebs solution. A Krebs liquid negative control group was set. Adding 15mL Krebs solution into the water bath, stabilizing the specimen for 1h, changing the solution every 15min during the period, starting the experiment after the muscle bar is stabilized, and recording the muscle tension (X) before administration. After 5min, the medicine is administrated, 6mL of the alcohol extract of the raw material medicine, 6mL of the positive control medicine, 6mL of the low, medium and high dose groups of the extract of the raw material medicine and 6mL of Krebs liquid are respectively added into the Krebs negative control group, so that the final concentration of the medicine in the water bath is 14.4mg/mL of the alcohol extract group of the raw material medicine, 0.024mL/mL of the positive control group, 7.2mg/mL of the low dose group of the extract of the raw material medicine, 14.4mg/mL of the medium dose group of the extract of the raw material medicine and 28.8mg/mL of the high dose group of the extract of the raw material medicine. Changes in muscle tone were observed after 5min of drug action and muscle tone was recorded. The rate of change of muscle tone is calculated according to the formula:

rate of change of muscle tone [ [ (X) _{After administration} -X _{Before administration} )/X _{Before administration} ]×100％

4. Results of the experiment

The muscle tone and its rate of change before and after each group administration are shown in Table 1. As can be seen from the results, there was no significant change in the muscular tension of Krebs group compared to that before administration, and the contractile tension of the gastric bands of the remaining groups was significantly decreased (P < 0.01). Compared with the Krebs group, the change rate of the muscle tension of each group is obviously increased (P is less than 0.01); compared with the bulk drug alcohol extract group, the positive control, the bulk drug extract high-dose group and the dose group in the bulk drug extract have obviously increased muscle tension change rate (P is less than 0.01). Therefore, the crude drug extract with a certain concentration has a direct relaxation effect on isolated gastric smooth muscle of a rat, can effectively reduce the contraction tension of the isolated gastric smooth muscle of the rat, has an effect superior to that of the crude drug alcohol extract, and has a certain concentration dependence.

TABLE 1 Change in muscle tone before and after administration: (

n＝6)

Before and after administration ^△△ P is less than 0.01, and the difference is extremely significant; compared with Krebs group ^** P is less than 0.01, and the difference is extremely significant; compared with the alcohol extract of the raw material medicine ^## P is less than 0.01, and the difference is very significant. n-6 indicates 6 replicates and the data in the table is the average of 6 replicates. n represents the number of repeated experiments, and the same applies below, unless otherwise specified.

(II) influence of crude drug extract on mouse auricle microcirculation blood flow

1. Grouping animals

50 clean ICR mice are selected, half of the mice are female and half of the mice, the weight of the mice is 18-22g, and the mice are fed adaptively for 7 days and are eaten with normal water. The weight of the medicine is randomly divided into 5 groups, namely a normal control group, a raw material medicine alcohol extract group, a raw material medicine extract low, medium and high dose group, and each group contains 10 medicines.

2. Method of administration

Perfusing the normal control group with distilled water for 7 days; the raw material medicine alcohol extract and the raw material medicine extract are dissolved by using a proper amount of distilled water, the gavage dose of the raw material medicine alcohol extract group is 1560mg/kg, the gavage doses of the raw material medicine extract groups with low, medium and high doses are 780mg/kg, 1560mg/kg and 3120mg/kg respectively, and the gavage is performed for 7 days.

3. Index detection

After 7 days of dosing, the mouse auricle microcirculation was detected by the moorflp pi speckle full-frame real-time scanning imaging system. Under the waking state, the auricle of the mouse is placed in the visual field right below the scanning head, 5 measurement areas with 5 × 5 pixel areas are selected, and the subcutaneous blood flow of 1mm is measured for 1 min.

4. Results of the experiment

As shown in Table 2, compared with the normal control group, the auricle microcirculation blood flow of mice in the bulk drug alcohol extract group, the bulk drug extract medium and high dose group is obviously increased, and the difference has statistical significance (P is less than 0.05, and P is less than 0.01). Compared with the group of the bulk drug alcohol extracts, the mouse auricle microcirculation blood flow of the bulk drug extract high-dose group is obviously increased, and the difference has statistical significance (P is less than 0.05).

TABLE 2 comparison of the microcirculation blood flow of mouse auricles

n＝10)

Compared with the normal control group, ^* P＜0.05， ^** p is less than 0.01; compared with the alcohol extract group of the raw material medicine, ^# P＜0.05。

example 8

The traditional Chinese medicine composition is prepared into effervescent granules, the effervescent granules can be quickly dissolved in water and can be uniformly dispersed in water, and the effervescent granules have the characteristics of convenient taking, quick response, high bioavailability and the like, have good mouthfeel, have the medicinal experience like drinking beverages, are easily accepted by patients, are suitable for people of different ages, and are particularly suitable for the old, children and patients with difficulty in swallowing solid preparations.

A preparation method of a traditional Chinese medicine composition for improving asthenopia comprises effervescent granules, and the preparation method of the effervescent granules comprises the following steps:

b1: because sodium bicarbonate is easily decomposed by heat at high temperature, and the effervescence effect is influenced, the PEG-6000 is heated and melted, and the sodium bicarbonate is added when the temperature is controlled to be 61-70 ℃, so that the decomposition of the sodium bicarbonate is reduced. In the embodiment, PEG-6000 is taken, heated and melted to 70 ℃, added with sodium bicarbonate, stirred evenly, cooled, crushed into fine powder, and sieved by a 80-mesh sieve to obtain sodium bicarbonate coated with PEG-6000; the mass ratio of PEG-6000 to sodium bicarbonate is 0.75: 1; in this example, the mass of PEG-6000 was 161.83g, and the mass of sodium bicarbonate was 215.77 g.

B2: taking 15 parts of the raw material medicine extract (enough raw material medicine extract is obtained by repeating experiments) in example 1, 15 parts of diluent D-mannitol and 70 parts of effervescent disintegrant; effervescent disintegrants include: citric acid, sodium bicarbonate coated with PEG-6000; the acid-base mass ratio of the citric acid to the sodium bicarbonate is 1.5:1, when the citric acid with crystal water is used, the crystal water in the citric acid is not counted when the acid-base mass ratio is calculated. In the embodiment, the mass of the raw material medicine extract is 150g, the mass of the diluent D-mannitol is 150g, and the mass of the effervescent disintegrant is 700 g; in the effervescent disintegrant, the mass of citric acid was 322.4g, and the mass of sodium bicarbonate coated with PEG-6000 was 377.6 g.

b4: mixing one part of the crude drug extract obtained in step B3 with diluent, adding citric acid, mixing, adding 85% ethanol as wetting agent, wet granulating, drying at 40 deg.C, and making into acid granule; mixing the other part of the extract of the raw material medicine obtained in the step B3 with diluent, adding sodium bicarbonate wrapped with PEG-6000, mixing, adding 85% ethanol as wetting agent, wet granulating, drying at 40 deg.C, and making into alkali granule;

b5: and D, uniformly mixing the acid granules and the alkali granules in the step B4, and finishing granules to obtain the effervescent granules. The shape of the effervescent granule of the Chinese medicinal composition is shown in figure 1, and is a dark yellow effervescent granule.

The invention adopts an acid-base separate granulation method, and the PEG-6000 is used for wrapping the sodium bicarbonate, thereby effectively isolating an acid source and an alkali source and ensuring the stability of the preparation. In view of the fact that the raw material medicine extract is large in viscosity and easy to absorb moisture, in order to reduce the moisture absorption of the product, the raw material medicine extract and the diluent in the step B3 are mixed uniformly, and then are respectively prepared into acid granules and alkali granules, and finally the prepared effervescent granules are excellent in moisture absorption resistance.

Example 9

b1: heating and melting PEG-6000 to 61 deg.C, adding sodium bicarbonate, stirring, cooling, and pulverizing into fine powder to obtain PEG-6000-coated sodium bicarbonate; the mass ratio of PEG-6000 to sodium bicarbonate is 0.65: 1;

b2: taking 10 parts of the raw material medicine extract, 10 parts of dextrin serving as a diluent and 50 parts of an effervescent disintegrant in the embodiment 1 by mass; effervescent disintegrants include: citric acid, sodium bicarbonate coated with PEG-6000; the mass ratio of the citric acid to the sodium bicarbonate is 1.3: 1;

b4: mixing one part of the crude drug extract obtained in step B3 with diluent, adding citric acid, mixing, adding 75% ethanol as wetting agent, wet granulating, drying at 40 deg.C, and making into acid granule; mixing the other part of the extract of the raw material medicine obtained in the step B3 with diluent, adding sodium bicarbonate wrapped with PEG-6000, mixing, adding 75% ethanol as wetting agent, wet granulating, drying at 40 deg.C, and making into alkali granule;

Example 10

b1: heating and melting PEG-6000 to 65 deg.C, adding sodium bicarbonate, stirring, cooling, and pulverizing into fine powder to obtain PEG-6000-coated sodium bicarbonate; the mass ratio of PEG-6000 to sodium bicarbonate is 0.85: 1;

b2: taking 20 parts of the raw material drug extract, 20 parts of diluent lactose and 62 parts of effervescent disintegrant in the example 1 according to the parts by weight; effervescent disintegrants include: citric acid, sodium bicarbonate coated with PEG-6000; the mass ratio of the citric acid to the sodium bicarbonate is 1.38: 1;

b3: b2, uniformly mixing the raw material medicine extract and the diluent, and dividing into two equal parts to obtain a mixture of two parts of raw material medicine extracts and the diluent;

b4: mixing one part of the crude drug extract obtained in step B3 with diluent, adding citric acid, mixing, adding 95% ethanol as wetting agent, wet granulating, drying at 40 deg.C, and making into acid granule; mixing the other part of the extract of the raw material medicine obtained in the step B3 with diluent, adding sodium bicarbonate wrapped with PEG-6000, mixing, adding 95% ethanol as wetting agent, wet granulating, drying at 40 deg.C, and making into alkali granule;

Example 11

b1: heating and melting PEG-6000 to 65 deg.C, adding sodium bicarbonate, stirring, cooling, and pulverizing into fine powder to obtain PEG-6000-coated sodium bicarbonate; the mass ratio of PEG-6000 to sodium bicarbonate is 0.5: 1;

b2: according to the mass parts, 5 parts of the raw material medicine extract, 2.5 parts of diluent lactose, 2.5 parts of diluent dextrin and 50 parts of effervescent disintegrant in the embodiment 1 are taken; effervescent disintegrants include: citric acid, sodium bicarbonate coated with PEG-6000; the mass ratio of the citric acid to the sodium bicarbonate is 1.0: 1;

Example 12

b1: heating and melting PEG-6000 to 65 deg.C, adding sodium bicarbonate, stirring, cooling, and pulverizing into fine powder to obtain PEG-6000-coated sodium bicarbonate; the mass ratio of PEG-6000 to sodium bicarbonate is 1: 1;

b2: taking 25 parts of the raw material medicine extract, 10 parts of diluent lactose, 15 parts of diluent dextrin and 70 parts of effervescent disintegrant in the embodiment 1 according to the parts by weight; effervescent disintegrants include: citric acid, sodium bicarbonate coated with PEG-6000; the mass ratio of the citric acid to the sodium bicarbonate is 1.7: 1;

b4: mixing one part of the crude drug extract obtained in step B3 with diluent, adding citric acid, mixing, adding 95% ethanol as wetting agent, wet granulating, drying at 40 deg.C, and making into acid granule; mixing the other part of the extract of the raw material medicine obtained in the step B3 with a diluent, adding sodium bicarbonate wrapped by PEG-6000, mixing, adding 95% ethanol as a wetting agent, granulating by a wet method, and drying at 40 ℃ to obtain alkali granules;

Example 13

1. Subject selection

24 asthenopia patients were selected, 15 men and 9 women, and the age was 22-48 years. The eye symptoms are: the vision can not be seen for a long time, the vision can be blurred for a short time, and the vision is dry and astringent, scorching, foreign body sensation, soreness and distending pain, photophobia and lacrimation. The following subjects were excluded: patients with infectious, traumatic, or other ocular disorders; patients with cardiovascular and cerebrovascular diseases, liver, kidney, hemopoietic system, etc.; taking the medicine or health product related to the tested function to influence the result judgment.

2 method of treatment

By adopting the preparation method of the traditional Chinese medicine composition in the embodiment 8, sufficient effervescent granules are repeatedly prepared, specifically, 800g of chrysanthemum morifolium ramat and 400g of safflower are used together, and 200 bags of effervescent granules are prepared. One package is taken in the morning and at night, and 7 days are a treatment course.

3 Observation index and therapeutic effect standard

The percentage of photopic persistence and symptom score were used as observation indicators. Symptoms are: non-durable vision, temporary blurred vision, dry eyes, burning sensation, lacrimation, itching and distending pain of eyes, dizziness and headache, soreness and weakness of waist and knees, insomnia and dreaminess and amnesia, and scoring is carried out according to the standard of no (0 point), slight (1 point), more obvious (2 points) and very obvious (3 points).

Referring to the standard of curative effect for diagnosis of disease in traditional Chinese medicine (standard docket number 94-0080), the curative effect index is [ (integral of symptoms before treatment-integral of symptoms after treatment)/integral of symptoms before treatment ] × 100%. And (3) curing: the curative effect index is more than or equal to 90 percent. The effect is shown: the curative effect index is 60-89%. The method has the following advantages: the curative effect index is 30-59%. And (4) invalidation: the curative effect index is less than 30%. The total effective rate is the cure rate + the significant efficiency + the effective rate. It is effective to increase the average photopic vision persistence by more than or equal to 10%.

4 therapeutic effects

The effect of the traditional Chinese medicine composition for improving asthenopia symptoms is shown in table 3:

TABLE 3 Effect of the Chinese medicinal composition of the present invention on improving asthenopia symptom

The result shows that the total effective rate of the traditional Chinese medicine composition for improving the asthenopia symptom is 95.8%, and the effect is obvious.

The effect of the traditional Chinese medicine composition for improving the photopic vision persistence percentage is shown in table 4:

TABLE 4 the effect of the Chinese medicinal composition of the present invention on improving the percent of photopic vision persistence (x + -s)

The result shows that the traditional Chinese medicine composition improves the photopic vision persistence by 19.34 percent, has obvious effect and has popularization and application values.

Typical cases

Patient 1, female, 31 years old, building design practitioner. The user can watch the computer for drawing pictures and the mobile phone for a long time, and often overtime, stay up to night and lack of sports. The subjective symptoms of non-durable vision, lacrimation, itching and distending pain of eyes and blurred vision often appear. After the traditional Chinese medicine composition is continuously taken for 1 course, the asthenopia symptom is relieved, the asthenopia symptom disappears after the traditional Chinese medicine composition is taken for 2 courses, and the curative effect is consolidated after the traditional Chinese medicine composition is continuously taken for 1 course.

Patient 2, male, age 27, student. After the traditional Chinese medicine composition is taken for 1 treatment course, the asthenopia symptoms disappear, and the curative effect is consolidated after the traditional Chinese medicine composition is continuously taken for 1 treatment course. No recurrence occurred in the follow-up month of 3.

Comparative example 1

The establishment of a method for measuring the content of total flavonoids containing rutin (C) ₂₇ H ₃₀ O ₁₆ ) And (6) counting.

1. Preparation of rutin control solution: rutin (purity: 98.00%, Kyormant Biotechnology Co., Ltd., batch number: MUST-19010202) reference substance 1.18mg is accurately weighed, placed in a 10mL volumetric flask, dissolved by 70% ethanol and diluted to scale, and the rutin reference substance solution with the concentration of 0.118mg/mL is obtained.

2. Preparation of a test solution: weighing 0.6g (0.4 g of chrysanthemum morifolium and 0.2g of safflower) of medicinal powder of a prescription screened by a 10-mesh sieve, precisely weighing, placing in a round-bottom flask, adding 50mL of 70% ethanol, precisely weighing, performing reflux extraction at 80 ℃ for 1 time, performing extraction for 1h, cooling, precisely weighing, supplementing the loss weight with 70% ethanol, shaking, filtering, and taking the filtrate.

3. Preparation of an analytical sample: taking a proper amount of rutin control solution or test solution, placing in a 10mL volumetric flask, adding 2mL of 10% aluminum trichloride solution and 3mL of 0.1mol/L potassium acetate solution, adding 70% ethanol to 10mL, shaking up, and standing for 15min to prepare an analysis sample.

4. Determination of detection wavelength: and respectively transferring a test sample solution and a rutin reference substance solution in proper amount to prepare an analysis sample according to the item 3 of the comparative example. Scanning was performed at a wavelength of 200-800nm with a UV-2450 UV-visible spectrophotometer (Shimadzu, Japan) without adding any solution as a blank, and a spectrum was recorded. The results show that: the maximum absorption wavelength of the test solution analysis sample and the rutin control solution analysis sample is determined to be 403nm because the maximum absorption is detected to be at 403 nm. The results are shown in FIG. 2.

5. Establishing a rutin standard curve: precisely measuring rutin control solutions 0mL, 0.5mL, 1.0mL, 1.5mL, 2.0mL and 3mL, respectively preparing rutin control solution analysis samples with concentrations of 0 μ g/mL, 5.90 μ g/mL, 11.80 μ g/mL, 17.70 μ g/mL, 23.60 μ g/mL and 35.40 μ g/mL according to the item "3" of the comparative example, and taking the rutin control solution analysis sample with concentration of 0 μ g/mL as a blank control. Measuring absorbance of each rutin control solution analysis sample at a wavelength of 403nm, performing linear regression with rutin concentration (μ g/mL) as abscissa and absorbance A as ordinate, and obtaining a regression equation: y ═ 0.0278X +0.0001, r ═ 0.9997, indicating that the rutin control solution is in good linear relationship in the range of 5.90 μ g/mL to 35.40 μ g/mL.

6. And (3) precision test: precisely measuring 3.0mL of rutin control solution, preparing analysis sample according to the item "3" of the comparative example, measuring absorbance at 403nm, and continuously measuring for 6 times. As a result, RSD of the absorbance was 0.08%.

7. And (3) stability test: 0.25mL of the test sample solution was precisely measured, and an analytical sample was prepared in the same manner as in the comparative example No. 3, and absorbance was measured at a wavelength of 403nm and every 10 min. The RSD of the absorbance of the test solution in 2h is 1.20 percent, and the result shows that the stability of the test solution in 2h is good after the color development.

8. And (3) repeatability test: 0.25mL of test solution is precisely measured, an analysis sample is prepared according to the item 3 of the comparative example, the absorbance is measured at the wavelength of 403nm, and the RSD is 2.07 percent, which indicates that the method has better repeatability.

The total flavone content in the present invention was measured by the above analytical method, and is applicable to all examples or comparative examples unless otherwise specified. And substituting the detected absorbance of the analysis solution into a regression equation to obtain the concentration of the total flavonoids in the analysis solution. And obtaining the concentration of the total flavone in the sample according to the concentration or dilution times of the analysis solution during treatment.

The invention adopts static and dynamic adsorption experiments on six kinds of macroporous adsorption resin, and takes the content of total flavone as an index, and the total flavone takes rutin (C) ₂₇ H ₃₀ O ₁₆ ) And (4) observing the adsorption and desorption capacity of different macroporous adsorption resins on the main components of the prescription.

Pretreatment of macroporous adsorption resin: six kinds of macroporous adsorption resin S-8, H1020, ADS-7, LS206, HPD600 and LSA-10 are weighed and soaked in ethanol of about 2BV for 24 hr. And (3) putting the soaked macroporous resin on a column by a wet method, flushing with 2.5-5BV of ethanol at the flow rate of 2BV/h until the effluent liquid is not white and turbid after being mixed with distilled water, and finally flushing with distilled water at the flow rate of 2BV/h until no alcohol smell exists for later use.

Sample loading liquid: the alcohol-free extract solution of example 1 was prepared to have a drug concentration of about 0.2g crude drug/mL.

Analysis solution before adsorption: 3.0mL of the sample liquid was precisely measured, and an analytical sample was prepared according to the item "3" of this comparative example.

Analysis solution after adsorption: precisely measuring 3.0mL of effluent liquid of the sample loading liquid after being adsorbed by macroporous adsorption resin, and preparing an analysis sample according to the item 3 of the comparative example.

Eluent analysis solution: 3.0mL of the eluate was precisely measured and analyzed according to the item "3" of this comparative example.

Static test: specific adsorption amount and adsorption rate measurement: weighing 4g of each of the 6 pretreated macroporous adsorption resins, placing the weighed materials into a 100mL conical flask with a plug, adding 20mL of sample liquid, placing the sample liquid into a constant-temperature oscillator (100r/min, 37 ℃) to oscillate for 24 hours, filtering, respectively measuring the content of total flavonoids in effluent liquid of the sample liquid and the sample liquid adsorbed by the macroporous adsorption resins, and calculating specific adsorption capacity and adsorption rate, wherein the calculation formula is as follows:

specific adsorption capacity m ₁ (mg/g)＝(C ₀ ×V ₀ -C ₁ ×V ₁ ) (ii) a/M; adsorption rate (%) - (C) ₀ ×V ₀ -C ₁ ×V ₁ )/(C ₀ ×V ₀ )

In the formula, C ₀ : the concentration of total flavonoids before adsorption; c ₁ : the concentration of the adsorbed total flavonoids; v ₀ : volume of solution before adsorption, V ₁ : volume of solution after adsorption; m: weight of macroporous resin

Determination of specific elution amount and elution rate: taking the adsorbed resin, carrying out suction filtration to dry, placing the resin in a 100mL conical flask, precisely adding 20mL of 60% ethanol, placing the resin in a constant temperature oscillator (100r/min, 37 ℃) for oscillation for 24h, filtering, measuring the content of total flavonoids in eluent, and calculating the specific elution amount and the elution rate according to the following formula:

specific elution amount m ₂ (mg/g)＝(C ₂ ×V ₂ ) (ii) a/M; (iv) elution rate (%) - (C) ₂ ×V ₂ )/(C ₀ ×V ₀ -C ₁ ×V ₁ )。

In the formula, C ₂ : the concentration of the total flavone after desorption; v ₂ : volume of solution after desorption.

The results are shown in Table 5.

TABLE 5 static test results of Total Flavonoids: (

n＝3)

The results show that the HPD600 and LSA-10 macroporous adsorption resins have the best elution capacity but poor adsorption capacity, the H1020 and LS206 macroporous adsorption resins have the better adsorption capacity and elution capacity for the total flavone, S-8 and ADS-7 with the worst elution capacity are eliminated comprehensively, and the remaining HPD600, H1020, LSA-10 and LS206 macroporous adsorption resins are selected for the next dynamic test.

Dynamic test: weighing 10g of each of four macroporous resins including HPD600, H1020, LSA-10 and LS206 screened by static tests, filling the macroporous resins into a column by a wet method, adding 20mL (about 1BV) of sample loading liquid at the speed of 0.7mL/min (about 2BV/H), firstly washing the macroporous resins by 100mL (about 5BV) of ultrapure water at the speed of 0.7mL/min (about 2BV/H), then eluting the macroporous resins by 100mL (about 5BV) of 60% ethanol at the speed of 0.7mL/min (about 2BV/H), collecting eluent, and calculating specific adsorption capacity, adsorption rate, specific elution capacity and elution rate. The results are shown in Table 6.

TABLE 6 dynamic test results of Total Flavonoids: (

n＝3)

The result shows that the LS206 type macroporous resin has better adsorption and elution performance.

Comparative example 2

Weighing 10g of LS206 macroporous resin, loading the LS206 macroporous resin into a column by a wet method, adjusting the pH of the sample solution in the example 1 to 2-3, 4-5, 6-7 and 8-9 by using 2mol/LHCl and 2mol/LNaOH, respectively loading 20mL (about 1BV) of sample solution at the speed of 0.7mL/min (about 2BV/h), washing by using 100mL (about 5BV) of ultrapure water at the speed of 0.7mL/min (about 2BV/h), eluting by using 100mL (about 5BV) of 60% ethanol at the speed of 0.7mL/min (about 2BV/h), collecting eluent, and calculating the specific adsorption capacity, adsorption rate, specific elution capacity and elution rate. The results are shown in Table 7.

TABLE 7 influence of pH of the loading solution on the adsorption and elution performance of macroporous adsorbent resins ((

n＝3)

From the results, it was found that the adsorption capacity and elution capacity of LS206 macroporous resin to total flavonoids were sufficient when the pH of the sample solution was 4 to 5, and it was preferable to purify the sample solution without adjusting the pH because the pH of the alcohol-free extract solution itself was between 4 and 5. Simple steps and good purification effect.

Comparative example 3

LS206 macroporous adsorbent resin was subjected to wet column packing (diameter/height ratio 1:3, 3 cm. times.9 cm), the solutions (pH 4.90, pH was not adjusted; total flavone concentration 11.69mg/mL) obtained in example 1 were subjected to each of 120mL (about 2BV) at 1BV/h, 2BV/h and 3BV/h, and 10 mL/portion, and the total flavone content in each portion was measured, as shown in Table 8.

Table 8 results of investigation of sample loading amount and sample loading speed (

n＝3)

When the leakage amount of the total flavonoids in the effluent liquid reaches 10% of the loading concentration, the adsorption end point is regarded. According to the results, when the sample loading speed is 1BV/h, the total flavone obviously leaks from the 12 th effluent; when the sample loading speed is 2BV/h and 3BV/h, the total flavone content in the effluent of the 9 th part exceeds 10 percent of the sample loading concentration, and obvious leakage occurs. In view of efficiency and time cost, it is preferable that the loading amount is 80mL (about 1.3BV) and the loading speed is 2 BV/h.

Comparative example 4

Loading LS206 macroporous adsorption resin into a column according to a diameter-height ratio wet method of 1:3(3cm x 9cm), 1:5(3cm x 15cm) and 1:7(3cm x 15cm), loading 80mL, 133mL and 187mL (about 1.3BV) of sample loading liquid in example 1 at a speed of 2BV/h respectively, then removing impurities by using 126mL, 212mL and 296mL (about 2BV) of ultrapure water at a speed of 2BV/h, eluting by using 444mL, 318mL and 189mL (about 3BV) of 60% ethanol solution as an eluent at a speed of 2BV/h, collecting eluent, measuring the content of total flavonoids, performing vacuum drying to obtain extract, and calculating the yield and purity of the total flavonoids. The results are shown in Table 9.

The calculation formula of the extract yield is as follows: the mass of the extract/the total mass of the raw material medicaments is 100 percent.

TABLE 9 investigation results of aspect ratio: (

n＝3)

The results show that the transfer rate of total flavonoids is more than 80% when the ratio of diameter to height is 1:3 to 1:7, and the average purity of total flavonoids obtained is the highest when the ratio of diameter to height is 1: 5.

Comparative example 5

At present, pure water or low-concentration ethanol solution is generally adopted for impurity removal, and a new method is developed for impurity removal by using NaCl solution and compared with the impurity removal effect of pure water or low-concentration ethanol or other impurity removal solvents. The LS206 macroporous resin is loaded on a column by a wet method with the diameter-height ratio of 1:5(3cm x 15cm), 133mL (about 1.3BV) of sample loading liquid is loaded at the speed of 2BV/h, 212mL (about 2BV) of ultrapure water, 10% ethanol, 0.1mol/L LNaCl solution, 0.1mol/L HCl solution and 0.1mol/L NaOH solution are respectively used for impurity removal at the speed of 2BV/h, impurity removal effluent liquid is collected, the content of total flavone is measured, 50mL of impurity removal effluent liquid is taken for vacuum drying, and the weight of condensate is calculated. The results are shown in Table 10.

The total flavone loss rate (%) (total flavone amount in impurity-removed effluent/total flavone amount in sample-removed effluent) was 100%.

The impurity content (%) in the impurity-removed condensate was (total condensate weight-total flavone amount in condensate)/total condensate weight was 100%.

TABLE 10 loss rates of total flavonoids and the weight of the cured product in the presence of various solvents (

n＝3)

According to results, compared with other impurity removal solutions, when the impurity removal is carried out by using 0.1mol/L NaCl solution and 0.1mol/L HCl solution, the total flavone loss rate is obviously lower, but the HCl solution is more complex to process in the actual industrial production and can cause certain influence on the environment, the NaCl solution is used for impurity removal, the problems of complex post-treatment, environmental pollution and the like are solved, and the three advantages of low total flavone loss rate, more impurity removal total condensate and high impurity content in the impurity removal condensate in the impurity removal process are realized.

Comparative example 6

By using a controlled variable method, the experimental conditions of example 1 are taken as basic conditions (if no special description is made, the experimental conditions except the variables are the same as those of example 1), the LS206 macroporous resin is loaded on a column by a 1:5(3cm by 15cm) diameter-height ratio wet method, 133mL (about 1.3BV) of loading liquid is loaded at the speed of 2BV/h, 0.1mol/L NaCl solution is taken as an impurity removal solvent, the effluent is collected every 20 mL/part, the effluent is weighed after vacuum drying, the weight of a solidified substance is obtained, and the content of total flavonoids in the solidified substance is measured. The results are shown in Table 11.

TABLE 11 weight of condensate of impurity-removed effluent and total flavone content in condensate

n＝3)

From the results, it is found that when the amount of NaCl solution is 100mL, the weight gain of the solidified material is small, the total flavone content in the solidified material is higher and higher, and the increase of the occupied ratio is large, therefore, 100mL (about 0.9BV) of NaCl solution 0.1mol/L is preferably used as the impurity removal solvent.

Comparative example 7

The treated LS206 macroporous resin is loaded into a column by a 1:5(3cm by 15cm) diameter-height ratio wet method, 133mL (about 1.3BV) of sample loading liquid is loaded at the speed of 2BV/h, 100mL (about 0.9BV) of 0.1mol/L NaCl solution is used for removing impurities at the speed of 2BV/h, 60% ethanol is used for eluting at the speed of 2BV/h, and eluent is collected by 5 methods in sections. The specific operation mode is as follows:

group 1: all the materials are collected;

group 2: the first 20mL (about 0.2BV) was discarded before collection;

group 3: the first 40mL (about 0.4BV) was discarded and collected;

group 4: the first 50mL (about 0.5BV) was discarded and collected;

group 5: the first 60mL (about 0.6BV) was discarded and collected;

collecting the eluents, vacuum drying to obtain dry extract, measuring the content of total flavone, and calculating the transfer rate and purity of total flavone. The results are shown in Table 12.

TABLE 12 influence of different collection modes of the eluate on the purification parameters of Total Flavonoids ((

n＝3)

As is clear from the results, the transfer rate of total flavonoids decreased and the purity increased with the increase of the amount of the discarded eluate, and the transfer rate of all 5 groups was more than 80%, and the purity was more than 60% after discarding 60mL, so that it is preferable to discard the first 60mL of the eluate.

Comparative example 8

The diluent for the effervescent granules is generally selected to be water soluble and includes: dextrin, lactose, D-mannitol; the effervescent granule is easy to react and lose the effervescence when being wetted, so a diluent with strong moisture absorption resistance is selected.

The experimental conditions of example 8 were used as basic conditions by a controlled variation method (unless otherwise specified, the experimental conditions except for the variables were the same as those of example 8), and three kinds of excipients, lactose, D-mannitol, and dextrin were used as indexes for examination, and the results are shown in Table 13.

TABLE 13 selection results for diluent types

As a result, it was found that dextrin gives a sticky soft material, and lactose and D-mannitol have satisfactory properties without much difference in granulation easiness, clarity and effervescence time, but D-mannitol also acts as a taste-masking agent, and D-mannitol is preferred as a diluent.

Comparative example 9

The fixed acid-base ratio is 1.3:1, PEG-6000: NaHCO 2 ₃ The effect of the amount of disintegrant on effervescent granules was examined using the dissolution time, pH and granule forming ratio as indices under the same conditions as in example 8 except that D-mannitol was used as the diluent and 85% ethanol was used as the wetting agent, and the results are shown in Table 14.

TABLE 14 examination of disintegrant dosage

From the results, it is understood that the dissolution time is related to the amount of the disintegrant, and the larger the amount, the more rapid the dissolution, the less the change in the pH value, and the smaller the change in the granule molding rate.

Comparative example 10

Using the controlled variable method, the experimental conditions of example 8 were used as basic conditions (unless otherwise specified, the experimental conditions except for the variables were the same as in example 8), and granulation was performed at different ratios (PEG: base), and the results of examination were shown in Table 15 using inclusion state, dissolution time, pH and granule formation rate as indices.

TABLE 15 investigation of PEG-6000 and sodium bicarbonate ratio

From the results, it is known that when the ratio of PEG-6000 to sodium bicarbonate is 0.5:1, the sodium bicarbonate is too large to be uniformly mixed in a short time, and the hardness of the dry paste is large, increasing the difficulty of pulverization; when the ratio of PEG-6000 to sodium bicarbonate is 0.75:1 and 1:1, the inclusion condition is better, the hygroscopicity of PEG-6000 is stronger, namely the smaller the dosage is, the better the dosage is, meanwhile, PEG pharmaceutic adjuvants may have similar laxative effect, and the use of too high concentration is not suitable. Therefore, the ratio of PEG-6000 to sodium bicarbonate is preferably 0.75: 1.

Comparative example 11

Using the controlled variable method, the experimental conditions of example 8 were used as basic conditions (unless otherwise specified, the experimental conditions except for the variables were the same as in example 8), and granulation was performed at different acid-base ratios, and the results were examined using the dissolution time and pH as indices, and are shown in Table 16.

TABLE 16 examination of the acid-base ratio

From the results, it is understood that when the acid-base ratio is 1.5:1, the dissolution time is shortest, the molar ratio of acid-base which dissolves most quickly is 0.6:1, and the ratio is preferably 1.5:1 in terms of mass ratio.

Comparative example 12

The results of examination by the controlled variable method using the experimental conditions of example 8 as the basic conditions (unless otherwise specified, the experimental conditions except for the variables were the same as in example 8), changing the concentration of the wetting agent in the ethanol solution, and using the melting time and the granule forming ratio as indices, are shown in Table 17.

TABLE 17 examination of wetting agents

The commonly used wetting agent is water, and ethanol is selected as the wetting agent due to the fact that the extract of the raw material medicine is high in viscosity and easy to absorb moisture. Whereas the lower the concentration of wetting agent, i.e. the higher the water content, the lower the granule formation rate, the moisture will induce hygroscopicity of the extract, so that the mixture becomes viscous to the detriment of granulation. As a result, it was found that when 85% ethanol was used as the wetting agent, the melting time was short and the molding rate was high.

The foregoing has outlined rather broadly the preferred embodiments and principles of the present invention and it will be appreciated that those skilled in the art may devise variations of the present invention that are within the spirit and scope of the appended claims.

Claims

1. The traditional Chinese medicine composition for improving the asthenopia is characterized by comprising the following raw material medicines in parts by mass: 16-24 parts of Hangzhou white chrysanthemum and 8-12 parts of safflower;

the preparation of the traditional Chinese medicine composition comprises effervescent granules;

the effervescent granule comprises the following components in parts by weight: 5-25 parts of crude drug extract, 5-25 parts of diluent and 50-70 parts of effervescent disintegrant;

the diluent comprises: at least one of dextrin, lactose, and D-mannitol;

the effervescent disintegrant comprises: citric acid, sodium bicarbonate coated with PEG-6000; in the sodium bicarbonate coated with the PEG-6000, the mass ratio of the PEG-6000 to the sodium bicarbonate is (0.75-1) to 1; the mass ratio of the citric acid to the sodium bicarbonate is (1.0-1.7) to 1;

the raw material medicine extract is prepared by the following steps:

a2: loading macroporous adsorption resin into a column, then sampling alcohol-free extract on the macroporous adsorption resin column, removing impurities from the macroporous adsorption resin column by using NaCl solution, then eluting by using ethanol to obtain eluent, and drying the eluent to obtain a crude drug extract; the macroporous adsorption resin is at least one of H1020, LS206, HPD600 and LSA-10.

2. The traditional Chinese medicine composition for improving asthenopia according to claim 1, which is prepared from the following raw materials in parts by mass: 19-21 parts of Hangzhou white chrysanthemum and 9.5-10.5 parts of safflower;

the diluent comprises: at least one of dextrin, lactose, and D-mannitol;

the raw material medicine extract is prepared by the following steps:

a1: taking 19-21 parts by weight of chrysanthemum morifolium and 9.5-10.5 parts by weight of safflower, heating and refluxing the chrysanthemum morifolium and the safflower by using ethanol to obtain an ethanol extract, and removing the ethanol from the ethanol extract to obtain an ethanol-free extract;

3. A preparation method of a crude drug extract for improving asthenopia is characterized by comprising the following steps:

s2: loading macroporous adsorption resin into a column by a wet method according to the diameter-height ratio of 1: 3-7, loading 0.5BV-2BV of alcohol-free extract on the macroporous adsorption resin column at the flow rate of 1BV/h-3BV/h, removing impurities from the macroporous adsorption resin column by 0.72BV-1.26BV of NaCl solution with the concentration of 0.05mol/L-0.2mol/L at the flow rate of 1BV/h-3BV/h, eluting by 1.8BV-2.7BV of 40-80% ethanol at the flow rate of 1BV/h-3BV/h, discarding the former 0.5BV-0.6BV, collecting the rest of effluent as eluent, and performing vacuum drying on the eluent to obtain a crude drug extract; the macroporous adsorption resin is at least one of H1020, LS206, HPD600 and LSA-10.

4. The preparation method of the traditional Chinese medicine composition for improving asthenopia according to claim 1, wherein the preparation method of the effervescent granule comprises the following steps:

b1: heating and melting PEG-6000, adding sodium bicarbonate, stirring, cooling, and pulverizing into fine powder to obtain PEG-6000-coated sodium bicarbonate; the mass ratio of the PEG-6000 to the sodium bicarbonate is (0.75-1) to 1;

b2: taking 5-25 parts of raw material medicine extract, 5-25 parts of diluent and 50-70 parts of effervescent disintegrant according to the mass parts; the effervescent disintegrant comprises: citric acid, sodium bicarbonate coated with PEG-6000; the mass ratio of the citric acid to the sodium bicarbonate is (1.0-1.7) to 1;

5. The preparation method of the traditional Chinese medicine composition for improving asthenopia according to claim 4, wherein the preparation method of the effervescent granule comprises the following steps:

b1: heating and melting PEG-6000 to 61-70 deg.C, adding sodium bicarbonate, stirring, cooling, and pulverizing into fine powder to obtain PEG-6000-coated sodium bicarbonate; the mass ratio of the PEG-6000 to the sodium bicarbonate is (0.75-0.85) to 1;

b2: taking 10-20 parts of raw material medicine extract, 10-20 parts of diluent and 50-70 parts of effervescent disintegrant according to parts by weight; the effervescent disintegrant comprises: citric acid, sodium bicarbonate coated with PEG-6000; the mass ratio of the citric acid to the sodium bicarbonate is (1.3-1.5) to 1;

6. The preparation method of the traditional Chinese medicine composition for improving asthenopia according to claim 5, which is characterized by comprising the following steps:

the diluent is D-mannitol; the wetting agent is 75-95% ethanol.

7. Use of the traditional Chinese medicine composition according to claims 1-2, or the crude drug extract prepared by the method according to claim 3, or the traditional Chinese medicine composition prepared by the method according to any one of claims 4-6, for preparing a medicament for improving asthenopia.