Chitosan microspheres, a preparation method thereof and application of the chitosan microspheres in the preparation of Baoyuan oral liquid

Abstract

The application relates to the field of traditional Chinese medicine oral liquid preparations, in particular to a chitosan microsphere and a preparation method thereof and application of the chitosan microsphere in preparation of Baoyuan oral liquid preparations. The chitosan microsphere comprises, in weight parts, 80-120 parts of chitosan, 2-5 parts of an emulsifier and 30-40 parts of an oil phase; the chitosan microsphere is obtained by dissolving in an acid solution and then reacting with the emulsifier and the oil phase. The application adds a clarifying agent, i.e. a mixture of chitosan microspheres with different particle sizes, carries out flocculation adsorption, and adds a stabilizer sodium carboxymethyl cellulose CMC-Na; through the synergistic effect of the two, the quality and safety of the oral liquid are remarkably improved. The colloids after adsorption of the clarifying agent can be more stably dissolved in the medicinal liquid under the synergistic effect of the stabilizer sodium carboxymethyl cellulose, the preparation precipitation problem is solved, and the prepared Baoyuan oral liquid has good stability and immunoregulatory activity.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of traditional Chinese medicine oral liquid preparation, and particularly relates to a chitosan microsphere, a preparation method thereof and application of the chitosan microsphere in Baoyuan oral liquid preparation. BACKGROUND

[0002] The immune system is the most effective weapon to defend against pathogen invasion, but due to individual genetic and disease effects, the patient's own immune capacity varies greatly. Baoyuan decoction, as a famous qi-tonifying prescription, plays an important role in supplementing the body's vital qi and thus improving the body's immune function.

[0003] The composition of Baoyuan decoction in the Ming Dynasty's Wei Zhi's Boai Xinjian is: ginseng, astragalus, licorice, cassia, plus a piece of ginger, decocted with water, and taken at any time. It is used to treat deficiency and weakness, and insufficient vital qi. Symptoms include fatigue, lack of energy, shortness of breath, aversion to cold, as well as infantile smallpox, yang deficiency, blood deficiency, and difficulty in convergence. The literature research shows that Baoyuan decoction can increase the cell-mediated immune function of infants after atrial septal defect repair, and significantly increase the T lymphocyte colony count, CD 3+ , CD 4+ / CD 8+ expression rate, and significantly enhance the cell-mediated immunity.

[0004] As a classical prescription in ancient China, Baoyuan decoction is widely used and has definite curative effect. The traditional decoction has been used since ancient times, but the composition of Baoyuan oral liquid is complex. In addition to containing active ingredients, the traditional decoction also contains some high molecular impurity ingredients such as starch, gum, mucilage, tannin and pigment. These ingredients have strong solubility when heated, and gradually precipitate after cooling, or produce precipitates due to changes in temperature, oxidation and pH value. Therefore, the precipitation problem of Baoyuan oral liquid after development and preparation has been a key point that has puzzled the development of the preparation. SUMMARY

[0005] The purpose of the present application is to provide a chitosan microsphere and a preparation method thereof, and application of the chitosan microsphere in the preparation of stable Baoyuan oral liquid preparation, so as to completely retain the efficacy of the classical prescription.

[0006] To achieve the above-mentioned purpose, the technical solutions provided by the present application are as follows.

[0007] A chitosan microsphere, by weight, comprises: chitosan 80-120 parts, emulsifier 2-5 parts, and oil phase 30-40 parts. The chitosan microsphere is obtained by dissolving in an acidic solution and then reacting with the emulsifier and the oil phase.

[0008] In one preferred embodiment, the pH value of the acidic solution is 2-4.

[0009] The cationic and positive chitosan microspheres prepared by the acidic solution can strongly adsorb the anionic suspended substances in the Baoyuan oral liquid.

[0010] In one preferred embodiment, the acidic solution comprises one or both of an acetic acid solution or a phosphoric acid solution.

[0011] In one preferred embodiment, the emulsifier comprises one or both of sodium dodecyl sulfate or Tween.

[0012] In one preferred embodiment, the Tween comprises Tween 80.

[0013] In one preferred embodiment, the oil phase comprises one or more of soybean oil, olive oil or sunflower oil.

[0014] In one preferred embodiment, the chitosan microspheres comprise 10-20 wt% of chitosan microspheres with a particle size less than 0.1 μm, 20-40 wt% of chitosan microspheres with a particle size of 0.1-40 μm and 45-70 wt% of chitosan microspheres with a particle size of 45-105 μm.

[0015] The chitosan microspheres with a particle size of 45-105 μm are used to adsorb and remove macromolecular impurities such as starch and pectin in the medicinal liquid, to improve the clarity of the medicinal liquid, and to be filtered out during the preparation of the oral liquid. The chitosan microspheres with a particle size less than 0.1 μm still exist in the oral liquid after preparation, and are used to maintain the stability of the oral liquid and to effectively retain the effective components in the oral liquid, such as part of the polysaccharides, so that the efficacy of the oral liquid is largely retained. The chitosan microspheres with a particle size of 0.1-40 μm can adsorb phenolic acid and tannin components in the medicinal liquid to improve the stability of the medicinal liquid.

[0016] The preparation method of the chitosan microspheres comprises the following steps: dissolving chitosan in an acidic solution, adding an emulsifier, stirring, then adding an oil phase dropwise, heating the reaction system to 70-90 °C after the dropwise addition is completed, purifying the chitosan microspheres after 30-50 min of reaction; the emulsifier is added dropwise in multiple times, and each dropwise addition is separated from the previous dropwise addition by 20-40 min.

[0017] The reaction temperature of the system can affect the solubility of chitosan and the viscosity of the solution. If the temperature is too high, the viscosity of the chitosan solution will decrease, and if the temperature is too low, the cross-linking may not be complete, affecting the stability and performance of the microspheres.

[0018] In one preferred embodiment, the emulsifier is added in three times, 50-60wt% of the total amount of emulsifier is added in the first time, 30-39wt% of the total amount of emulsifier is added in the second time, and 1-20wt% of the total amount of emulsifier is added in the third time, and the interval between each time is 20-40min.

[0019] The adding process and the amount of each addition of the emulsifier have a significant influence on the particle size and morphology of the microspheres. The multiple times of adding and the amount of the emulsifier ensure the existence of a certain range of particle sizes of the chitosan microspheres in the chitosan microspheres, thereby ensuring the implementation of the function.

[0020] The size of the particle size has an important influence on the application and performance of chitosan. If the particle size is too large, the solubility is reduced, and if the particle size is too small, the adsorption performance is reduced.

[0021] The chitosan microspheres prepared by the method of the present application have stronger adsorption performance than the prior art and a wider pH range. The addition of the chitosan microspheres needs to be able to play an adsorption role and strengthen the retention of active ingredients, and on the other hand, it cannot cause the settlement of Baoyuan oral liquid. The chitosan microspheres synthesized by the prior art for adsorbing tetracycline, Cr or phosphorus elements will obviously cause the settlement of Baoyuan oral liquid and are not suitable for the system of the present application.

[0022] The present application also claims the use of the chitosan microspheres in the preparation of Baoyuan oral liquid preparations.

[0023] Based on the same inventive concept, the present application also claims a preparation method of the Baoyuan oral liquid preparation, comprising the following steps:

[0024] S1, mixing ginseng 140-160 parts, astragalus 300-320 parts, licorice 70-85 parts, cassia bark 28-35 parts, and ginger 110-130 parts, baking, then adding 12-18 times the amount of water, soaking for 40-70min, boiling, then keeping it slightly boiling for 1.2-1.5h, filtering to obtain filtrate 1 and dregs;

[0025] S2, adding 10-14 times the amount of water to the dregs to boil, then keeping it slightly boiling for 35-65min, filtering, combining the filtrates, concentrating, and adjusting the pH value to 4.3-5.8;

[0026] S3, adding 2-25 parts of clarifying agent and 0.2-1.2 parts of stabilizer, mixing uniformly, filtering to obtain the Baoyuan oral liquid preparation;

[0027] The clarifying agent is the chitosan microspheres described above.

[0028] In one preferred embodiment, the filtering is carried out by using a ceramic inorganic membrane.

[0029] In one preferred embodiment, the filtering is carried out by using a ceramic inorganic membrane with a pore size of 0.1-0.45 μm.

[0030] In one preferred embodiment, the concentration is carried out by vacuum concentration at 60-80 °C to 350-450 ml.

[0031] In step S2, the pH value is adjusted to be in the range of 4.3-5.8, and the Baoyuan oral liquid is more stable and has better clarity. If the pH value is too acidic or too alkaline, the stability of the Baoyuan oral liquid becomes poor. As a preferred embodiment of the present application, the concentration of the stabilizer sodium carboxymethyl cellulose CMC-Na is 0.06-0.15%.

[0032] If the amount of the clarifying agent is too much, the sample liquid will be too viscous, affecting the operability and subsequent processing. If the amount of the clarifying agent is too little, the adsorption performance will be poor, and precipitation will occur.

[0033] In one preferred embodiment, the stabilizer is one or both of sodium carboxymethyl cellulose and xanthan gum, and sodium carboxymethyl cellulose is preferred in consideration of the effect of improving texture.

[0034] In one preferred embodiment, the ginseng is grown for 5-8 years.

[0035] In one preferred embodiment, the astragalus is cultivated in a wild-like manner.

[0036] In one preferred embodiment, the cinnamon bark is selected from the bark of a cinnamon tree with a tree age of 10-15 years.

[0037] In one preferred embodiment, the ginger variety is small ginger "Xiaohuangjiang".

[0038] In one preferred embodiment, the Baoyuan oral liquid preparation further comprises a flavoring agent 25-35 parts, which is added after the stabilizer is added.

[0039] In one preferred embodiment, the flavoring agent comprises one or both of steviol glycoside and xylitol.

[0040] In one preferred embodiment, the flavoring agent comprises steviol glycoside 0.1-0.5 parts and xylitol 25-35 parts.

[0041] In one preferred embodiment, the flavoring agent is one or both of steviol glycoside and xylitol.

[0042] Based on the same inventive concept, the present application also claims the Baoyuan oral liquid preparation prepared by the above preparation method.

[0043] The present application has the following beneficial effects:

[0044] (1)The application provides a preparation method of Baoyuan oral liquid, avoids loss of medicinal substances caused by drying process, reduces the dosage of traditional decoction, retains the medicinal substance basis of traditional decoction, and improves the bioavailability of Chinese medicine oral liquid.

[0045] (2)The components of Chinese medicine oral liquid are quite complex, and the oral liquid is a mixture of solution, colloid, suspension and emulsion. The particles in the colloid and suspension are prone to sedimentation and coalescence when standing, and the emulsion is prone to creaming and rupture when the conditions change. Meanwhile, the particles in the colloid, suspension and emulsion have different charges, and thus are attracted to each other, resulting in a large amount of precipitation after standing. Therefore, the application adds the clarifying agent chitosan microspheres for flocculation and adsorption, and the stabilizer sodium carboxymethyl cellulose (CMC-Na). The stabilizer can promote the effect of the clarifying agent, and the clarifying agent can also promote the effect of the stabilizer. The main principle may be that the chitosan microspheres with a particle size of less than 0.1 μm can improve the permeability of the drug, the chitosan microspheres with a particle size of 0.1-40 μm can adsorb phenolic acid and tannin components in the medicinal liquid to improve the stability of the medicinal liquid, and the chitosan microspheres with a particle size of 45-105 μm can adsorb macromolecular impurities such as starch and pectin in the medicinal liquid to improve the clarity of the medicinal liquid. The clarifying agent and the stabilizer have synergistic effects in two aspects of adsorbing impurities and buffering water quality; the clarifying agent mainly removes impurities in the oral liquid through adsorption, and the stabilizer mainly buffers and stabilizes the water quality to balance the environmental factors such as pH, salinity and osmotic pressure in the water. The clarifying agent removes impurities through adsorption, and the stabilizer maintains the stability of the oral liquid by buffering the water quality, and the synergistic effect of the two can significantly improve the quality and safety of the oral liquid. The colloids adsorbed by the clarifying agent can be more stably dissolved in the medicinal liquid under the synergistic effect of the stabilizer sodium carboxymethyl cellulose, thereby solving the problem of preparation precipitation and making the prepared Baoyuan oral liquid have good stability and immunoregulatory activity. BRIEF DESCRIPTION OF DRAWINGS

[0046] Figure 1 The state of the Baoyuan oral liquid prepared in Example 1 after standing at a temperature of 25 and a humidity of 60% for 30 days;

[0047] Figure 2 The state of the Baoyuan oral liquid prepared in Examples 1 and 2 after standing at a temperature of 25 and a humidity of 60% for 30 days;

[0048] Figure 3 The state of the Baoyuan oral liquid prepared in Comparative Example 1 after standing at a temperature of 25 and a humidity of 60% for 30 days;

[0049] Figure 4 The state of the Baoyuan oral liquid prepared in Comparative Example 2 after standing at a temperature of 25 and a humidity of 60% for 30 days;

[0050] Figure 5 The state of the Baoyuan oral liquid prepared for Comparative Example 12 after standing at temperature 25 and humidity 60% for 30 days. DETAILED DESCRIPTION

[0051] The application will be further described in connection with the following specific examples, which are in no way intended to limit the scope of the application. The molecular weight of the chitosan used in the examples and comparative examples is 161.2.

[0052] Example 1

[0053] The chitosan microspheres are prepared as follows: first, 100 g of chitosan is dissolved in an acetic acid solution with pH value of 3.2, keeping the pH value of the system at 3.2-3.3. Then, an emulsifier (sodium dodecyl sulfate) is added, the amount of the emulsifier being 2.5% of the mass of the chitosan, the emulsifier being added dropwise in three times, 50 wt% of the total amount of the emulsifier being added in the first time, 35 wt% of the total amount of the emulsifier being added in the second time, and 15 wt% of the total amount of the emulsifier being added in the third time, the interval between each time of dropwise addition being 20-40 min; and the emulsifier is uniformly distributed in the solution by fully stirring. Next, 35 g of an oil phase (soybean oil) is slowly added dropwise into the solution while continuously stirring, forming tiny oil droplets. Then, the mixture is heated to 85°C for 35-40 min, so that the water in the oil droplets evaporates, forming a hollow structure. Finally, the unreacted substances and impurities are removed by filtration, washing and other steps, obtaining pure chitosan microspheres. In the chitosan microspheres, 20 wt% of the chitosan microspheres have a particle size less than 0.1 μm, 30 wt% of the chitosan microspheres have a particle size of 0.1-40 μm, and 50% of the chitosan microspheres have a particle size of 45-105 μm.

[0054] Example 2

[0055] On the basis of Example 1, 60 wt% of the total amount of the emulsifier is added dropwise in the first time, 39 wt% of the total amount of the emulsifier is added dropwise in the second time, and 1 wt% of the total amount of the emulsifier is added dropwise in the third time, the interval between each time of dropwise addition being 20-40 min. As a result, in the obtained chitosan microspheres, 10 wt% of the chitosan microspheres have a particle size less than 0.1 μm, 35 wt% of the chitosan microspheres have a particle size of 0.1-40 μm, and 55% of the chitosan microspheres have a particle size of 45-105 μm.

[0056] Example 3

[0057] On the basis of Example 1, adjust the first drop of 60wt% of the total amount of emulsifier, the second drop of 30wt% of the total amount of emulsifier, the third drop of 10wt% of the total amount of emulsifier, each drop interval 20-40min from the last drop. So that the resulting chitosan microspheres, including 15wt% of the particle size of less than 0.1 μm chitosan microspheres, 25wt% of the particle size of 0.1-40 μm chitosan microspheres and 60% of the particle size of 45-105 μm chitosan microspheres.

[0058] Comparative Example 1

[0059] On the basis of Example 1, adjust the amount of emulsifier is 0.375% of the mass of chitosan and drop in one time, so that the average particle size of chitosan microspheres is less than 0.1 μm.

[0060] Comparative Example 2

[0061] On the basis of Example 1, adjust the amount of emulsifier is 0.875% of the mass of chitosan and drop in one time, so that the particle size of chitosan microspheres is 0.1-40 μm.

[0062] Comparative Example 3

[0063] On the basis of Example 1, adjust the amount of emulsifier is 1.25% of the mass of chitosan and drop in one time, so that the particle size of chitosan microspheres is 45-105 μm.

[0064] Example 4

[0065] On the basis of Example 1, adjust the mixture heating to 70℃, get chitosan microspheres.

[0066] Example 5

[0067] On the basis of Example 1, adjust the mixture heating to 90℃, get chitosan microspheres.

[0068] Comparative Example 3

[0069] On the basis of Example 1, adjust the mixture heating to 60℃, get chitosan microspheres. The viscosity of the resulting chitosan microspheres will decrease, the adsorption performance decreases.

[0070] Comparative Example 4

[0071] On the basis of Example 1, adjust the mixture heating to 100℃, get chitosan microspheres. Chitosan microspheres crosslinking is not complete, affecting the stability and performance of the microspheres.

[0072] Example 6

[0073] On the basis of Example 1, adjust the amount of emulsifier is 1% of the mass of chitosan, get chitosan microspheres.

[0074] Example 7

[0075] On the basis of Example 1, the amount of emulsifier was adjusted to 5% of the mass of chitosan to obtain chitosan microspheres.

[0076] Comparative Example 5

[0077] On the basis of Example 1, the total amount of emulsifier was adjusted to 2.125% of the mass of chitosan, and was added dropwise twice. The amount of emulsifier added dropwise the first time and the second time was the same as in Example 1, and the interval between each dropwise addition was 20-40 min. Chitosan microspheres were obtained. The chitosan microspheres included 37.5 wt% of chitosan microspheres with a particle size of 0.1-40 μm and 62.5% of chitosan microspheres with a particle size of 45-105 μm.

[0078] Comparative Example 6

[0079] On the basis of Example 1, the total amount of emulsifier was adjusted to 1.75% of the mass of chitosan, and was added dropwise twice. The amount of emulsifier added dropwise the first time was the same as in Example 1, and the amount of emulsifier added dropwise the second time was the same as the amount of emulsifier added dropwise the third time in Example 1, and the interval between each dropwise addition was 20-40 min. Chitosan microspheres were obtained. The chitosan microspheres included 28.6 wt% of chitosan microspheres with a particle size of less than 0.1 μm and 71.4% of chitosan microspheres with a particle size of 45-105 μm.

[0080] Comparative Example 7

[0081] On the basis of Example 1, the total amount of emulsifier was adjusted to 1.25% of the mass of chitosan, and was added dropwise twice. The amount of emulsifier added dropwise the first time was the same as the amount of emulsifier added dropwise the second time in Example 1, and the amount of emulsifier added dropwise the second time was the same as the amount of emulsifier added dropwise the third time in Example 1, and the interval between each dropwise addition was 20-40 min. Chitosan microspheres were obtained. The chitosan microspheres included 40 wt% of chitosan microspheres with a particle size of less than 0.1 μm and 60 wt% of chitosan microspheres with a particle size of 0.1-40 μm.

[0082] Comparative Example 8

[0083] First, chitosan and polyethyleneimine (PEI) (mass ratio of chitosan to polyethyleneimine 1:3) were dissolved in 15 mL of a 15 wt% acetic acid solution as an aqueous phase system. 90 mL of liquid paraffin and 6 mL of Span 80 were uniformly mixed in a 250 mL three-necked flask to form an oil phase system. The three-necked flask was equipped with a condenser tube and mechanical stirring. The temperature of the water bath was adjusted to 35°C, and the mechanical stirring was controlled at 300 r / min -1。The aqueous phase solution was added drop by drop to the oil phase system, and emulsified for 1 h to form a white emulsion. Then 1.5 mL of 50 wt% glutaraldehyde solution was added dropwise, and the water phase monomers wrapped in the oil phase were crosslinked for 2.5 h. The reactants gradually changed from milky white to brown. The product was centrifuged and washed with petroleum ether, 0.5 M NaOH solution, deionized water, and anhydrous ethanol in sequence. The washed product was placed in a drying oven at 80°C for 12 h, and the product (CP) was collected and placed in a drying machine for standby.

[0084] The chitosan microspheres prepared in the above examples and comparative examples were used to prepare Baoyuan oral liquid, and the specific process was as follows:

[0085] Ginseng (five years old) 149.2 parts, Huangqi (wild-like) 298.4 parts, Gancao 74.8 parts, Rougui (fifteen years old) 30.0 parts, Shengjiang (small yellow ginger) 120.0 parts; After the above five traditional Chinese medicines were cleaned, soaked, cut, and dried, they were soaked in 12 times the amount of ultrapure water for 55 minutes, boiled with strong fire, and then transferred to a weak fire to keep it slightly boiling for 1.2 hours. The filtrate was filtered through a 0.45 um ceramic inorganic membrane, and the residue was added with 11 times the amount of ultrapure water and boiled with strong fire, then transferred to a weak fire to keep it slightly boiling for 45 minutes. The filtrate was filtered through a 0.45 um ceramic inorganic membrane, and the filtrate was combined and concentrated under vacuum at 65°C to 400 ml. The pH value of the system was adjusted to 4.5, and the flavoring agent steviol glycoside 0.2 g and xylitol 32 g were added. The filtrate was filtered, adjusted to a total volume of 500 ml, stirred uniformly, filled, sterilized, and lamp checked to obtain the product.

[0086] The amount of chitosan microspheres added was 0.75% of the sample solution (400 ml).

[0087] The amount of carboxymethyl cellulose sodium CMC-Na as stabilizer was 0.09% of the sample solution (400 ml).

[0088] The light transmittance, soluble solids and anti-immune antibody level of Baoyuan oral liquid were tested, and the test method was as follows: the absorbance of the clarified liquid was measured at 623 nm wavelength by ultraviolet spectrophotometer, and the light transmittance was calculated according to the conversion formula of absorbance and light transmittance. Reference: Lu M, Zhang HX, Fu J, et al. Relationship between carbon black surface energy and toluene extract light transmittance [J. Rubber Industry, 2020, 67(9): 709-712.

[0089] The test method of the anti-immune antibody level is as follows: a 24-well cell culture plate is taken, 950 μL of mouse spleen lymphocyte (LP) suspension is added to each well, and three replicate wells are set for each group. After pre-incubation for 1 h, anti-mouse CD3 antibody (1.25 μg / mL, purchased from Biopure) is added to stimulate the spleen lymphocytes (10 μL of PBS is used for the control group), and the plate is cultured in a 37℃, 5% CO2 incubator, and gently shaken once after 30 min to remove the unadherent cells. After 3 h, the unadherent cells are removed by washing the culture solution twice, and the culture solution containing Baoyuan oral liquid (the concentration of the Baoyuan oral liquid is 3 mg / mL) is added, and the plate is continuously cultured for 48 h. The supernatant is collected by centrifugation at 3000 r / min, and diluted with the culture solution at a ratio of 1:5. The OD value is measured at 450 nm in a microplate reader, and the anti-immune antibody level is calculated according to the standard curve.

[0090] The results are as follows:

[0091] Table 1 Properties of the Baoyuan oral liquid prepared from the chitosan microspheres prepared in Examples 1-7 and Comparative Examples 1-8

[0092] Sample Transmittance (%) Soluble solids (%) Anti-CD3 antibody (μg / mL) Example 1 92 4.6 21.2 Example 2 91 4.3 21.1 Example 3 91 4.5 20.9 Example 4 90 4.2 21.1 Example 5 92 4.3 20.8 Example 6 91 4.2 20.7 Example 7 90 4.3 21.1 Comparative Example 1 84 3.6 18.2 Comparative Example 2 82 3.4 18.1 Comparative Example 3 83 3.3 17.9 Comparative Example 4 85 3.2 17.8 Comparative Example 5 82 3.4 18.3 Comparative Example 6 84 3.3 18.2 Comparative Example 7 83 3.2 17.9 Comparative Example 8 82 3.5 17.6

[0093] The states of the Baoyuan oral liquids prepared in Examples 1 and 2 after standing at a temperature of 25 and a humidity of 60% for 30 d are shown in Figs. 1 and 2, respectively. The states of the Baoyuan oral liquids prepared in Comparative Examples 1 and 2 after standing at a temperature of 25 and a humidity of 60% for 30 d are shown in Figs. 3 and 4, respectively. It can be seen that the stability and clarity of the oral liquids are obviously different. Figure 1 Figure 2 Figure 3 Figure 4 It can be seen that the stability and clarity of the oral liquids are obviously different.

[0094] Example 8

[0095] A Baoyuan oral liquid for regulating immune function and a preparation method thereof, comprising the following raw materials in parts by weight: ginseng (five years old) 134.3 parts, astragalus (wild-like) 268.6 parts, licorice 67.3 parts, cassia (fifteen years old) 27.0 parts, and ginger (small yellow ginger) 108.0 parts; the above five ingredients are cleaned, soaked, cut, and dried, and then soaked in 11 times the amount of ultrapure water for 50 minutes. After boiling with a strong fire, the temperature is reduced to a weak fire to maintain a slight boiling for 1.3 hours. The liquid is filtered with a 0.45 um ceramic inorganic membrane. The residue is boiled with 10 times the amount of ultrapure water, and then the temperature is reduced to a weak fire to maintain a slight boiling for 55 minutes. The liquid is filtered with a 0.45 um ceramic inorganic membrane. The filtrates are combined, vacuum concentrated to 400 ml at 65℃, the pH value of the system is adjusted to 4.5, and then 0.2 g of sweetener steviol glycoside and 29 g of wood sugar are added. The mixture is filtered, adjusted to a total amount of 500 ml, stirred uniformly, filled, sterilized, and light checked to obtain the product.

[0096] ​​​The preparation method of the chitosan microspheres is as follows: first, 100 g of chitosan is dissolved in an acetic acid solution with a pH value of 3.8, and the pH value of the system is kept at 3.2-3.3. Then, an emulsifier (sodium dodecyl sulfate) is added, and the amount of the emulsifier is 3.5% of the mass of the chitosan. The emulsifier is added in three times, 50 wt% of the total amount of the emulsifier is added in the first time, 35 wt% of the total amount of the emulsifier is added in the second time, and 15 wt% of the total amount of the emulsifier is added in the third time. The interval between each time of adding is 20-40 min. The emulsifier is uniformly distributed in the solution by fully stirring. Then, 35 g of an oil phase (soybean oil) is slowly added into the water phase while continuously stirring to form small oil droplets. The mixture is heated to 80℃ and kept for 35-40 min to evaporate the water in the oil droplets and form a hollow structure. Finally, the unreacted substances and impurities are removed by filtration, washing and other steps to obtain pure chitosan microspheres. In the chitosan microspheres, 20 wt% of the chitosan microspheres have a particle size of less than 0.1 μm, 20 wt% of the chitosan microspheres have a particle size of 0.1-40 μm, and 60% of the chitosan microspheres have a particle size of 45-105 μm. Among the chitosan microspheres with a particle size of 45-105 μm, the particle size is mostly above 80 μm.

[0097] The amount of the chitosan microspheres added is 0.15% of the sample solution (400 ml).

[0098] The amount of the stabilizer sodium carboxymethyl cellulose CMC-Na added is 0.12% of the sample solution (400 ml).

[0099] Comparative Example 9 (diatomite, compared with Example 8)

[0100] A Baoyuan oral liquid for regulating immune function and a preparation method thereof, including the following raw materials by weight: 134.3 parts of ginseng (five years old), 268.6 parts of radix astragali (wild-like), 67.3 parts of licorice, 27.0 parts of cassia (fifteen years old), and 108.0 parts of ginger (small yellow ginger); after the above five ingredients are cleaned, soaked, cut, and baked, 11 times the amount of ultrapure water is added for soaking for 50 minutes. After boiling with a strong fire, the temperature is kept at a slight boil with a weak fire for 1.3 hours. The liquid is filtered with a 0.45 um ceramic inorganic membrane. The residue is boiled with 10 times the amount of ultrapure water with a strong fire, and then the temperature is kept at a slight boil with a weak fire for 55 minutes. The liquid is filtered with a 0.45 um ceramic inorganic membrane. The filtrates are combined and concentrated to 400 ml under vacuum at 65℃. The pH value of the system is adjusted to 4.5. Sweetener steviol glycoside 0.2 g, xylitol 29 g, clarifying agent diatomite 0.6% (purchased from Hebei Yousheng Refractory Materials), stabilizer sodium carboxymethyl cellulose CMC-Na are added. After filtration, the total amount is adjusted to 500 ml. After stirring, the liquid is filled, sterilized, and checked with a light. The Baoyuan oral liquid is obtained.

[0101] Comparative Example 10 (ZTC1+1 clarifying agent)

[0102] A Baoyuan oral liquid for regulating immune function and a preparation method thereof, comprising the following raw materials in parts by weight: Radix ginseng (five years old) 134.3 parts, Radix astragali (pseudo-wild) 268.6 parts, Glycyrrhiza 67.3 parts, Cinnamomum cassia (fifteen years old) 27.0 parts, and Zingiber officinale (small yellow ginger) 108.0 parts; after the above five ingredients are cleaned, dampened, cut, and baked, 11 times the amount of ultrapure water is added for soaking for 50 minutes, and then boiled with a strong fire, followed by keeping a slight boil with a weak fire for 1.3 hours; the medicinal liquid is filtered with a 0.45 um ceramic inorganic membrane, the residue is boiled with 10 times the amount of ultrapure water with a strong fire, and then kept a slight boil with a weak fire for 55 minutes; the medicinal liquid is filtered with a 0.45 um ceramic inorganic membrane, the filtrates are combined, vacuum concentrated to 400 ml at 65℃, the pH value of the system is adjusted to 4.5, 0.2 g of sweetener steviol glycoside, 29 g of xylitol, 0.6% of ZTC1+1 clarifier (purchased from Tianjin Zhengtiancheng Clarification Technology Co., Ltd.), and stabilizer sodium carboxymethyl cellulose CMC-Na are added, filtered, adjusted to a total amount of 500 ml, stirred uniformly, filled, sterilized, and lamp checked to obtain the product.

[0103] Comparative Example 11 (conventional chitosan gel, without acidification and microspherization operation)

[0104] A Baoyuan oral liquid for regulating immune function and a preparation method thereof, comprising the following raw materials in parts by weight: Radix ginseng (five years old) 134.3 parts, Radix astragali (pseudo-wild) 268.6 parts, Glycyrrhiza 67.3 parts, Cinnamomum cassia (fifteen years old) 27.0 parts, and Zingiber officinale (small yellow ginger) 108.0 parts; after the above five ingredients are cleaned, dampened, cut, and baked, 11 times the amount of ultrapure water is added for soaking for 50 minutes, and then boiled with a strong fire, followed by keeping a slight boil with a weak fire for 1.3 hours; the medicinal liquid is filtered with a 0.45 um ceramic inorganic membrane, the residue is boiled with 10 times the amount of ultrapure water with a strong fire, and then kept a slight boil with a weak fire for 55 minutes; the medicinal liquid is filtered with a 0.45 um ceramic inorganic membrane, the filtrates are combined, vacuum concentrated to 400 ml at 65℃, the pH value of the system is adjusted to 4.5, 0.2 g of sweetener steviol glycoside, 29 g of xylitol, 0.6% of ZTC1+1 clarifier (purchased from Tianjin Zhengtiancheng Clarification Technology Co., Ltd.), and stabilizer sodium carboxymethyl cellulose CMC-Na are added, filtered, adjusted to a total amount of 500 ml, stirred uniformly, filled, sterilized, and lamp checked to obtain the product.

[0105] Comparative Example 12 (without adding stabilizer)

[0106] Compared with Example 8, the only difference is that no stabilizer sodium carboxymethyl cellulose CMC-Na is added, and the others are the same.

[0107] Comparative Example 13 (without adding clarifier)

[0108] Compared with Example 8, the only difference is that no clarifier acidic chitosan microspheres is added, and the others are the same. The state of the Baoyuan oral liquid prepared in Comparative Example 12 after standing for 30 days at a temperature of 25 and a humidity of 60% is as follows:Figure 5 as shown.

[0109] Performance tests were carried out on Example 8 and Comparative Examples 9-13, and the test methods were as above. The test results are shown in the following table:

[0110] Table 2 Performance tests of Baoyuan oral liquid of Example 8 and Comparative Examples 9-13

[0111] Sample Transmittance (%) Soluble solids (%) Anti-CD3 antibody (μg / mL) Example 1 92 4.6 21.2 Example 8 90 4.3 21.3 Comparative Example 9 82 3.5 17.9 Comparative Example 10 88 3.2 17.8 Comparative Example 11 84 3.7 17.6 Comparative Example 12 87 3.9 18.1 Comparative Example 13 82 3.2 17.7

[0112] From the above table, the performance of the examples is better than that of the comparative examples, and the main reasons may be: traditional Chinese medicine decoction is mostly a compound preparation, and water is mostly used as a solvent during extraction, so some high molecular substances such as starch, gum, protein, and sugar are also extracted. The traditional Chinese medicine decoction is essentially a mixture of solution, colloid, suspension and emulsion, and the particles in the colloid and suspension will settle and coalesce when standing, and the emulsion will easily emulsify and break when the conditions change. At the same time, due to the different charges carried by the particles of colloid, suspension and emulsion, they will also attract each other, resulting in a large amount of precipitation after standing. Although there are so many reasons for the formation of the precipitate, involving the acid-base combination of chemical components of traditional Chinese medicine, tannin precipitation, hydrolysis degradation and polymerization, etc. However, in general, the principle of treatment is to remove the precipitate to the maximum extent without affecting the clinical efficacy. The selection of oral liquid clarifying agent should first meet the safety, and then the retention rate of active ingredients, clarity, etc. Common clarifying agents include diatomite, ZTC1+1 clarifying agent, water-soluble chitosan, etc. Diatomite is not suitable for filtering oral liquid, and it is difficult to filter oral liquid with high concentration using diatomite. ZTC1+1 has good filtering effect on oral liquid, but ZTC1+1 is relatively expensive. Compared with acid-soluble chitosan, water-soluble chitosan has loose flocculation, and the effect is not as good as that of acid-soluble chitosan, so the acid-soluble chitosan is obtained by acidifying chitosan in the present application, and the acid-soluble chitosan is microspherized to strengthen its porous adsorption function, so that the clarity effect is better. In addition, the stabilizer carboxymethyl cellulose sodium CMC-Na is added in the preparation to ensure the clarity and stability of the preparation. The present application can not only solve the problem of Baoyuan oral liquid precipitation, but also improve the efficacy of the drug by using new pharmaceutical technology and more reasonable preparation process, which also provides a new idea for the application and development of traditional Chinese medicine oral liquid dosage forms.

Claims

1. Chitosan microspheres characterized in that, Chitosan 80-120 parts by weight, emulsifier 2-5 parts, oil phase 30-40 parts; the chitosan microspheres are obtained by dissolving in an acidic solution and then reacting with the emulsifier and the oil phase; the chitosan microspheres include 10-20 wt% of chitosan microspheres with a particle size of less than 0.1 µm, 20-40 wt% of chitosan microspheres with a particle size of 0.1-40 µm, and 45%-70% of chitosan microspheres with a particle size of 45-105 µm; The pH value of the acidic solution is 2-4.

2. The chitosan microspheres according to claim 1, characterized in that, The acidic solution includes one or both of an acetic acid solution or a phosphoric acid solution; the emulsifier includes one or both of sodium dodecyl sulfate or Tween; and the oil phase is one or more of soybean oil, olive oil, or sunflower seed oil.

3. The method of claim 1 or 2, wherein the method is characterized by, The method comprises the following steps: Chitosan is dissolved in an acidic solution, an emulsifier is added, stirring is performed, then an oil phase is added dropwise, after the dropwise addition is completed, the reaction system is heated to 70-90°C, and chitosan microspheres are obtained by purification after 30-50 min of reaction; the emulsifier is added dropwise in multiple times, and each dropwise addition is separated from the previous dropwise addition by 20-40 min; the emulsifier is added dropwise in three times, 50-60 wt% of the total amount of the emulsifier is added dropwise in the first time, 30-39 wt% of the total amount of the emulsifier is added dropwise in the second time, and 1-20 wt% of the total amount of the emulsifier is added dropwise in the third time, and each dropwise addition is separated from the previous dropwise addition by 20-40 min.

4. Use of the chitosan microspheres according to claim 1 or 2 in the preparation of a Baoyuan oral liquid preparation.

5. A method for preparing a Baoyuan oral liquid preparation, characterized in that, The method comprises the following steps: S1. Ginseng 140-160 parts, Radix Astragali 300-320 parts, Radix Glycyrrhizae 70-85 parts, Cinnamomum cassia bark 28-35 parts, and raw ginger 110-130 parts are mixed and baked, then 12-18 times the amount of water is added, the mixture is soaked for 40-70 min, boiled, then kept at a slight boil for 1.2-1.5 h, filtered, and a filtrate 1 and a residue are obtained; S2. The residue is boiled with 10-14 times the amount of water, then kept at a slight boil for 35-65 min, filtered, and the filtrates are combined, concentrated, and adjusted to a pH value of 4.3-5.8; S3. A clarifying agent 2-25 parts and a stabilizer 0.2-1.2 parts are added and mixed uniformly, filtered, and the Baoyuan oral liquid preparation is obtained; The clarifying agent is the chitosan microspheres according to claim 1 or 2.

6. The preparation method according to claim 5, characterized in that, The filtering is performed by using a ceramic inorganic membrane.

7. The preparation method according to claim 6, characterized in that, The filtering is performed by using a ceramic inorganic membrane with a membrane pore size of 0.1-0.45 µm.

8. A Baoyuan oral liquid preparation prepared by the method according to claim 6 or 7.

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