Composition of piveronium bromide and fructus rosae laevigatae extract as well as preparation process and application of composition

Abstract

The invention belongs to the technical field of pharmaceutical preparations, and particularly relates to a composition of piveronium bromide and a cherokee rose fruit extract, a preparation process and application. The tablet is composed of piveronium bromide, a rosa laevigata michx extract, a disintegrating agent, a lubricant and a filler, and pharmacological experiments of rats find that the tablet has a significant treatment effect on diarrhea-type irritable bowel syndrome, can be completely dissolved out within 15 min, has low content of related substances, is higher than the requirements of pharmacopeia, and is suitable for further industrial mass production.

Description

Technical Field

[0001] The invention belongs to the technical field of pharmaceutical preparations, and particularly relates to a composition of pinaverium bromide and a Rosa laevigata extract, a preparation process and uses thereof. Background Art

[0002] Pinaverium bromide was developed by the French pharmaceutical company Solvay and first marketed in 1975. Its structural formula is as follows:

[0003]

[0004] Pinaverium bromide is a calcium antagonist with highly selective antispasmodic effect on the gastrointestinal tract. Its mechanism of action on smooth muscle is the same as other calcium antagonists, but it has a highly selective effect on colon smooth muscle. Pinaverium bromide blocks the influx of calcium ions into the smooth muscle cells of the intestinal wall, preventing excessive muscle contraction and achieving antispasmodic effect. It can eliminate the high reactivity of intestinal smooth muscle and increase intestinal motility.

[0005] The Latin name of Rosa laevigata Michx is Rosa laevigata Michx. It has a long history of medicinal use and is recorded in ancient medical classics such as "Compendium of Materia Medica", "Chinese Materia Medica", "Lingnan Herbal Medicine", "Jiangxi Folk Herbal Prescriptions", and "Hunan Herbal Medicine". It is widely distributed in Guangxi, Guangdong, Hunan and other places. It is the main raw material for Gynecological Qianjin Tablets, Jinji Capsules, Sanjin Tablets, and Wanglaoji Herbal Tea. The chemical components of its roots are mainly triterpenoid saponins, flavonoids, carbohydrates, etc., but the research on its material basis and mechanism of action is not in-depth enough.

[0006] In the prior art, it is not seen that pinaverium bromide and Rosa laevigata are prepared into a pharmaceutical composition for use, and the two are not prepared into tablets for related experiments. Summary of the invention

[0007] Based on the deficiencies in the prior art, a composition, preparation process and use of pinaverium bromide and Rosa laevigata extract are provided, which solve the problem that the active substance of pinaverium bromide is unstable during storage and is easily decomposed when exposed to light and heat, and is suitable for further large-scale industrial production.

[0008] Specifically, the inventors solved the problem through the following technical solutions:

[0009] The first object of the present invention is to provide a tablet containing pinaverium bromide, which is composed of the following raw materials: 10 parts by weight of pinaverium bromide, 0.5-1.5 parts by weight of Rosa laevigata extract, 2-10 parts by weight of disintegrant, 0.1-1.1 parts by weight of lubricant, and 40-130 parts by weight of filler.

[0010] Furthermore, the pinaverium bromide tablet is composed of the following raw materials: 10 parts by weight of pinaverium bromide, 1 part by weight of Rosa laevigata extract, 3 parts by weight of disintegrant, 0.6 parts by weight of lubricant, and 85 parts by weight of filler.

[0011] Furthermore, the filler is selected from one or two of microcrystalline cellulose, lactose, mannitol, and starch; more preferably, the filler is selected from a combination of microcrystalline cellulose and lactose.

[0012] In a preferred embodiment of the present invention, the ratio of microcrystalline cellulose to lactose is 2-6:2-7 by weight; more preferably, the ratio of microcrystalline cellulose to lactose is 8:9.

[0013] Furthermore, the disintegrant is selected from one of low-substituted hydroxypropyl cellulose and sodium carboxymethyl starch, preferably sodium carboxymethyl starch; the lubricant is selected from any one of magnesium stearate, talc, and sodium dodecyl sulfate, preferably magnesium stearate.

[0014] In a preferred embodiment of the present invention, the pinaverium bromide tablets are composed of the following raw materials: 10 parts by weight of pinaverium bromide, 0.5-1.5 parts by weight of Rosa laevigata extract, 2-10 parts by weight of sodium carboxymethyl starch, 0.1-1.1 parts by weight of magnesium stearate, 20-60 parts by weight of microcrystalline cellulose, and 20-70 parts by weight of lactose.

[0015] In a preferred embodiment of the present invention, the pinaverium bromide tablets are composed of the following raw materials: 10 parts by weight of pinaverium bromide, 1 part by weight of Rosa laevigata extract, 3 parts by weight of sodium carboxymethyl starch, 0.6 parts by weight of magnesium stearate, 40 parts by weight of microcrystalline cellulose, and 45 parts by weight of lactose.

[0016] In a preferred embodiment of the present invention, the pinaverium bromide tablets may be coated.

[0017] The second object of the present invention is that the present invention also provides a method for extracting the Rosa Laevigata extract:

[0018] 1) Take the fruit powder of Rosa laevigata and add 30-60% sodium chloride to soak for 30-50 minutes, then heat to 70-80° C. to extract the medicinal liquid, and keep the residue and the extract for later use;

[0019] 2) Repeat step 1) with the residue and extract twice; combine the filtrates from the three extractions;

[0020] 3) collecting the filtrate from step 2), concentrating under reduced pressure at 50-60° C. to an alcohol content of 80-90%; standing at room temperature for 24 hours, centrifuging, collecting the precipitate and supernatant, and the precipitate is the Rosa laevigata extract;

[0021] 4) Deproteinizing the polysaccharide extract obtained in step 3) by Sevag method, centrifuging, concentrating the supernatant, precipitating with ethanol, collecting the precipitate, and vacuum drying to obtain the Rosa laevigata extract.

[0022] Step 4) is specifically as follows: using the Sevag method to remove protein from the Rosa laevigata extract obtained in step 3), centrifuging, concentrating the supernatant, taking the mixed Rosa laevigata extract solution and adding 95% ethanol in a volume ratio of 1:4, placing it in a refrigerator (4°C) and standing it for 24 hours overnight, centrifuging it at 4000r / min for 15min, collecting the precipitate and adding an appropriate amount of deionized water to dissolve it to obtain the Rosa laevigata extract.

[0023] The third object of the present invention is that the present invention also provides a method for preparing the pinaverium bromide tablets:

[0024] 1) Dissolve glyceryl monostearate in a mixed solution of chloroform and 95% or more ethanol, stirring continuously to mix well;

[0025] 2) adding pinaverium bromide to step 1) and mixing, stirring to dissolve;

[0026] 3) adding the Rosa laevigata extract evenly to the mixed solution of step 2), maintaining the temperature at 40-50° C. and performing rotary evaporation under reduced pressure to dryness, adding microcrystalline cellulose, lactose, and sodium carboxymethyl starch, blending, sieving, adding magnesium stearate, and tableting to obtain plain tablets;

[0027] 4) Weigh the coating premix powder, add water and mix to prepare a film coating premix, apply film coating to the above plain tablets, the coating weight gain is 3-10%, and dry to obtain tablets containing pinaverium bromide.

[0028] The fourth object of the present invention is to mix the pinaverium bromide of the present invention with the extract of Rosa Laevigata, and provide the use of the pinaverium bromide tablets in the preparation of drugs for treating diarrhea-type irritable bowel syndrome. The inventors created the combined use of the Rosa Laevigata extract and pinaverium bromide. In the pharmacological experiment on rats, it was found that on the 21st and 35th days, the novelty inhibition of feeding latency of rats in Example 1-3 groups was significantly shortened compared with the positive group, Comparative Example 1 group and Comparative Example 2 group, and the content of D-lactic acid and TNF-α, IFN-γ, and IL-6 in serum was significantly reduced, indicating that the combined use of pinaverium bromide and the Rosa Laevigata extract of the present invention has achieved unexpected technical effects.

[0029] The fifth object of the present invention is to provide a tablet containing a Rosa Laevigata extract and pinaverium bromide, wherein the dissolution of pinaverium bromide in 15 minutes can be more than 95%, which is better than the requirements of the pharmacopoeia. In the accelerated test at 0, 1, 2, 3, and 6 months, the solubility of Examples 1-5 under the condition of pH 1.0 HCl solution was more than 99%, which was significantly higher than the dissolution requirements of the Chinese Pharmacopoeia. In the accelerated test, the solubility of Comparative Examples 1, 3, 4, 5, and 6 at 15 minutes was less than 85%, but Comparative Examples 3-5 were already less than 80%, which could not meet the requirements of the pharmacopoeia. At the same time, the Rosa Laevigata extract was optimized, and it was found that the pretreatment and extraction process had a greater impact on it; a filler different from the present invention was selected, and during storage, the Rosa Laevigata extract had varying degrees of degradation.

[0030] The content of related substances in the tablet of the present invention is low, and the content of related substances in Examples 1-5 is within 0.5%. In comparison, Comparative Example 1 does not contain the extract of Rose Laevigata, and the extract is particularly important for the stability of Pinaverium Bromide. The inventor believes that polysaccharides can form a protective film on the surface of the drug. This film can isolate the contact of drug molecules with the external environment, slow down the influence of moisture, oxygen and light on Pinaverium Bromide, thereby reducing or avoiding the rate of hydrolysis reaction, thereby improving the stability of the drug. The extract of Rose Laevigata in Comparative Example 2 is too high, and other substances in it may react with Pinaverium Bromide to cause degradation. The different extracts used in Comparative Example 6 also have an impact on the related substances of the final product. Comparative Example 5 uses different preparation processes, and the difference in preparation processes has a greater impact on Pinaverium Bromide. It can be seen from this that Pinaverium Bromide is relatively sensitive to light, temperature and other auxiliary materials, and not taking relevant protection methods will accelerate its degradation. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 : Comparison of dissolution rates of each example at 0, 1, 2, 3 and 6 months.

[0032] Figure 2 : Comparison of related substances of each example at 0, 1, 2, 3 and 6 months.

[0033] Figure 3 : Comparison of novelty-suppressed food intake latency in rats of each group in the 0th experiment.

[0034] Figure 4 :Comparison of novelty-suppressed food intake latency in rats of each group in the 21st experiment.

[0035] Figure 5 :Comparison of novelty-suppressed food intake latency in rats of each group in the 35th experiment.

[0036] Figure 6 :Comparison of D-lactic acid content in rats of each experimental group.

[0037] Figure 7 : Comparison of D-lactic acid and TNF-α content in serum of rats in each group of the experiment.

[0038] Figure 8 : Comparison of D-lactic acid and IL-6 content in serum of rats in each group of the experiment.

[0039] Fig. 9 : Comparison of D-lactic acid and IFN-γ content in serum of rats in each experimental group. DETAILED DESCRIPTION

[0040] The present invention discloses an infant formula milk powder capable of improving the immunity of the body and a preparation method thereof. Those skilled in the art can refer to the content of this article and appropriately change the process parameters. It should be particularly noted that all similar replacements and modifications are obvious to those skilled in the art, and they are all considered to be included in the present invention. The method and application of the present invention have been described through a comparative embodiment, and relevant personnel can obviously modify or appropriately change and combine the method and application described herein without departing from the content, spirit and scope of the present invention to implement and apply the technology of the present invention.

[0041] Example 1: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0042] Pinaverium bromide 10g, Rosa laevigata extract 0.5g, sodium carboxymethyl starch 2g, magnesium stearate 0.1g, microcrystalline cellulose 20g, lactose 20g.

[0043] Preparation process:

[0044] 1) dissolving glyceryl monostearate in a mixed solution of chloroform and 95% or more ethanol, stirring constantly and mixing; wherein the weight volume ratio of glyceryl monostearate is 1:100, and the volume ratio of chloroform to ethanol is 2:1;

[0045] 2) adding pinaverium bromide to step 1) and mixing, stirring to dissolve;

[0046] 3) adding the Rosa laevigata extract evenly to the mixed solution of step 2), maintaining the temperature at 40-50° C. and performing rotary evaporation under reduced pressure to dryness, adding microcrystalline cellulose, lactose, and sodium carboxymethyl starch, blending, sieving, adding magnesium stearate, and tableting to obtain plain tablets;

[0047] 4) Weigh 10 mg of coating premix powder and add 40 mg of water to make a film coating premix, and film coat the above plain tablets. The coating weight gain is 3-10%, and then dried. The material temperature is set to 35° C. and continued to dry until the free moisture is below 3%, to obtain tablets containing pinaverium bromide.

[0048] Example 2: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0049] Pinaverium bromide 10g, Rosa laevigata extract 1.5g, sodium carboxymethyl starch 10g, magnesium stearate 1.1g, microcrystalline cellulose 60g, lactose 70g.

[0050] The preparation process is the same as that of Example 1.

[0051] Example 3: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0052] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0053] The preparation process is the same as that of Example 1.

[0054] Example 4: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0055] Pinaverium bromide 10g, Rosa laevigata extract 1g, low-substituted hydroxypropyl cellulose 3g, talc 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0056] The preparation process is the same as that of Example 1.

[0057] Example 5: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0058] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, micro powder silica magnesium 0.6g, microcrystalline cellulose 40g, mannitol 45g.

[0059] The preparation process is the same as that of Example 1.

[0060] Example 6: A Pinaverium Bromide Tablet and Its Preparation Method

[0061] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, micronized silica magnesium 0.6g, sorbitol 40g, lactose 45g.

[0062] The preparation process is the same as that of Example 1.

[0063] Comparative Example 1: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0064] Pinaverium bromide 10g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0065] The preparation process is the same as that of Example 1.

[0066] Comparative Example 2: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0067] Pinaverium bromide 10g, Rosa laevigata extract 2g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0068] The preparation process is the same as that of Example 1.

[0069] Comparative Example 3: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0070] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 85g.

[0071] The preparation process is the same as that of Example 1.

[0072] Comparative Example 4: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0073] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, sucrose 45g.

[0074] The preparation process is the same as that of Example 1.

[0075] Comparative Example 5: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0076] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0077] Preparation process:

[0078] Add pinaverium bromide and Rosa laevigata extract to a mixed solution of ethanol with a concentration of more than 95%, and stir continuously to mix well; maintain the temperature at 40-50°C and perform reduced pressure rotary evaporation to dryness, add microcrystalline cellulose, lactose, and sodium carboxymethyl starch, blend them, sieve them, add magnesium stearate, and press them into tablets to obtain plain tablets; weigh 10 mg of coating premix powder, add 40 mg of water and mix them to prepare a film coating premix, film-coat the above plain tablets, dry them, set the material temperature to 35°C, and continue drying until the free moisture is below 3%, to obtain tablets containing pinaverium bromide.

[0079] Comparative Example 6: A pinaverium bromide tablet and a preparation method thereof

[0080] Pinaverium bromide 10g, Panax notoginseng polysaccharide 1g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0081] 1. Dissolution of Pinaverium Bromide Tablets and Determination of Related Substances

[0082] 1.1 Dissolution test and results of pinaverium bromide tablets

[0083] This experimental example investigates the dissolution changes of commercially available pinaverium bromide tablets (Beijing Fuyuan Pharmaceutical Co., Ltd., batch number national medicine standard H20133036), Examples 1-6 and Comparative Examples 1-6 containing pinaverium bromide tablets under pH 1.0 HCl solution conditions.

[0084] Specific dissolution determination method: pH 1.0 HCl solution is used as the medium, and the dissolution is carried out at a rotation speed of 50 rpm / min. The ultraviolet spectrophotometry method is used for detection, and the detection wavelength is 243nm (based on pinaverium bromide). The test results are shown in Figure 1 .

[0085] According to the conventional requirements of tablets, the dissolution rate should be above 80% within 15 minutes. Figure 1 It can be seen that the dissolution of Examples 1-5 in the HCl solution at pH 1.0 was above 99% at 0, 1, 2, 3, and 6 months of the accelerated test, and the tablets containing pinaverium bromide prepared in Examples 1-5 all met the dissolution requirements of the Chinese Pharmacopoeia. However, in the accelerated test, the dissolution of Comparative Examples 1, 3, 4, 5, and 6 at 15 minutes was lower than 85%, and Comparative Examples 3-5 were lower than 80%, which could not meet the requirements of the Pharmacopoeia.

[0086] 1.2 Determination of related substances in pinaverium bromide tablets and results

[0087] Determination method of related substances: Determined by high performance liquid chromatography, refer to the 2020 edition of the Chinese Pharmacopoeia

[0088] Chromatographic conditions: octadecylsilane bonded silica gel as filler, acetonitrile-sodium dodecyl sulfate buffer (70:30) as mobile phase, detection wavelength of 245nm, column temperature of 25℃, flow rate of 1.0mL / min, injection volume of 20μL. Preparation method of sodium dodecyl sulfate buffer: Take 13.0g sodium dodecyl sulfate, add 1500mL of water to dissolve, add 5.0mL of phosphoric acid, mix well, adjust pH to 2.5 with triethylamine, and filter to obtain.

[0089] Depend on Figure 2It can be seen that compared with comparative examples 1-6, the total impurity content of the tablets of embodiments 1-6 of the present invention in the first accelerated test is substantially unchanged, while the total impurity content of comparative examples 1, 2, 3, and 5 is significantly increased, indicating that the stability of the tablets containing pinaverium bromide prepared by the method of the present invention is significantly improved, wherein the stability of embodiment 3 is the best. Comparative example 1 does not contain the extract of the fruit of golden rosa laevigata, and the extract is particularly important for the stability of pinaverium bromide. The inventor believes that polysaccharides can form a protective film on the surface of the drug. This film can isolate the contact of drug molecules with the external environment, slow down the influence of moisture, oxygen and light on pinaverium bromide, thereby reducing or avoiding the rate of hydrolysis reaction, thereby improving the stability of the drug. The extract of the fruit of golden rosa laevigata of comparative example 2 is too high, and other substances inside may react with pinaverium bromide to cause degradation, and the different adjuvants used in comparative example 3 also have an impact on the final product, and comparative example 5 uses different preparation processes, and the difference in preparation processes also has an impact on pinaverium bromide, and pinaverium bromide is relatively sensitive to light and temperature, and does not take relevant means to accelerate its degradation.

[0090] 2. Pharmacological Experiment on Rats of Tablets of the Composition of the Present Invention

[0091] 2.1 Modeling and grouping

[0092] 2.1.1 Modeling

[0093] A total of 72 SPF Wistar female rats were adaptively fed for 1 week, and 8 rats were randomly selected as the blank group using a random number table. The remaining rats were treated with a compound modeling method of intestinal perfusion of acetic acid + tail clamp stress + mold restriction to prepare a diarrhea-predominant irritable bowel syndrome model.

[0094] From day 1 to day 14, all rats were fasted 12 hours before enema. 3% sodium pentobarbital was injected intraperitoneally at 50 mg / kg at 8:30 every day for anesthesia. After anesthesia, an 8F catheter lubricated with paraffin oil was inserted 8 cm through the anus. 1 mL of 4% acetic acid was injected into the model rats. After 30 seconds, an equal amount of phosphate buffered saline was injected. The blank group was injected with 1 mL of saline. From day 7 to day 21, the model rats were restrained in a mold for 1 hour at 15:00 every day, and then the plastic clip was clamped at the 1 / 3 of the rat's tail for 1 hour. The blank group was not intervened. The significant increase in fecal water content was regarded as the standard for successful modeling (the fecal water content of the model group and the blank group was significantly different P<0.01, indicating that the modeling was successful).

[0095] 2.1.2 Grouping

[0096] The rats were divided into 5 groups according to the random number table: model group, positive control group, Example 1-3 groups, Comparative Example 1 group, Comparative Example 6 group, 8 rats in each group, and a blank group with no treatment.

[0097] 2.3 Administration

[0098] The pinaverium bromide tablets and positive control drugs prepared in each example were ground into 50 mg of pinaverium bromide and dissolved in 25 mL of distilled water to prepare a 2.50 mg / mL suspension. The above examples and suspensions prepared as needed were placed in a 4°C refrigerator for standby use and gavaged twice a day in the morning and evening; the positive control group and each example group were given 5.40 mL·kg of pinaverium bromide suspension -1 ·d -1 (i.e. 13.50 mg kg -1 ), once a day by gavage; the blank group and the model group were given 10 mL·kg of normal saline -1 ·d -1 The drug was administered by intragastric gavage for 14 consecutive days.

[0099] 2.4 Specimen collection and processing

[0100] After the last administration of each group, the rats were fasted but not watered for 24 hours. After weighing their body weight, they were anesthetized by intraperitoneal injection of 3% sodium pentobarbital at 50 mg / kg. About 4 mL of blood was quickly collected from the heart and placed in equal amounts in red skull tubes (dry vacuum tubes without additives) and purple skull tubes (blood collection tubes containing ethylenediaminetetraacetic acid and its salts) and centrifuged at 2500 r / min for 10 minutes. Serum and plasma were aspirated with a pipette and packaged, and frozen at -80°C for later use. 3-4 cm of colon tissue was cut from 6-7 cm above the anal verge, and cut along the longitudinal axis of the intestinal tube, rinsed with physiological saline, and the intestinal mucosal morphology was observed. 1 cm of colon was cut and quickly fixed in paraformaldehyde fixative, and embedded in paraffin after 24 hours; another 1 cm of colon was quickly fixed in electron microscope solution at room temperature in the dark, and stored in a 4°C refrigerator after 2 hours. The remaining tissue was stored in a -80°C refrigerator for subsequent experiments.

[0101] 2.5 Experimental testing indicators

[0102] 2.5.1 Novelty-suppressed feeding experiment

[0103] Novelty suppression of food intake experiments were conducted on days 0, 21, and 35 of the experiment. Rats were fasted for 24 h before the experiment and placed in a 50 cm × 50 cm × 50 cm plastic box with an open top. The bottom of the box was covered with bedding and the same feed was placed. The latency of the rats to start eating was observed and calculated within 5 min. The criterion for starting to eat was when the rats started to chew the food.

[0104] 2.5.2 Elisa method for detection of D-lactic acid in plasma and TNF-α, IL-6, and IFN-γ in serum

[0105] According to the operating steps on the kit, complete sample addition, incubation-washing, enzyme addition-incubation-washing, color development-stopping, and OD value determination in sequence. Substitute the OD value of the sample into the linear regression equation of the standard curve (calculated based on the concentration of the standard and the OD value) to obtain the sample concentration × 5, which is the actual concentration of the sample.

[0106] 2.6 Results

[0107] 2.6.1 Comparison of novelty-suppressed food intake latency in rats of different groups

[0108] Before modeling, the novelty inhibition of food intake latency of rats in each group was compared. On day 0, there was no significant difference (P>0.05). Figure 3 At the end of modeling, compared with the blank group, the latency of novelty inhibition of food intake of rats in the modeling group was significantly prolonged (P<0.01), indicating that the modeling was successful; after treatment, on the 21st day, compared with the positive group, the comparative example 1 group and the comparative example 2 group, the latency of novelty inhibition of food intake of rats in the Example 1-3 group was significantly shortened (P<0.05 or P<0.01), see Figure 4 After treatment, on the 35th day, the novelty inhibition of food intake latency of rats in Example 1-3 groups was significantly shortened compared with the positive group, Comparative Example 1 group and Comparative Example 2 group (P<0.01), as shown in Figure 5 2.6.2 Comparison of D-lactic acid in rat plasma and TNF-α, IFN-γ, and IL-6 in serum

[0109] Compared with the blank group, the D-lactic acid in the plasma and the TNF-α, IFN-γ, and IL-6 content in the serum of the rats in the model group increased significantly (P < 0.05, P < 0.01). Compared with the model group and the pinaverium bromide group, the D-lactic acid and the TNF-α, IFN-γ, and IL-6 content in the serum of the Example 1-3 groups decreased significantly (P < 0.05, P < 0.01). Figure 6-9 . DETAILED DESCRIPTION

[0111] The present invention discloses an infant formula milk powder capable of improving the immunity of the body and a preparation method thereof. Those skilled in the art can refer to the content of this article and appropriately change the process parameters. It should be particularly noted that all similar replacements and modifications are obvious to those skilled in the art, and they are all considered to be included in the present invention. The method and application of the present invention have been described through a comparative embodiment, and relevant personnel can obviously modify or appropriately change and combine the method and application described herein without departing from the content, spirit and scope of the present invention to implement and apply the technology of the present invention.

[0112] Example: Preparation of Rosa Laevigata Extract

[0113] Method 1: Preparation of Rosa Laevigata Extract

[0114] 1) Take 500 g of Rosa laevigata fruit pulp powder, add 30% sodium chloride and soak for 30 minutes, then heat to 70° C., extract the medicinal liquid, and set aside the medicinal residue and the extract;

[0115] 2) Repeat step 1) with the medicinal residue and extract twice; combine the filtrates from the three extractions;

[0116] 3) collecting the filtrate from step 2), concentrating under reduced pressure at 50-60° C. to an alcohol content of 80%; standing at room temperature for 24 hours, centrifuging, collecting the precipitate and supernatant, and the precipitate is the Rosa laevigata extract;

[0117] 4) The protein of the Rosa laevigata extract obtained in step 3) was removed by Sevag method, centrifuged, and the supernatant was concentrated. The mixed Rosa laevigata extract was added with 95% ethanol in a volume ratio of 1:4, placed in a refrigerator (4°C) and allowed to stand for 24 hours overnight, centrifuged at 4000 r / min for 15 minutes, and the precipitate was collected and dissolved with an appropriate amount of deionized water to obtain 243.23 g of Rosa laevigata extract.

[0118] Method 2: Preparation of Rosa Laevigata Extract

[0119] 1) taking 500 g of Rosa laevigata fruit pulp powder, adding 60% sodium chloride and soaking for 50 minutes, then heating to 80° C., extracting the medicinal liquid, and setting aside the medicinal residue and the extract;

[0120] 2) Repeat step 1) with the residue and extract twice; combine the filtrates from the three extractions;

[0121] 3) collecting the filtrate from step 2), concentrating under reduced pressure at 50-60° C. to an alcohol content of 90%; standing at room temperature for 24 hours, centrifuging, collecting the precipitate and supernatant, and the precipitate is the Rosa laevigata extract;

[0122] 4) The protein of the Rosa laevigata extract obtained in step 3) was removed by Sevag method, centrifuged, and the supernatant was concentrated. The mixed Rosa laevigata extract was added with 95% ethanol in a volume ratio of 1:4, placed in a refrigerator (4°C) for 24 hours overnight, centrifuged at 4000 r / min for 15 minutes, and the precipitate was collected and dissolved with an appropriate amount of deionized water to obtain 239.67 g of Rosa laevigata extract.

[0123] Method 3: Preparation of Rosa Laevigata Extract

[0124] 1) taking 500 g of Rosa laevigata fruit pulp powder, adding 45% sodium chloride and soaking for 40 minutes, then heating to 75° C., extracting the medicinal liquid, and setting aside the medicinal residue and the extract;

[0125] 2) Repeat step 1) with the residue and extract twice; combine the filtrates from the three extractions;

[0126] 3) collecting the filtrate from step 2), concentrating under reduced pressure at 50-60° C. to an alcohol content of 85%; standing at room temperature for 24 hours, centrifuging, collecting the precipitate and supernatant, and the precipitate is the Rosa laevigata extract;

[0127] 4) The protein of the Rosa laevigata extract obtained in step 3) was removed by Sevag method, centrifuged, and the supernatant was concentrated. The mixed Rosa laevigata extract was added with 95% ethanol in a volume ratio of 1:4, placed in a refrigerator (4°C) for 24 hours overnight, centrifuged at 4000 r / min for 15 minutes, and the precipitate was collected and dissolved with an appropriate amount of deionized water to obtain 251.88 g of Rosa laevigata extract.

[0128] Method 4: Preparation of Rosa Laevigata Extract

[0129] 1) Take 500 g of Rosa laevigata fruit pulp powder, soak it in water for 40 minutes, then heat it to 75° C., extract the medicinal liquid, and keep the medicinal residue and the extract for later use;

[0130] 2) Repeat step 1) with the residue and extract twice; combine the filtrates from the three extractions;

[0131] 3) collecting the filtrate from step 2), concentrating under reduced pressure at 50-60° C. to an alcohol content of 85%; standing at room temperature for 24 hours, centrifuging, collecting the precipitate and supernatant, and the precipitate is the Rosa laevigata extract;

[0132] 4) The protein of the Rosa laevigata extract obtained in step 3) was removed by Sevag method, centrifuged, and the supernatant was concentrated. The mixed Rosa laevigata extract was added with 50% ethanol at a volume ratio of 1:7.6, placed in a refrigerator (4°C) and allowed to stand for 24 hours overnight, centrifuged at 4000 r / min for 15 minutes, and the precipitate was collected and dissolved with an appropriate amount of deionized water to obtain 187.40 g of Rosa laevigata extract.

[0133] Example 1: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0134] Pinaverium bromide 10g, Rosa laevigata extract 0.5g, sodium carboxymethyl starch 2g, magnesium stearate 0.1g, microcrystalline cellulose 20g, lactose 20g.

[0135] Preparation process:

[0136] 1) dissolving glyceryl monostearate in a mixed solution of chloroform and 95% or more ethanol, stirring constantly and mixing; wherein the weight volume ratio of glyceryl monostearate is 1:100, and the volume ratio of chloroform to ethanol is 2:1;

[0137] 2) adding pinaverium bromide to step 1) and mixing, stirring to dissolve;

[0138] 3) Add the Rosa Laevigata extract evenly into the mixed solution of step 2) and keep the temperature at 40-50°C

[0139] The mixture was evaporated to dryness under reduced pressure, microcrystalline cellulose, lactose and sodium carboxymethyl starch were added and blended, the mixture was sieved, magnesium stearate was added and tabletted to obtain plain tablets;

[0140] 4) Weigh 10 mg of coating premix powder and add 40 mg of water to make a film coating premix, and film coat the above plain tablets. The coating weight gain is 3-10%, and then dried. The material temperature is set to 35° C. and continued to dry until the free moisture is below 3%, to obtain tablets containing pinaverium bromide.

[0141] Example 2: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0142] Pinaverium bromide 10g, Rosa laevigata extract 1.5g, sodium carboxymethyl starch 10g, magnesium stearate 1.1g, microcrystalline cellulose 60g, lactose 70g.

[0143] The preparation process is the same as that of Example 1.

[0144] Example 3: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0145] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0146] The preparation process is the same as that of Example 1.

[0147] Example 4: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0148] Pinaverium bromide 10g, Rosa laevigata extract 1g, low-substituted hydroxypropyl cellulose 3g, talc 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0149] The preparation process is the same as that of Example 1.

[0150] Example 5: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0151] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, micro powder silica magnesium 0.6g, microcrystalline cellulose 40g, mannitol 45g.

[0152] The preparation process is the same as that of Example 1.

[0153] Example 6: A Pinaverium Bromide Tablet and Its Preparation Method

[0154] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, micronized silica magnesium 0.6g, sorbitol 40g, lactose 45g.

[0155] The preparation process is the same as that of Example 1.

[0156] Comparative Example 1: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0157] Pinaverium bromide 10g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0158] The preparation process is the same as that of Example 1.

[0159] Comparative Example 2: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0160] Pinaverium bromide 10g, Rosa laevigata extract 2g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0161] The preparation process is the same as that of Example 1.

[0162] Comparative Example 3: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0163] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 85g.

[0164] The preparation process is the same as that of Example 1.

[0165] Comparative Example 4: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0166] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, sucrose 45g.

[0167] The preparation process is the same as that of Example 1.

[0168] Comparative Example 5: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0169] Pinaverium bromide 10g, Rosa laevigata extract 1g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0170] Preparation process:

[0171] Add pinaverium bromide and Rosa laevigata extract to a mixed solution of ethanol with a concentration of more than 95%, and stir continuously to mix well; maintain the temperature at 40-50°C and perform reduced pressure rotary evaporation to dryness, add microcrystalline cellulose, lactose, and sodium carboxymethyl starch, blend them, sieve them, add magnesium stearate, and press them into tablets to obtain plain tablets; weigh 10 mg of coating premix powder, add 40 mg of water and mix them to prepare a film coating premix, film-coat the above plain tablets, dry them, set the material temperature to 35°C, and continue drying until the free moisture is below 3%, to obtain tablets containing pinaverium bromide.

[0172] Comparative Example 6: A Pinaverium Bromide Tablet and a Preparation Method Thereof

[0173] Pinaverium bromide 10g, Panax notoginseng polysaccharide 1g, sodium carboxymethyl starch 3g, magnesium stearate 0.6g, microcrystalline cellulose 40g, lactose 45g.

[0174] The preparation process is the same as that of Example 1.

[0175] 1) Dissolution of Pinaverium Bromide Tablets and Determination of Related Substances

[0176] 1.1 Dissolution test and results of pinaverium bromide tablets

[0177] This experimental example investigates the dissolution changes of commercially available pinaverium bromide tablets (Beijing Fuyuan Pharmaceutical Co., Ltd., batch number national medicine standard H20133036), Examples 1-6 and Comparative Examples 1-6 containing pinaverium bromide tablets under pH 1.0 HCl solution conditions.

[0178] Specific dissolution determination method: pH 1.0 HCl solution is used as the medium, and the dissolution is carried out at a rotation speed of 50 rpm / min. The ultraviolet spectrophotometry method is used for detection, and the detection wavelength is 243nm (based on pinaverium bromide). The test results are shown in Figure 1 .

[0179] According to the conventional requirements of tablets, the dissolution rate should be above 80% within 15 minutes. Figure 1 It can be seen that the dissolution of Examples 1-5 in the HCl solution at pH 1.0 was above 99% at 0, 1, 2, 3, and 6 months of the accelerated test, and the tablets containing pinaverium bromide prepared in Examples 1-5 all met the dissolution requirements of the Chinese Pharmacopoeia. However, in the accelerated test, the dissolution of Comparative Examples 1, 3, 4, 5, and 6 at 15 minutes was lower than 85%, and Comparative Examples 3-5 were lower than 80%, which could not meet the requirements of the Pharmacopoeia.

[0180] 1.2 Determination of related substances in pinaverium bromide tablets and results

[0181] Determination method of related substances: Determined by high performance liquid chromatography, refer to the 2020 edition of the Chinese Pharmacopoeia

[0182] Chromatographic conditions: octadecylsilane bonded silica gel as filler, acetonitrile-sodium dodecyl sulfate buffer (70:30) as mobile phase, detection wavelength of 245nm, column temperature of 25℃, flow rate of 1.0mL / min, injection volume of 20μL. Preparation method of sodium dodecyl sulfate buffer: Take 13.0g sodium dodecyl sulfate, add 1500mL of water to dissolve, add 5.0mL of phosphoric acid, mix well, adjust pH to 2.5 with triethylamine, and filter to obtain.

[0183] Depend on Figure 2It can be seen that compared with comparative examples 1-6, the total impurity content of the tablets of embodiments 1-6 of the present invention in the first accelerated test is substantially unchanged, while the total impurity content of comparative examples 1, 2, 3, and 5 is significantly increased, indicating that the stability of the tablets containing pinaverium bromide prepared by the method of the present invention is significantly improved, wherein the stability of embodiment 3 is the best. Comparative example 1 does not contain the extract of the fruit of golden rosa laevigata, and the extract is particularly important for the stability of pinaverium bromide. The inventor believes that polysaccharides can form a protective film on the surface of the drug. This film can isolate the contact of drug molecules with the external environment, slow down the influence of moisture, oxygen and light on pinaverium bromide, thereby reducing or avoiding the rate of hydrolysis reaction, thereby improving the stability of the drug. The extract of the fruit of golden rosa laevigata of comparative example 2 is too high, and other substances inside may react with pinaverium bromide to cause degradation, and the different adjuvants used in comparative example 3 also have an impact on the final product, and comparative example 5 uses different preparation processes, and the difference in preparation processes also has an impact on pinaverium bromide, and pinaverium bromide is relatively sensitive to light and temperature, and does not take relevant means to accelerate its degradation.

[0184] 2.1 Modeling and grouping

[0185] 2.1.1 Modeling

[0186] A total of 72 SPF Wistar female rats were adaptively fed for 1 week, and 8 rats were randomly selected as the blank group using a random number table. The remaining rats were treated with a compound modeling method of intestinal perfusion of acetic acid + tail clamp stress + mold restriction to prepare a diarrhea-predominant irritable bowel syndrome model.

[0187] From day 1 to day 14, all rats were fasted 12 hours before enema. 3% sodium pentobarbital was injected intraperitoneally at 50 mg / kg at 8:30 every day for anesthesia. After anesthesia, an 8F catheter lubricated with paraffin oil was inserted 8 cm through the anus. 1 mL of 4% acetic acid was injected into the model rats. After 30 seconds, an equal amount of phosphate buffered saline was injected. The blank group was injected with 1 mL of saline. From day 7 to day 21, the model rats were restrained in a mold for 1 hour at 15:00 every day, and then the plastic clip was clamped at the 1 / 3 of the rat's tail for 1 hour. The blank group was not intervened. The significant increase in fecal water content was regarded as the standard for successful modeling (the fecal water content of the model group and the blank group was significantly different P<0.01, indicating that the modeling was successful).

[0188] 2.1.2 Grouping

[0189] The rats were divided into 5 groups according to the random number table: model group, positive control group, Example 1-3 groups, Comparative Example 1 group, Comparative Example 6 group, 8 rats in each group, and a blank group with no treatment.

[0190] 2.3 Administration

[0191] The pinaverium bromide tablets and positive control drugs prepared in each example were ground into 50 mg of pinaverium bromide and dissolved in 25 mL of distilled water to prepare a 2.50 mg / mL suspension. The above examples and suspensions prepared as needed were placed in a 4°C refrigerator for standby use and gavaged twice a day in the morning and evening; the positive control group and each example group were given 5.40 mL·kg of pinaverium bromide suspension -1 ·d -1 (i.e. 13.50 mg kg -1 ), once a day by gavage; the blank group and the model group were given 10 mL·kg of normal saline -1 ·d -1 The drug was administered by intragastric gavage for 14 consecutive days.

[0192] 2.4 Specimen collection and processing

[0193] After the last administration of each group, the rats were fasted but not watered for 24 hours. After weighing their body weight, they were anesthetized by intraperitoneal injection of 3% sodium pentobarbital at 50 mg / kg. About 4 mL of blood was quickly collected from the heart and placed in equal amounts in red skull tubes (dry vacuum tubes without additives) and purple skull tubes (blood collection tubes containing ethylenediaminetetraacetic acid and its salts) and centrifuged at 2500 r / min for 10 minutes. Serum and plasma were aspirated with a pipette and packaged, and frozen at -80°C for later use. 3-4 cm of colon tissue was cut from 6-7 cm above the anal verge, and cut along the longitudinal axis of the intestinal tube, rinsed with physiological saline, and the intestinal mucosal morphology was observed. 1 cm of colon was cut and quickly fixed in paraformaldehyde fixative, and embedded in paraffin after 24 hours; another 1 cm of colon was quickly fixed in electron microscope solution at room temperature in the dark, and stored in a 4°C refrigerator after 2 hours. The remaining tissue was stored in a -80°C refrigerator for subsequent experiments.

[0194] 2.5 Experimental Results

[0195] 2.5.1 Behavioral experiment (novelty suppression of food intake experiment)

[0196] Novelty suppression of feeding experiments were conducted on days 0, 21, and 35 of the experiment. Rats were fasted for 24 h before the experiment and placed in a 50 cm × 40 cm × 30 cm plastic box with an open top. The bottom of the box was covered with bedding, and several grains of feed of the same size, quantity, and direction were placed in a fixed position. The latency period for the rats to start eating within 5 min was observed and calculated. The criterion for starting to eat was when the rats started to chew the food.

[0197] 2.5.2 Elisa method for detection of D-lactic acid in plasma and TNF-α, IL-6, and IFN-γ in serum

[0198] According to the operating steps on the kit, complete sample addition, incubation-washing, enzyme addition-incubation-washing, color development-stopping, and OD value determination in sequence. Substitute the OD value of the sample into the linear regression equation of the standard curve (calculated based on the concentration of the standard and the OD value) to obtain the sample concentration × 5, which is the actual concentration of the sample.

[0199] 2.6 Results 2.6.1

[0201] Before modeling, the novelty inhibition of food intake latency of rats in each group was compared. On day 0, there was no significant difference (P>0.05). Figure 3 At the end of modeling, compared with the blank group, the latency of novelty inhibition of food intake of rats in the modeling group was significantly prolonged (P<0.01), indicating that the modeling was successful; after treatment, on the 21st day, compared with the positive group, the comparative example 1 group and the comparative example 2 group, the latency of novelty inhibition of food intake of rats in the Example 1-3 group was significantly shortened (P<0.05 or P<0.01), see Figure 4 After treatment, on the 35th day, the novelty inhibition of food intake latency of rats in Example 1-3 groups was significantly shortened compared with the positive group, Comparative Example 1 group and Comparative Example 2 group (P<0.01), as shown in Figure 5 . 2.6.2

[0203] Compared with the blank group, the D-lactic acid in the plasma and the TNF-α, IFN-γ, and IL-6 content in the serum of the rats in the model group increased significantly (P < 0.05, P < 0.01). Compared with the model group and the pinaverium bromide group, the D-lactic acid and the TNF-α, IFN-γ, and IL-6 content in the serum of the Example 1-3 groups decreased significantly (P < 0.05, P < 0.01). Figure 6-9 .

Claims

1. A tablet containing pinaverium bromide, characterized in that: The pinaverium bromide tablet is composed of the following raw materials: 10 parts by weight of pinaverium bromide, 0.5-1.5 parts by weight of Rosa laevigata extract, 2-10 parts by weight of disintegrant, 0.1-1.1 parts by weight of lubricant, and 40-130 parts by weight of filler.

2. The Pinaverium Bromide tablet according to claim 1, characterized in that The pinaverium bromide tablet is composed of the following raw materials: 10 parts by weight of pinaverium bromide, 1 part by weight of Rosa laevigata extract, 3 parts by weight of disintegrant, 0.6 parts by weight of lubricant, and 85 parts by weight of filler.

3. The Pinaverium Bromide tablet according to claim 1, characterized in that The filler is selected from one or two of microcrystalline cellulose, lactose, mannitol, and starch; more preferably, the filler is selected from a combination of microcrystalline cellulose and lactose.

4. The Pinaverium Bromide tablet according to claim 3, characterized in that Calculated by weight, the ratio of microcrystalline cellulose to lactose is 2-6:2-7; more preferably, the ratio of microcrystalline cellulose to lactose is 8:

9.

5. The Pinaverium Bromide tablet according to claim 1, characterized in that The disintegrant is selected from one of low-substituted hydroxypropyl cellulose and sodium carboxymethyl starch, preferably sodium carboxymethyl starch; the lubricant is selected from any one of magnesium stearate, talc, and sodium dodecyl sulfate, preferably magnesium stearate.

6. The Pinaverium Bromide tablet according to claim 1, characterized in that The pinaverium bromide tablet is composed of the following raw materials: 10 parts by weight of pinaverium bromide, 0.5-1.5 parts by weight of Rosa laevigata extract, 2-10 parts by weight of sodium carboxymethyl starch, 0.1-1.1 parts by weight of magnesium stearate, 20-60 parts by weight of microcrystalline cellulose, and 20-70 parts by weight of lactose.

7. The Pinaverium Bromide tablet according to claim 1, characterized in that The pinaverium bromide tablet is composed of the following raw materials: 10 parts by weight of pinaverium bromide, 1 part by weight of Rosa laevigata extract, 3 parts by weight of sodium carboxymethyl starch, 0.6 parts by weight of magnesium stearate, 40 parts by weight of microcrystalline cellulose, and 45 parts by weight of lactose.

8. The Pinaverium Bromide tablet according to claim 1, characterized in that The pinaverium bromide tablets may be coated.

9. The Pinaverium bromide tablet according to any one of claims 1-2 or 6-7, characterized in that The extraction method of the Rosa Laevigata extract is: 1) Take the fruit powder of Rosa laevigata and add 30-60% sodium chloride to soak for 30-50 minutes, then heat to 70-80° C. to extract the medicinal liquid, and keep the residue and the extract for later use; 2) Repeat step 1) with the residue and extract twice; combine the filtrates from the three extractions; 3) collecting the filtrate from step 2), concentrating under reduced pressure at 50-60° C. to an alcohol content of 80-90%; standing at room temperature for 24 hours, centrifuging, collecting the precipitate and supernatant, and the precipitate is the Rosa laevigata extract; 4) Deproteinizing the polysaccharide extract obtained in step 3) by Sevag method, centrifuging, concentrating the supernatant, precipitating with ethanol, collecting the precipitate, and vacuum drying to obtain the Rosa laevigata extract.

10. The Pinaverium bromide tablet according to claim 8, characterized in that: The preparation method of the pinaverium bromide tablets: 1) Dissolve glyceryl monostearate in a mixed solution of chloroform and 95% or more ethanol, stirring continuously to mix well; 2) adding pinaverium bromide to step 1), mixing and stirring to dissolve; 3) adding the Rosa laevigata extract evenly to the mixed solution of step 2), maintaining the temperature at 40-50° C. and performing rotary evaporation under reduced pressure to dryness, adding microcrystalline cellulose, lactose, and sodium carboxymethyl starch, blending, sieving, adding magnesium stearate, and tableting to obtain plain tablets; 4) Weigh the coating premix powder, add water and mix to prepare a film coating premix, apply film coating to the above plain tablets, the coating weight gain is 3-10%, and dry to obtain tablets containing pinaverium bromide.

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