Method for detecting content of carbomer in medicinal preparation

Abstract

The invention relates to a method for detecting the content of carbomer in a medicinal preparation, which is used for solving the problems of high detection cost and auxiliary material interference in the prior art. According to the method, urea and guanidine organic alkali are added in sample pretreatment, so that the influence of preparation auxiliary materials on the neutralization reaction of carbomer is weakened; according to the method, excessive berberine is adopted to be fully combined with carboxylic acid groups in the carbomer, then the residual concentration of the berberine is detected, and a linear relation of the reaction of the carbomer and the berberine is established, so that the content of the carbomer in a sample is calculated. The detection method established by the invention is suitable for different preparation formulas, is more flexible in detection means, and is suitable for industrial popularization.

Description

Technical Field

[0002] This invention belongs to the field of analytical chemistry technology, and specifically relates to a method for detecting carbomer content in pharmaceutical preparations. Background Technology

[0004] Carbomer is a high molecular weight polymer formed by crosslinking polyalkyl sucrose or polyalkyl pentaerythritol with acrylic acid. Different combinations of substances and degrees of polymerization result in products with different properties and efficacy. Carbomer has strong hygroscopicity and is highly electrostatic. Due to the presence of 52%–68% carboxylic acid groups in its internal structure, it is acidic. Before contact with water, the polyacrylic acid in carbomer is coiled; when its powder is mixed with water, the coiled molecular chains slowly unwind. In an aqueous dispersion with a pH of 2.5–3.0, carbomer can be neutralized with alkaline substances, causing the carboxyl groups of carbomer to ionize and subsequently form a gel.

[0005] Carbomer possesses excellent gelling, emulsifying, solubilizing, thickening, suspending, and film-forming properties, exhibits no allergic reactions, and demonstrates stable and safe efficacy. It exhibits good compressibility under low pressure, high bioavailability, and strong in vitro-in vivo correlation, making it suitable for various formulation types, including gels, semi-solid dosage forms, solid dosage forms, and bioadhesives. Existing methods for determining carbomer content include acid-base titration, drying and weighing, and high-performance liquid chromatography (HPLC).

[0006] CN117630230A, "A Method for Determining Carbomer Content in Preparations by Evaporative Light Scattering in High Performance Liquid Chromatography" (Application Date: 2023.12.21; Publication Date: 2024.03.01), discloses the chromatographic conditions for the carbomer HPLC-ELSD method: an octadecylsilane-bonded silica column is used as the chromatographic column; water or 0.02% formic acid aqueous solution is used as mobile phase A; acetone is used as mobile phase B; and ethanol is used as mobile phase C, with gradient elution; the drift tube temperature of the evaporative light scattering detector is 45℃~65℃, and the nitrogen flow rate is 1.8~3.0 L / min. This technical solution utilizes the linear relationship between the logarithm of the peak area of ​​carbomer in the evaporative light scattering detector and the logarithm of its concentration to establish a standard curve, thereby achieving quantitative detection. However, this method requires sophisticated detection equipment and has high detection costs.

[0007] CN120293753A, "A Method for Determining Carbomer Content in Eye Drops" (Application Date: 2025.01.13; Publication Date: 2025.07.11), discloses a method for separating carbomer and brinzolamide from other excipients in eye drops through a two-stage washing and centrifugation process. The weights of carbomer and brinzolamide are obtained by drying and weighing, and then the weight of brinzolamide is subtracted to determine the carbomer content in the sample. However, the washing and centrifugation step in this method cannot eliminate the interference of other polymer excipients on the detection of carbomer content.

[0008] Acid-base titration primarily utilizes the reaction of a strong base with the carboxylic acid groups in carbomer. CN115015143A, "An Analytical Method for Carbomer Homopolymer in Transdermal Absorption Preparations" (Application Date: 2022.08.08; Publication Date: 2022.09.06), employs sodium hydroxide to dissociate the free carboxyl groups of carbomer homopolymer into carboxylate ions. Calcium ions then combine with these carboxylate ions to form a complex, thereby extracting and separating the carbomer. Flame atomic absorption spectrophotometry is used to detect the uncomplexed calcium, and the carbomer homopolymer content is calculated from the complexed calcium content. This method uses uncommon and expensive instruments, resulting in high testing costs; sample pretreatment time exceeds 2 hours, and the testing time is long.

[0009] CN112461826A, "A Method for Determining Carbomer Content in Carbomer-Containing Gels" (Application Date: 2017.06.29; Publication Date: 2021.03.09), uses a strong alkaline solution to ensure the carboxylic acid groups of carbomer in the gel exist in an ionic state. An excess metal salt solution is then added to allow the metal salt to fully combine with the carboxylic acid groups in the carbomer, forming a complex. After removing the complex, the unbound metal salt is titrated with a complexometric titrant to calculate the carbomer content in the sample. The disadvantages of the back titration method are that metal ion raw materials or excipients (such as EDTA) in the formulation can interfere with the detection results; moreover, the sample pretreatment time also exceeds 2 hours, resulting in a long detection time. Summary of the Invention

[0011] The purpose of this invention is to provide a method for detecting carbomer content in pharmaceutical preparations, addressing the problems of high detection costs and excipient interference in existing technologies, and providing accurate and stable results. This invention uses excess berberine to fully bind with the carboxylic acid groups in carbomer, then detects the remaining concentration of berberine, thereby calculating the carbomer content in the sample.

[0012] After in-depth research on carbomer formulations, the inventors discovered that pharmaceutical excipients are the core issue interfering with the reaction between berberine and the carboxylic acid group in carbomer. Eliminating interference from other excipients is the key step in solving the technical solution of this invention.

[0013] Furthermore, berberine is an organic alkaloid with relatively weak basicity, and the resulting ion pairs mainly exist in the form of weak ion pairs. The existing technology, "Preparation of Berberine-Carbomer Salt Complex and Evaluation of its Gastric Bioadhesiveness and Anti-Hp Activity," points out that carbomer is a common bioadhesive material. Even when berberine-carbomer salt is produced through neutralization, some carboxylic acid groups remain unreacted, and the remaining carboxyl groups ensure that berberine-carbomer salt still possesses strong gastrointestinal bioadhesive properties. The weak basicity of berberine itself is the reason why some carboxyl groups in carbomer do not participate in the reaction. In addition, CN115015143A and CN112461826A, when using a strong base as an acid-base titrant, require approximately 2 hours of water bath heating to ensure complete reaction between carbomer and the strong base. Therefore, when using berberine as a titrant for carbomer determination, it is necessary to address the issue of establishing a linear relationship between the amount of carbomer added and berberine.

[0014] This invention is achieved using the following technical solution:

[0015] A method for detecting carbomer content in a pharmaceutical preparation, comprising the following steps:

[0016] (1) Preparation of blank test sample: Take a drug preparation that does not contain carbomer, add 2 to 8 times the amount of urea or guanidine organic base solution, heat to dissolve, centrifuge, and take the lower layer solution as the blank test sample solution;

[0017] (2) Preparation of linear sample: Add carbomer solution to blank test sample solution, add berberine, let stand, centrifuge and take the supernatant to obtain linear sample; the ratio of berberine to carbomer is not less than 18:1;

[0018] (3) Standard curve plotting: The berberine content of the linear sample solution was detected, and the berberine consumption of the linear sample was determined by comparing the amount of berberine added in step (2). A standard curve of carbomer addition and berberine consumption was established.

[0019] (4) Test sample determination: The test drug preparation is processed according to the method in step (1); berberine is added, and the supernatant is taken by centrifugation. The amount of berberine used is more than 18 times the carbomer content in the test drug preparation; the berberine content is detected, and the carbomer content is calculated using the standard curve in step (3).

[0020] Furthermore, in step (1), the amount of urea or guanidine organic base used is 4 times the amount of the drug preparation.

[0021] Furthermore, the guanidine organic bases are selected from one or more of guanidine hydrochloride, guanidine sulfate, guanidine carbonate, guanidine nitrate, and guanidine phosphate.

[0022] Furthermore, in step (2), the ratio of berberine to carbomer is 18 to 42:1; preferably, the ratio of berberine to carbomer is 20:1. The standing time in step (2) is 10 to 20 minutes.

[0023] Furthermore, in step (1), the heating is carried out in a water bath for 10 to 20 minutes, and the centrifugation speed is 10,000 rpm to 14,000 rpm.

[0024] Furthermore, the method for detecting berberine content is selected from one of the following: high performance liquid chromatography, ultra-high performance liquid chromatography, ultraviolet-visible spectrophotometry, and fluorescence spectroscopy.

[0025] Furthermore, the chromatographic column used in the high-performance liquid chromatography is an octadecylsilane-bonded silica gel packed column, mobile phase A is a 0.4% triethylamine solution with a pH of 4.5, and mobile phase B is selected from acetonitrile or methanol.

[0026] Furthermore, the berberine content in the sample is calculated in steps (3) and (4) by high performance liquid chromatography or ultra-high performance liquid chromatography using the concentration and peak area of ​​berberine reference standard.

[0027] Furthermore, the berberine content detection method adopts ultraviolet-visible spectrophotometry, and the berberine content in the sample is calculated in steps (3) and (4) by using the concentration and absorbance of the berberine reference standard.

[0028] This invention has the following advantages:

[0029] (1) The present invention employs a specific pretreatment method to reduce the influence of pharmaceutical excipients on the quantitative detection of carbomer, rapidly establish a linear relationship between the reaction of carbomer and berberine, and realize the quantitative detection of carbomer.

[0030] Existing technologies indicate that the yield of berberine-carbomer salt [precipitate dry mass / (carbomer feed amount + berberine feed amount) * 100%] is not linearly related to the increase of berberine content; the drug loading (berberine content in precipitate / precipitate mass * 100%) increases with the increase of berberine dosage, but it is difficult to achieve a high drug loading due to the incomplete reaction of carboxylic acid groups in carbomer.

[0031] Pharmaceutical excipients are numerous and complex in composition. Examples 2 and 4, corresponding to the scope of this invention, weaken the influence of pharmaceutical excipients on the carbomer neutralization reaction by adding urea and guanidine hydrochloride during sample pretreatment; simultaneously, they alter the reaction system of berberine and carbomer to ensure sufficient reaction and a stable drug loading. The resulting carbomer quantification method exhibits good linearity (R0). 2>0.99); In addition, the acid-base reaction water bath time in the method of the present invention is about 20 min, which greatly shortens the pretreatment time of the preparation sample compared with the prior art, and helps to quickly establish a quantitative detection method.

[0032] Different pretreatment methods, such as the use of ethyl acetate to treat the same cream in Example 3, cannot completely eliminate the interference of excipients on carbomer, directly affecting the content detection results and preventing the formation of a linear relationship. Combining Examples 2-4, it can be seen that specific pretreatment steps are crucial for the detection of carbomer content in formulations.

[0033] In the detection method of this invention, the linear range of carbomer content is 0.09%~0.21%. Within the linear range, the linear relationship between berberine consumption y and carbomer addition x is good, and the linear equation is y = 9.154x + 0.0156, R0 2 =0.9932.

[0034] (2) This invention establishes a carbomer content detection method that is applicable to different formulations, has more flexible detection methods, and is suitable for industrial promotion.

[0035] This invention uses ivermectin cream and diclofenac gel to study the effect of excipient differences on detection results. The experimental results of Example 10 show that this technical solution can achieve the detection of carbomer content in the above-mentioned pharmaceutical preparations, and the detection results are consistent with the drug prescription. The technical solution provided by this invention meets the detection requirements of carbomer content in pharmaceutical preparations with different excipient compositions, and has a wide range of applications and high accuracy.

[0036] Furthermore, this invention investigated the effect of different liquid phase conditions on the quantitative detection of different carbomer types. Examples 11-13 used different liquid phase conditions to detect the content of two carbomer types, 974P and 980NF. The experimental results showed that there was a linear relationship between the amount of different carbomer types added and the amount of berberine consumed under different liquid phase conditions (R0). 2 >0.99), which allows for quantitative analysis. Attached Figure Description

[0038] The accompanying drawings, which constitute a part of this specification, are provided to further illustrate the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an undue limitation of the invention.

[0039] Figure 1 Linearity results of carbomer TR-1 liquid chromatography detection in Example 8. The y-axis represents the amount of berberine consumed, and the x-axis represents the amount of carbomer added; the same applies below.

[0040] Figure 2 Linearity results of Carbomer 974P liquid phase detection in Example 11.

[0041] Figure 3 Linearity results of Carbomer 980NF liquid phase detection in Example 11. Detailed Implementation

[0043] The present invention will be further described below with reference to specific embodiments. These embodiments are provided to better illustrate the invention and are not intended to limit it. Unless otherwise specified, the experimental methods used in the embodiments are conventional methods; the materials and reagents used are commercially available unless otherwise specified. Therefore, non-essential improvements and adjustments made by those skilled in the art based on the above-described invention are still within the scope of protection of this invention.

[0044] Example 1: Preparation of blank cream

[0045] 1. Test materials

[0046] Table 1. Formula for Carbomer-Free Cream

[0047] prescription Dosage per gram of formulation (mg) Ivermectin 10 glycerin 40 Isopropyl palmitate 40 Dimethicone 5 Disodium edetate (EDTA) 0.5 Citric acid monohydrate 0.5 cetyl alcohol 35 Stearyl alcohol 25 Polyethylene glycol hexadecyl ether 30 Stearosyl sorbitan 20 Methylparaben 2 Propylparaben 1 Phenoxyethanol 10 Propylene glycol 20 Oil alcohol 20 10% sodium hydroxide Appropriate amount Purified water Increase to 1000mg

[0048] 2. Preparation method

[0049] Aqueous phase preparation: Mix the prescribed amounts of glycerol, EDTA, citric acid monohydrate, and methylparaben, heat to 65℃-70℃, and stir at 800 rpm until homogeneous.

[0050] Oil phase preparation: Take the prescribed amounts of hexadecyl alcohol, octadecyl alcohol, propylparaben, dimethicone, polyethylene glycol hexadecyl ether, isopropyl palmitate, and stearyl sorbitan, mix them, heat to 70℃-75℃, and stir at 400rpm to homogenize.

[0051] Preparation of active phase: Mix the prescribed amounts of phenoxyethanol, oleyl alcohol, and propylene glycol, stir at 500 rpm until homogeneous, then add the prescribed amount of ivermectin and stir until dissolved.

[0052] Cream preparation: Add the oil phase to the aqueous phase and stir at 900 rpm for 10 min at 70℃ until homogeneous; cool to 40℃ and add the active phase, stir at 900 rpm for 10 min; cool the cream to 30℃ at 700 rpm, add purified water to make up the volume, and adjust the pH value to 6.3±0.3 with 10% sodium hydroxide solution to obtain the final product.

[0053] Example 2: Detection Method 1

[0054] 1. Reagent preparation

[0055] Preparation of carbomer solution: Take an appropriate amount of carbomer TR-1, add water and let it swell overnight, add an appropriate amount of 1 mol / L sodium hydroxide solution (8 μL for every 1 mg of carbomer), dilute with water to the mark, and shake well to obtain the solution.

[0056] Preparation of berberine solution: Add water to berberine, sonicate to dissolve, and make up to a final volume to prepare a berberine solution with a concentration of 0.5 mg / mL.

[0057] Preparation of blank sample solution: Take 10g of blank cream, add 40g of urea, add 180mL of water, heat in a water bath for 10~20min, shake to dissolve, centrifuge at 12000 rpm for 15min, take the lower layer solution, repeat twice, and combine to obtain the blank sample solution.

[0058] Preparation of linear sample solutions: Weigh 3.0 mL, 4.0 mL, 5.0 mL, 6.0 mL, and 7.0 mL of 0.2 mg / mL carbomer solution respectively, and place each in a 100 mL volumetric flask. Add 25 mL of blank sample solution, and add 25 mL, 30 mL, 40 mL, 50 mL, and 60 mL of 0.5 mg / mL berberine solution respectively. After standing for 20 min, make up to volume, shake well, and centrifuge at 12000 rpm for 15 min to obtain the linear sample solution.

[0059] 2. Experimental Procedure

[0060] Chromatographic conditions: HPLC was performed using an Ultimate XB-C18 C18 column (150 mm × 4.6 mm, 5 µm); Mobile phase A: 1 L of purified water was mixed with 4 mL of triethylamine, and the pH was adjusted to 4.5 with glacial acetic acid. The mixture was then filtered. Mobile phase B: Acetonitrile; isocratic elution was performed between 70% mobile phase A and 30% mobile phase B. Flow rate: 1.3 mL / min; Detection wavelength: 235 nm; Column temperature: 30 °C; Injection volume: 10 μL.

[0061] The berberine content of the above linear samples was detected to determine the residual berberine content in the samples, thereby calculating the linear relationship between the amount of carbomer added (x) and the amount of berberine consumed (y).

[0062] 3. Test Results

[0063] The experimental results show that there is a linear relationship between the amount of carbomer added (x) and the amount of berberine consumed (y), which can be quantitatively analyzed.

[0064] Table 2 Linear Relationship Data for Detection Method 1

[0065]

[0066] Example 3: Detection Method 2

[0067] 1. Reagent preparation

[0068] Preparation of the formulation solution: Take five 1.0g blank creams and add them to 20mL volumetric flasks. Add 2.5mL, 5mL, 10mL, 15mL, and 17.5mL of 0.2mg / mL carbomer solution respectively. Make up to volume with purified water, extract with an equal volume of ethyl acetate, centrifuge, and take 2mL of the aqueous layer. Add 3mL of 0.5mg / mL berberine solution, shake manually for 2min, and let the precipitate form. Take the supernatant and dilute it 5 times with purified water to obtain the sample.

[0069] 2. Experimental Procedure

[0070] Berberine content was detected by high performance liquid chromatography according to Example 2, and the linear relationship between carbomer addition amount x and berberine consumption amount y was calculated.

[0071] 3. Test Results

[0072] The experimental results are shown in Table 3.

[0073] Table 3 Linear Relationship Data for Detection Method 2

[0074]

[0075] The results showed that the linear relationship between the berberine and carbomer consumed in the reaction was poor, making accurate quantification impossible. This may be because the formulation excipients are numerous and complex, and ethyl acetate treatment of the cream cannot completely eliminate the interference of excipients on carbomer, resulting in insufficient enrichment of carbomer and directly affecting the content detection results.

[0076] Based on the experimental results of Examples 2 and 3, it was found that urea reduced the influence of formulation excipients on the neutralization reaction of carbomer; at the same time, by changing the reaction system, berberine and carbomer reacted fully to form a stable drug loading, thereby making the carbomer dosage and berberine consumption linearly related, and realizing the quantitative determination of carbomer.

[0077] Example 4: Detection Method 3

[0078] 1. Reagent preparation

[0079] The preparation method of Example 2 was used, except that guanidine hydrochloride was used instead of urea in the preparation of the blank sample solution. The preparation method is as follows:

[0080] Take 10g of blank cream, add 40g of guanidine hydrochloride, add 180mL of water, heat in a water bath for 10-20min, shake to dissolve, centrifuge at 12000 rpm for 15min, take the lower layer solution, repeat twice, and combine to obtain the final product.

[0081] 2. Experimental Procedure

[0082] Berberine content was detected by high performance liquid chromatography according to Example 2, and a linear equation was established between the amount of carbomer added (x) and the amount of berberine consumed (y).

[0083] 3. Test Results

[0084] The experimental results are shown in Table 4.

[0085] Table 4 Linear Relationship Data for Detection Method 3

[0086]

[0087] The results show that when the cream is treated with guanidine hydrochloride, the linear relationship between the berberine consumed in the reaction and the added carbomer is good, allowing for quantitative analysis. The experimental results indicate that guanidine hydrochloride, like urea, can reduce the influence of pharmaceutical excipients on the carbomer neutralization reaction. The above sample pretreatment steps are a key step in the detection of carbomer content in pharmaceutical preparations.

[0088] Example 5: Investigation of Additive Dosage

[0089] 1. Reagent preparation

[0090] Preparation of blank sample solution: Take 10g of blank cream, add 40g of urea, add 180mL of water, heat in a water bath for 10~20min, shake to dissolve, centrifuge at 12000rpm for 15min, take the lower layer solution and put it into a 500mL volumetric flask, repeat twice, make up to volume, and shake well to obtain the solution.

[0091] Preparation of linear sample solutions: Refer to Example 2 for preparation.

[0092] Recovery rate sample preparation: Take 1g blank cream, add 2g, 4g, and 8g urea respectively, add 10mL of 0.2 mg / mL carbomer solution, add 25mL of water, heat in a water bath for 10-20min, shake to dissolve, centrifuge at 12000 rpm for 15min, take the lower layer solution and put it into a 50mL volumetric flask, repeat twice, make up to volume and shake well; take 25mL of the above solution, add 40mL of 0.5 mg / mL berberine solution, let stand for 20min, make up to volume, shake well, centrifuge at 12000 rpm for 15min to obtain the recovery rate sample.

[0093] 2. Experimental Procedure

[0094] Berberine content was detected by high performance liquid chromatography (HPLC) according to Example 2, and the linear relationship between carbomer addition amount x and berberine consumption amount y was calculated. The recovery rate of berberine consumption in the sample was detected by HPLC, and the amount of carbomer measured was calculated by linear equation.

[0095] 3. Test Results

[0096] Table 5 Results of the study on additive dosage

[0097]

[0098] The experimental results show that the urea dosage corresponding to each gram of cream is between 2 and 8g, and the average method recovery rate is between 80% and 120%, all of which meet the requirements. The dosage of 4g of urea is the preferred one.

[0099] Example 6: Investigation of Berberine Dosage

[0100] 1. Reagent preparation

[0101] The preparation method of Example 2 was used, the difference being the amount of carbomer used in the preparation of the linear sample solution.

[0102] Preparation of linear sample solutions: Weigh 3 mL, 4.0 mL, 5.0 mL, 6.0 mL, and 7.0 mL of 0.2 mg / mL carbomer solution, and add 25 mL of blank sample solvent to each solution. Prepare a total of 3 groups in this manner.

[0103] Take 20 mL of 0.5 mg / mL berberine solution from the first group of samples and add it to each sample.

[0104] Take 50 mL of 0.5 mg / mL berberine solution from the second group of samples;

[0105] Take the third group of samples and add 25 mL, 30 mL, 40 mL, 50 mL and 60 mL of 0.5 mg / mL berberine solution respectively; after the three groups of samples stand for 20 min, make up the volume, shake well, and centrifuge at 12000 r / min for 15 min to obtain the sample.

[0106] 2. Experimental Procedure

[0107] Berberine content was detected by high performance liquid chromatography according to Example 2, and a linear equation was established between the amount of carbomer added (x) and the amount of berberine consumed (y).

[0108] 3. Test Results

[0109] Table 6 Results of the investigation of different berberine dosages

[0110]

[0111] The experimental results are shown in Table 6. When the amount of berberine added is 7-16 times that of carbomer, the linear relationship between the amount of berberine consumed in the reaction and the amount of carbomer added is poor, R 2Less than 0.99. The carboxylic acid groups in carbomer range from 52% to 68%, resulting in varying drug loading and yields during the reaction with berberine. Existing techniques indicate that the yield of berberine-carbomer salt [precipitate dry mass / (carbomer feed amount + berberine feed amount) * 100%] is not linearly related to increasing berberine content; the drug loading (berberine content in the precipitate / precipitate mass * 100%) increases with increasing berberine dosage, but high drug loading is difficult to achieve due to incomplete reaction of the carboxylic acid groups in the carbomer. Experimental results show that this technique, by adding urea and guanidine hydrochloride during sample pretreatment to alter the reaction system between berberine and carbomer, achieves a stable drug loading when the amount of berberine added is 18-42 times that of carbomer. The resulting quantitative method for carbomer exhibits good linearity. To improve the reproducibility of the method, the ratio of berberine to carbomer was kept constant at around 20 times during the linear study.

[0112] Example 7: Investigation of Carbomer Dosage

[0113] 1. Reagent preparation

[0114] Preparation of blank sample solution: Weigh 10g of the blank cream prepared in Example 1, add 40g of guanidine hydrochloride, add 180mL of water, heat in a water bath for 10~20min, shake to dissolve, centrifuge at 12000rpm for 15min, take the lower layer solution, repeat twice, make up to volume, and shake well to obtain the solution.

[0115] Linear sample preparation: Weigh 1.5 mL, 2.0 mL, 2.5 mL, 3.0 mL, and 3.5 mL of 0.1 mg / mL carbomer solution respectively, and add 6.25 mL of blank sample solution to each; add 6.25 mL, 7.5 mL, 10 mL, 12.5 mL, and 15 mL of 0.5 mg / mL berberine solution respectively, let stand for 20 min, then make up to volume, shake well, and centrifuge at 12000 rpm for 15 min to obtain the linear sample.

[0116] 2. Experimental Procedure

[0117] Berberine content was detected using the liquid phase method described in Example 2, and a linear equation was established between the amount of carbomer added (x) and the amount of berberine consumed (y).

[0118] 3. Test Results

[0119] Table 7 Results of the Carbomer Range Study

[0120]

[0121] The experimental results are shown in Table 7. When the carbomer content is 0.15 mg and the amount of raw materials added is 0.173 g, the proportion of carbomer in the formulation is as low as 0.09%. The linear range of carbomer content in this technical solution is 0.09%~0.21%. Within the linear range, the linear relationship between berberine consumption and carbomer addition is good, and quantitative analysis can be performed.

[0122] Example 8: Methodological Validation

[0123] 1. Specificity

[0124] Preparation of blank sample solution: Take 1g of blank cream, add 4g of urea, add 25mL of water, heat in a water bath for 10~20min, shake to dissolve, centrifuge at 12000rpm for 15min, take the lower layer solution, repeat twice, and combine to obtain the blank sample solution.

[0125] Take 25 mL of blank sample solution, add 40 mL of 0.5 mg / mL berberine solution, let stand for 20 min, make up to volume, invert and shake well, centrifuge at 12000 rpm for 15 min, take the solution and inject it. It was found that the measured amount of berberine was basically consistent with the added amount, indicating that other excipients in ivermectin cream do not participate in the reaction of berberine. This method has strong specificity.

[0126] 2. Linear relationship

[0127] Transfer 3 mL, 4 mL, 5 mL, 6 mL, and 7 mL of carbomer solution to each 100 mL volumetric flask. Add 25 mL of blank preparation solution, and then add 20 times the amount of carbomer in 0.5 mg / mL berberine solution to each flask. Let stand for 20 min, then dilute to volume, invert and shake well, centrifuge at 12000 rpm for 15 min, and inject the sample. Calculate the linear relationship between the amount of carbomer added (x) and the amount of berberine consumed (y). The experimental results show that carbomer has a linear relationship with the amount of berberine consumed in the range of 0.6–1.41 mg.

[0128] Table 8. Methodological Validation of Linear Data

[0129]

[0130] 3. Accuracy

[0131] Weigh 1g of blank cream and add 4g of urea to each. Place them in a 50mL centrifuge tube. Accurately transfer 8mL, 10mL, and 12mL of 0.2mg / mL carbomer concentrated solution to prepare solutions containing low, medium, and high concentrations of carbomer. Add 25mL of water, heat in a water bath and shake to dissolve. Centrifuge at 12000rpm for 15min. Take the lower layer solution. Repeat twice. Combine and shake well to obtain the carbomer-added preparation solution.

[0132] Take 25 mL of carbomer additive solution, add different volumes of 0.5 mg / mL berberine solution, react for 20 min, make up to volume, invert and shake well, centrifuge at 12000 rpm for 15 min, inject the solution, calculate the amount of berberine consumed, and then calculate the amount of carbomer measured according to the above linear equation. Calculate the ratio of the measured amount to the added amount to obtain the recovery rate. The results are shown in the table below.

[0133] Table 9. Method validation accuracy data

[0134]

[0135] The experimental results show that the recovery rate of carbomer at different concentrations is greater than 90%, meeting the methodological requirements. This technical solution can be used for the detection of carbomer at different concentrations in formulations.

[0136] Example 9: Testing of Multiple Batches of Samples

[0137] 1. Reagent preparation

[0138] 1.1 Preparation of Carbomer-Containing Creams

[0139] Homemade samples: Carbomer was added to a concentration of 2 mg / g according to the cream formulation and preparation method in Example 1 to obtain samples Homemade-1 and Homemade-2.

[0140] Reference sample: Commercially available ivermectin cream (manufacturer: GALDERMA; batch numbers: 3414227, 3414246) was selected.

[0141] 1.2 Preparation of the test sample

[0142] Take 1g of the above sample, add 4g of urea, add 25mL of water, heat in a water bath and shake for 10-20min to dissolve, centrifuge at 12000rpm for 15min, take the lower layer solution and put it into a 500mL volumetric flask, repeat twice, make up to volume, and shake well to obtain the final product.

[0143] 2. Experimental Procedure

[0144] According to the linear relationship between the amount of carbomer added (x) and the amount of berberine consumed (y) when the amount of urea added is 4g in Example 5, the amount of berberine consumed in the sample was detected by high performance liquid chromatography, and the amount of carbomer was calculated by linear equation.

[0145] 3. Test Results

[0146] Table 10. Detection results of carbomer content in different samples

[0147]

[0148] The test results show that this technical solution is applicable to the detection of carbomer content in different batches of ointment samples.

[0149] Example 10: Detection of other samples to be tested

[0150] To investigate the applicability of this method to other carbomer-containing products with formulations significantly different from those used in this study, the carbomer content in commercially available diclofenac diethylamine emulsion (Voltaren) was determined. The product specification was 1%, manufacturer: Haleon CH SARL, batch number: UN9D-A.

[0151] 1. Sample formulation and blank sample preparation

[0152] Patent 202380019803.9 provides a formulation for diclofenac gel containing: diclofenac diethylamine, isopropanol, propylene glycol, carbomer 974P, Silmargin 1000, cocoyl capryloyl decanoate, diethylamine, liquid paraffin, fragrance cream, and purified water, indicating that each 100mg of this product contains 1.2mg of carbomer.

[0153] Table 11. Composition of 1% Diclofenac Emulsion Formula

[0154]

[0155] Blank sample preparation method: Weigh appropriate amounts of each component (except carbomer) according to the prescription ratio, so that the total amount is about 50g, add it to a 100mL beaker, and mechanically stir at 200rpm for 30min until the components are mixed evenly.

[0156] 2. Reagent preparation

[0157] Preparation of blank sample solution for diclofenac gel: Weigh 0.8g of blank sample without carbomer, add 3.2g of urea, add 180mL of water, heat in a water bath and shake to dissolve, centrifuge at 12000rpm for 15min, take the lower layer solution and put it into a 500mL volumetric flask, repeat twice, make up to volume, and shake well to obtain the solution.

[0158] Preparation of linear samples: Weigh 3.0 mL, 4.0 mL, 5.0 mL, 6.0 mL, and 7.0 mL of 0.2 mg / mL carbomer solution respectively, and place each in a 100 mL volumetric flask. Add 25 mL of blank sample solvent, and then add 25 mL, 30 mL, 40 mL, 50 mL, and 60 mL of 0.5 mg / mL berberine solution respectively. After standing for 20 min, make up to volume, shake well, and centrifuge at 12000 rpm for 15 min to obtain the linear sample.

[0159] Preparation of the test sample: Take 200 mg of commercially available diclofenac gel into a 50 mL centrifuge tube, add 0.4 g of urea, add 25 mL of water, heat in a water bath and shake for 10-20 min to dissolve, centrifuge at 12000 rpm for 15 min, take the lower layer solution and transfer to a 50 mL volumetric flask, repeat twice, make up to volume and shake well. Take 25 mL of the test sample solution, add 40 mL of 0.5 mg / mL berberine, react for 20 min, make up to volume, invert and shake well, centrifuge at 12000 rpm for 15 min to obtain the test sample.

[0160] 3. Test Methods

[0161] Berberine content was detected by high performance liquid chromatography (HPLC) according to Example 2, and the linear relationship between carbomer addition amount x and berberine consumption amount y was calculated. The recovery rate of berberine consumption in the sample was detected by HPLC, and the amount of carbomer measured was calculated by linear equation.

[0162] 4. Test Results

[0163] Table 12. Linear Relationship between Carbomer and Berberine in Diclofenac Diethylamine Latex

[0164]

[0165] Table 13. Determination of Carbomer Content in Diclofenac Diethylamine Latex Cream

[0166]

[0167] The test results showed that the carbomer content in the test sample was 1.2 mg / 100 mg, consistent with the patent information. This technical solution can detect the carbomer content in different carbomer-containing drug formulations, and the detection results are consistent with the drug prescription. This method has a wide range of applications and high accuracy.

[0168] Example 11: Detection of Carbomer Content in Different Types

[0169] 1. Reagent preparation

[0170] Berberine solution and blank sample were prepared according to the method in Example 2.

[0171] Preparation of Carbomer 974P and Carbomer 980NF solutions: Take 50 mg of each of the two types of carbomer, add water to swell, sonicate for 30 min to prepare a 1 mg / mL stock solution, transfer the entire solution to a 250 mL volumetric flask, add 400 μL of 1 mol / L sodium hydroxide solution, dilute with water to the mark, and shake well.

[0172] Preparation of linear solutions: Weigh out carbomer concentrate according to the table below, place each in a 100mL volumetric flask, add 25mL blank sample solvent and 0.5mg / mL berberine solution to each, let stand for 20min, make up to volume, invert and shake well, centrifuge at 12000rpm for 15min to obtain the solution.

[0173] Table 14 Carbomer Addition Amount and Berberine Solution Addition Amount

[0174]

[0175] 2. Experimental Procedure

[0176] Chromatographic conditions: HPLC was performed using an Agilent Zorbax SB C18 column (4.6 mm * 150 mm, 5 μm); Mobile phase A: 1 L of purified water was mixed with 4 mL of triethylamine, and the pH was adjusted to 4.5 with glacial acetic acid. The mixture was then filtered. Mobile phase B: Acetonitrile; isocratic elution was performed between 70% mobile phase A and 30% mobile phase B. Flow rate: 1.3 mL / min; Detection wavelength: 235 nm; Column temperature: 30 °C; Injection volume: 10 μL.

[0177] The above samples were tested for berberine content to determine the residual berberine content in the samples, and then the linear relationship between the amount of carbomer added (x) and the amount of berberine consumed (y) was calculated.

[0178] 3. Test Results

[0179] The experimental results are shown in Table 15. A linear relationship exists between the amount of carbomer added (x) and the amount of berberine consumed (y), allowing for quantitative analysis. Common carbomer types contain carboxylic acid groups ranging from 52% to 68%, resulting in different drug loadings after carbomer and berberine form salts. This technical solution can ensure stable drug loadings for berberine with different types of carbomer, enabling quantitative detection of different carbomer types.

[0180] Table 15 Linearity data for carbomer content detection of different models

[0181]

[0182] Example 12: Quantitative Detection by Different Liquid Chromatography Methods

[0183] 1. Reagent preparation

[0184] Berberine solution, blank sample, carbomer solution, and linear solution were prepared according to the method in Example 11.

[0185] 2. Experimental Procedure

[0186] Chromatographic conditions 1: HPLC was performed using an InfinityLab Poroshell 120 EC-C18 column (4.6 mm × 50 mm, 2.7 μm); Mobile phase A: 1 L of purified water was mixed with 4 mL of triethylamine, and the pH was adjusted to 4.5 with glacial acetic acid. The mixture was then filtered. Mobile phase B: Acetonitrile; isocratic elution was performed between 75% mobile phase A and 25% mobile phase B. Flow rate: 0.8 mL / min; Detection wavelength: 235 nm; Column temperature: 30 °C; Injection volume: 10 μL.

[0187] Chromatographic conditions 2: HPLC was performed using a ZORBAX SB-C8 column (4.6 mm × 150 mm, 3.5 μm); Mobile phase A: 1 L of purified water was mixed with 4 mL of triethylamine, and the pH was adjusted to 4.5 with glacial acetic acid. The mixture was then filtered. Mobile phase B: methanol; isocratic elution was performed from 45% mobile phase A to 55% mobile phase B. Flow rate: 0.8 mL / min; Detection wavelength: 235 nm; Column temperature: 30 °C; Injection volume: 10 μL.

[0188] The berberine content of the above samples was detected by chromatographic conditions 1 and 2, respectively, to determine the residual berberine content in the samples, and then the linear relationship between the amount of carbomer added (x) and the amount of berberine consumed (y) was calculated.

[0189] 3. Test Results

[0190] The detection results under the two chromatographic conditions are shown in Tables 16-17. Both chromatographic conditions enabled quantitative detection of carbomer, demonstrating good linearity.

[0191] Table 16 Linearity of Detection Results under Chromatographic Condition 1

[0192]

[0193] Table 17 Linearity of Detection Results under Chromatographic Condition 2

[0194]

Claims

1. A method for detecting carbomer content in a pharmaceutical preparation, characterized in that, The detection method includes the following steps: (1) Preparation of blank test sample: Take a drug preparation that does not contain carbomer, add 2 to 8 times the amount of urea or guanidine organic base solution, heat to dissolve, centrifuge, and take the lower layer solution as the blank test sample solution; (2) Preparation of linear sample: Add carbomer solution to blank test sample solution, add berberine, let stand, centrifuge and take the supernatant to obtain linear sample solution; the ratio of berberine to carbomer is not less than 18:1; (3) Standard curve plotting: The berberine content of the linear sample solution was detected, and the berberine consumption of the linear sample was determined by comparing the amount of berberine added in step (2). A standard curve of carbomer content versus berberine consumption was established. (4) Test sample determination: The drug preparation to be tested is processed according to the method in step (1); berberine is added, and the supernatant is taken by centrifugation. The amount of berberine used is more than 18 times the amount of carbomer in the drug preparation to be tested; the berberine content is detected, and the carbomer content is calculated using the standard curve in step (3).

2. The detection method according to claim 1, characterized in that, In step (1), the amount of urea or guanidine organic base used is 4 times the amount of the drug preparation.

3. The detection method according to claim 1 or 2, characterized in that, The guanidine organic base is selected from one or more of guanidine hydrochloride, guanidine sulfate, guanidine carbonate, guanidine nitrate, and guanidine phosphate.

4. The detection method according to claim 1, characterized in that, In step (2), the ratio of berberine to carbomer is 18-42:1, and the standing time is 10-20 minutes.

5. The detection method according to claim 4, characterized in that, The ratio of berberine to carbomer is 20:

1.

6. The detection method according to claim 1, characterized in that, In step (1), the heating is performed by water bath heating for 10-20 minutes, and the centrifugation speed is 10000 rpm to 14000 rpm.

7. The detection method according to claim 1, characterized in that, The method for detecting berberine content is selected from one of the following: high performance liquid chromatography, ultra-high performance liquid chromatography, ultraviolet-visible spectrophotometry, and fluorescence spectroscopy.

8. The detection method according to claim 7, characterized in that, The chromatographic column used in the high-performance liquid chromatography method is an octadecylsilane-bonded silica gel packed column, mobile phase A is a 0.4% triethylamine solution with a pH of 4.5, and mobile phase B is selected from acetonitrile or methanol.

9. The detection method according to claim 7, characterized in that, The high performance liquid chromatography or ultra-high performance liquid chromatography method calculates the berberine content in the sample in steps (3) and (4) by using the concentration of berberine reference standard and peak area.

10. The detection method according to claim 7, characterized in that, The method for detecting berberine content adopts ultraviolet-visible spectrophotometry, and calculates the berberine content in the sample in steps (3) and (4) by using the concentration and absorbance of berberine reference standard.

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