In-vitro release analysis method of fusidic acid cream

Abstract

The invention relates to a fusidic acid cream in-vitro release analysis method, which relates to the technical field of medicine, and comprises the following steps: taking a fusidic acid cream sample to be detected, smearing the fusidic acid cream sample on a filter membrane, placing the filter membrane on a transdermal instrument, enabling a sample-free layer of the filter membrane to face downwards and face a receiving pool, carrying out closed-loop sample loading, adding a receiving medium into the receiving pool, and operating according to a method set by the transdermal instrument, and taking the receiving liquid in the receiving pool at different times for detection. According to the in-vitro release method, a vertical diffusion cell is selected, a cellulose acetate membrane is selected as a filter membrane, and a phosphate buffer solution containing an organic solvent is selected as a receiving medium, so that the release rate and degree of fusidic acid from a matrix can be accurately evaluated, the quality of fusidic acid cream can be controlled, the performance of a preparation can be evaluated, and the safety of a product can be improved. The method can quickly evaluate the drug performance, and is low in cost, short in period, green and environment-friendly.

Description

Technical Field

[0001] The invention relates to the technical field of medicines, and in particular to an in vitro release analysis method of fusidic acid cream. Background Art

[0002] Fusidic acid cream, the main component of which is fusidic acid, has a chemical name of trans-16α-carboxy-3β, 11β-dihydroxy-4β, 8β, 14α-trimethyl-18-demethyl-5β, 10α-cholest-(17Z)-17(20), 24-diene-21-acid hemihydrate, and a molecular formula of:

[0003] C 31 H 48 O6·1 / 2H2O. Fusidic acid cream is an anti-infective drug for external use on the skin. It is suitable for various bacterial skin infections, mainly for skin infections caused by Gram-positive cocci, such as impetigo, furuncle, paronychia, traumatic infection, scurvy, hidradenitis, erythrasma, folliculitis, acne vulgaris, eczema infection, ulcer infection, etc.

[0004] In Vitro Release Test (IVRT) and In Vitro Permeation Test are important contents of in vitro critical quality attribute research of generic drugs such as ointments, creams, gels, etc. Among them, IVRT is used to evaluate the rate and extent of drug release from the matrix and characterize the steady-state in vitro release rate of drugs in the preparation under specific conditions (i.e. given method parameters). It is an important indicator in quality research and stability investigation, and can be used as an effective method to control drug quality and evaluate preparation performance.

[0005] CN115267004A discloses a method for detecting the content of related substances in fusidic acid, using LC-MS method to detect the content of impurity M in fusidic acid, and detecting the content of impurity M in the test solution by external standard one-point method, but does not involve the in vitro release-related data detection method of fusidic acid cream. CN117705983A discloses an evaluation method for in vitro transdermal release of a topical composition containing azelaic acid, using Strat-M artificial membrane as a permeation barrier, using phosphate buffer as a receiving medium, and combining ultra-high performance liquid chromatography analysis method, which can quickly and efficiently determine the transdermal rate and cumulative permeation amount of azelaic acid. The in vitro transdermal test is mainly used to evaluate the rate and degree of the drug reaching the skin action site or near it, or to evaluate the behavior of the drug permeating through the skin and entering the body, simulating the transdermal process of the drug under physiological conditions, which is different from the in vitro release test for evaluating the drug release rate of the preparation. There is no relevant report on the in vitro release test method of fusidic acid cream.

[0006] Therefore, it is necessary to develop a simple and accurate in vitro release method for fusidic acid cream. Summary of the invention

[0007] The purpose of the present invention is to provide an in vitro release analysis method of fusidic acid cream. The method is simple and has high accuracy, and solves the problem that there is no in vitro release analysis method of fusidic acid cream.

[0008] The technical solution provided by the present invention is as follows:

[0009] The present invention provides an in vitro release analysis method of fusidic acid cream, comprising the following steps:

[0010] Take the fusidic acid cream sample to be tested and smear it on the filter membrane and place it on the transdermal instrument. The filter membrane without the sample layer faces downward to the receiving pool. Close the loop and add the receiving medium to the receiving pool. Run the transdermal instrument according to the method set. Take the receiving solution in the receiving pool for detection at different times.

[0011] As an embodiment of the present invention, the transdermal instrument is a vertical diffusion cell.

[0012] As an embodiment of the present invention, the filter membrane is a polyethersulfone membrane, a cellulose acetate membrane, a polyvinylidene fluoride membrane or a polytetrafluoroethylene membrane, preferably a cellulose acetate membrane.

[0013] As an embodiment of the present invention, the receiving medium is a phosphate buffer containing an organic solvent.

[0014] As an embodiment of the present invention, the receiving medium is a phosphate buffer containing an organic solvent, wherein the organic solvent is selected from methanol or ethanol.

[0015] As an embodiment of the present invention, the receiving medium is a phosphate buffer containing methanol, the volume ratio of the phosphate buffer to methanol is 7:3, and the pH is 7.4.

[0016] As an embodiment of the present invention, the temperature of the transdermal instrument is 26-37°C, preferably 32°C.

[0017] As an embodiment of the present invention, the rotation speed of the transdermal instrument is 200-750 rpm, preferably 600 rpm.

[0018] As an embodiment of the present invention, the detection is carried out by high performance liquid chromatography, and the chromatographic column of the high performance liquid chromatography is Waters Symmetry C18, 4.6 mm×15 cm, 3.5 μm.

[0019] As an embodiment of the present invention, the mobile phase of the high performance liquid chromatography is methanol-0.05M phosphoric acid-acetonitrile in a volume ratio of 10:40:50.

[0020] Compared with the prior art solutions, the present invention has the following beneficial effects:

[0021] The in vitro release method of the present invention selects a vertical diffusion cell, the filter membrane selects a cellulose acetate membrane, and the receiving medium selects a phosphate buffer containing an organic solvent, which can accurately evaluate the rate and degree of release of fusidic acid from the matrix, is beneficial to control the quality of fusidic acid cream and evaluate the performance of the preparation, and improve the safety of the product. The present invention can quickly evaluate the performance of the drug, has low cost, short cycle, and is green and environmentally friendly. The method is simple, and realizes the high-precision evaluation of the quality difference of fusidic acid cream by the in vitro release evaluation method, which is of great significance. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 is the pH solubility curve calculated by the standard curve method;

[0023] Figure 2 It is the release rate curve of cellulose acetate membrane in Table 3 "Filter membrane screening results";

[0024] Figure 3 It is the release rate curve of the polyethersulfone membrane in Table 3 "Filter membrane screening results";

[0025] Figure 4 It is the release rate curve of the polytetrafluoroethylene membrane in Table 3 "Filter membrane screening results";

[0026] Figure 5 It is the release rate curve of the polyvinylidene fluoride membrane in Table 3 "Filter membrane screening results";

[0027] Figure 6 It is the release rate curve of nitrocellulose membrane in Table 3 "Filter membrane screening results";

[0028] Figure 7 It is the release rate curve of the glass cellulose membrane in Table 3 "Filter membrane screening results";

[0029] Figure 8 is the cumulative release profile of the six membranes;

[0030] Fig. 9 It is the release rate curve of pH 7.4 phosphate buffer-methanol (7:3) in Table 4 "Study results of pH 7.4 phosphate buffer and mixed solutions of organic phases in different proportions";

[0031] Fig.10It is the release rate curve of pH 7.4 phosphate buffer-methanol (6:4) in Table 4 "Study results of pH 7.4 phosphate buffer and mixed solutions of organic phases in different proportions";

[0032] Fig.11 It is the release rate curve of pH 7.4 phosphate buffer-methanol (5:5) in Table 4 "Study results of pH 7.4 phosphate buffer and mixed solutions of organic phases in different ratios";

[0033] Fig.12 It is the release rate curve of pH 7.4 phosphate buffer-ethanol (7:3) in Table 4 "Study results of pH 7.4 phosphate buffer and mixed solutions of organic phases in different proportions";

[0034] Fig.13 It is the release rate curve of pH 7.4 phosphate buffer-isopropanol (7:3) in Table 4 "Study results of pH 7.4 phosphate buffer and mixed solutions of organic phases in different ratios";

[0035] Fig.14 is the release rate curve of the reference preparation and the self-developed preparation of Example 1;

[0036] Fig.15 is the release rate curve of Example 2;

[0037] Fig.16 is the release rate curve of Example 3;

[0038] Fig.17 is the release rate curve of Comparative Example 1;

[0039] Fig.18 It is the release rate curve of Comparative Example 2. DETAILED DESCRIPTION

[0040] The present invention is further described below in conjunction with examples, but the embodiments of the present invention are not limited thereto.

[0041] The raw materials, reagents, materials and equipment used in the examples can all be purchased through commercial channels; among them, the raw material of fusidic acid comes from Jiangsu Jiuyang Biopharmaceutical Co., Ltd., the fusidic acid cream (reference preparation) comes from Ireland Leo Pharmaceuticals Co., Ltd., and the fusidic acid cream (self-developed preparation) comes from Nanjing Hanxin Pharmaceutical Technology Co., Ltd. The reference substance is diethanolamine fusidic acid, which comes from the British Pharmacopoeia Commission Laboratory.

[0042] Filter membrane treatment: Take the filter membrane and soak it in the receiving medium for at least 30 minutes, take it out and dry it for later use.

[0043] Place the treated filter membrane on weighing paper, place a 2mm gasket in the middle of the filter membrane, weigh 300 mg of fusidic acid cream into the gasket, scrape it flat and install it on the transdermal instrument, with the filter membrane without drug layer facing downward to the receiving pool, close the loop and load the sample, and fill the receiving pool with degassed receiving medium. Run the transdermal instrument according to the set method, take samples at different time points, and use HPLC to determine the drug concentration of fusidic acid in the receiving medium, calculate the cumulative drug release per unit area, and plot the release rate curve with the square root of time as the X-axis and the cumulative release per unit area as the Y-axis. The slope of the linear equation is the release rate.

[0044] Diffusion cell parameters (IVRT parameters): The transdermal instrument in this application is a vertical diffusion cell, using Logan SYSTEM918-12,

[0045] Stirring speed: 600 rpm / min;

[0046] Diffusion cell temperature: 32°C

[0047] Diffusion cell volume: 12mL

[0048] Sampling time points: 0.5h, 1h, 2h, 3h, 4h, 5h, 6h.

[0049] Chromatographic conditions:

[0050] instrument High performance liquid chromatography Detector UV detector Detection wavelength 235nm Chromatographic columns Waters Symmetry C18, 4.6mm×15cm, 3.5μm Column temperature 25℃ Flow rate 1.2mL / min Injection volume 10μL Run time 5min Mobile phase Methanol-0.05M phosphoric acid-acetonitrile (10:40:50)

[0051] Receiving media screening

[0052] 1. Study on the solubility of fusidic acid API

[0053] Method reference: Chinese Pharmacopoeia 2020 Edition Part IV, General Rules 15 (2). Take 5 mg of fusidic acid API for the test, and the diluent is the mobile phase. The preparation method of phosphate buffer solution refers to the "Guidelines for Determination and Comparison of Dissolution Curves of Ordinary Oral Solid Preparations" published by the Drug Review Center of the State Food and Drug Administration.

[0054] Receptor medium Dissolution Normal saline Not completely dissolved in 100mL of diluent pH 6.0 phosphate buffer Not completely dissolved in 100mL of diluent pH 6.8 phosphate buffer Completely dissolved in 50mL of diluent pH 7.4 phosphate buffer Completely dissolved in 10mL of diluent 10% methanol in water Not completely dissolved in 100mL of diluent 20% methanol in water Not completely dissolved in 100mL of diluent 30% methanol in water Not completely dissolved in 100 mL of diluent 40% methanol in water Not completely dissolved in 100mL of diluent 50% methanol in water Completely dissolved in 5 mL of diluent

[0055] From the above results, it can be seen that the solubility of fusidic acid raw material is relatively high in alkaline solution and 50% methanol solution. Since high-concentration methanol solution is not suitable for use as a receiving medium for IVPT, phosphate buffer is tentatively used for further pH solubility curve research.

[0056] 2. pH solubility curve study

[0057] Solution preparation:

[0058] Diluent: Mobile phase.

[0059] Preparation of standard curve: Take an appropriate amount of reference substance and dilute it with diluent to prepare linear solutions of different concentrations. The concentrations of fusidic acid are 0.5 mg / mL, 0.25 mg / mL, 0.1 mg / mL, 0.05 mg / mL, 0.025 mg / mL, 0.01 mg / mL and 0.005 mg / mL respectively.

[0060] Preparation of solubility curve solution of fusidic acid API: Accurately weigh 6 portions of 10 mg fusidic acid API and place them in 20 mL headspace bottles, add 10 mL of pH 5.0 phosphate buffer, pH 5.8 phosphate buffer, pH 6.5 phosphate buffer, pH 6.8 phosphate buffer, and pH 7.4 phosphate buffer, respectively, press the cap, shake at 32 ° C for 24 h, take samples, filter, and detect the content by HPLC.

[0061] Chromatographic conditions were as described above.

[0062] Calculation method:

[0063] Standard curve method: use the concentration of the linear solution as the horizontal coordinate and the peak area as the vertical coordinate to fit the standard curve. The linear correlation coefficient R 2 ≥0.998, substitute the peak area of ​​the test solution in the solubility study into the standard curve to calculate its concentration.

[0064] Table 1: pH solubility curve study results

[0065]

[0066] As shown in Table 1, the solubility of fusidic acid increases with the increase of pH. The solubility of fusidic acid raw material in pH 7.4 phosphate buffer is 0.53 mg / mL. The cumulative release of fusidic acid cream is generally less than 30%. When the cumulative release is 30%, the cumulative concentration of fusidic acid is 30%×300 mg×2%÷12 mL=0.15 mg / mL. Therefore, pH 7.4 phosphate buffer as a receiving medium can meet the sink condition of IVRT.

[0067] Membrane confirmation

[0068] Experimental process:

[0069] Fusidic acid solution: Weigh an appropriate amount of fusidic acid API and prepare 2.5 μg / mL fusidic acid solution with pH 7.4 phosphate buffer.

[0070] Control solution: Take 12 mL of fusidic acid solution and place it at 32°C, shake it for 6 hours and 24 hours respectively. Prepare three times in parallel.

[0071] Membrane solution: Take 12 mL of fusidic acid solution and add glass cellulose membrane, polyethersulfone membrane, nitrocellulose membrane, cellulose acetate membrane, nylon membrane, polyvinylidene fluoride membrane, and polytetrafluoroethylene membrane to cover, place at 32°C, and shake for 6 hours and 24 hours respectively. Prepare 3 copies in parallel.

[0072] Chromatographic conditions were as described above.

[0073] Calculation method:

[0074] According to the peak area external standard method, the peak area of ​​fusidic acid in the control solution was used as the reference to calculate the recovery rate of fusidic acid in each membrane-added solution. The results are shown in Table 2 Membrane confirmation results.

[0075] Table 2 Membrane confirmation results

[0076] Filter membrane Recovery rate (6h) Recovery rate (24h) Glass cellulose membrane 96.2％ 95.8％ Polyethersulfone membrane 100.6％ 100.8％ Nitrocellulose membrane 95.6％ 96.9％ Cellulose acetate membrane 104.7％ 103.5％ Nylon membrane 89.6％ 86.8％ Polyvinylidene fluoride film 98.4％ 97.0％ Polytetrafluoroethylene membrane 99.4％ 100.8％

[0077] As shown in Table 2, except for the nylon membrane, the recovery rates of the other membrane solutions were between 95% and 105%, and all of them could be used as membranes for IVRT of fusidic acid cream for further screening.

[0078] Membrane screening

[0079] Experimental process:

[0080] The filter membranes include glass cellulose membrane, polyethersulfone membrane, cellulose acetate membrane, polyvinylidene fluoride membrane, polytetrafluoroethylene membrane and nitrocellulose membrane.

[0081] Receiving medium selection: pH 7.4 phosphate buffer.

[0082] Filter membrane treatment: Soak the filter membrane in the receiving medium for 30 minutes, dry it and set aside.

[0083] Place the treated filter membrane on weighing paper, take a 2 mm gasket and place it in the middle of the filter membrane, weigh about 300 mg of fusidic acid cream (reference preparation) into the gasket, and scrape it flat; run the transdermal instrument according to the set method and take samples at different time points.

[0084] IVRT parameters and chromatographic conditions were as described above.

[0085] The results of membrane screening are shown in Table 3

[0086] Table 3 Filter membrane screening results

[0087]

[0088]

[0089] From Table 3 and Figure 8The results showed that among the above six membranes, the linear relationship of the release rate of fusidic acid cream within 6 hours was good, and the cumulative release amount on cellulose acetate membrane (CA) was the highest. Therefore, cellulose acetate membrane (CA) was selected for the development of IVRT method of fusidic acid cream.

[0090] Further screening of receiving medium - pH 7.4 phosphate buffer mixed with different proportions of organic phase

[0091] Filter membrane: Cellulose acetate membrane (CA)

[0092] Place the treated filter membrane on weighing paper, take a 2 mm gasket and place it in the middle of the filter membrane, weigh about 300 mg of fusidic acid cream (reference preparation) into the gasket, and scrape it flat; run the transdermal instrument according to the set method and take samples at different time points.

[0093] IVRT parameters and chromatographic conditions were as described previously.

[0094] The results are shown in Table 4, which are the research results of mixed solutions of phosphate buffer and organic phase in different proportions.

[0095] Table 4 Study results of pH 7.4 phosphate buffer and mixed solutions of organic phase in different proportions

[0096]

[0097] As shown in Table 4, adding an organic phase to the receiving medium can significantly increase the release rate. However, in the receiving medium containing 50% methanol and 30% isopropanol, the cumulative release of fusidic acid exceeds 30% (the cumulative release generally does not exceed 30%), and the cumulative release of 30% methanol is slightly greater than that of 30% ethanol. Therefore, the mixed solution containing 30% methanol is selected as the receiving medium, and the R of the linear equation of the release rate is 2 It is 0.9964, which meets the acceptance standard (NLT 0.97).

[0098] Example 1

[0099] Membrane treatment: Take the cellulose acetate membrane and soak it in the receiving medium for 30 minutes, take it out and dry it for later use; the receiving medium is pH 7.4 phosphate buffer-methanol (7:3).

[0100] Place the treated filter membrane on weighing paper, take a 2mm gasket and place it in the middle of the filter membrane, weigh 300mg of fusidic acid cream (reference preparation) and fusidic acid cream (self-made preparation) into the gasket, scrape it flat and install it in the transdermal instrument. The filter membrane without drug layer faces downward to the receiving pool, and the receiving pool is filled with degassed receiving medium to measure the in vitro release rate of the reference preparation and the self-made preparation. Operate according to the set transdermal instrument method, take samples at different time points, use HPLC to determine the drug concentration of fusidic acid in the receiving medium, calculate the cumulative drug release per unit area, and plot the release rate curve with the square root of time as the X-axis and the cumulative release per unit area as the Y-axis. The slope of the linear equation is the release rate. The release rate curve is shown in the figure. Fig.14 .

[0101] IVRT parameters and chromatographic conditions were as described previously.

[0102] Acceptable criteria for equivalence: The reference batch is called the reference batch (R), and the self-developed batch is called the self-developed batch (T). The amount of drug released from each diffusion cell in R is plotted against the square root of time. The resulting slope is the reference slope. Repeat the process for the self-developed batch (T). Combine each self-developed (T)-reference (R) pair and calculate all T / R slope ratios. This process is summarized in a table, where TS represents the slope of the self-developed formulation and RS is the slope of the reference formulation. Table 5 shows the ratio of the slope of the self-developed batch (TS) and the slope of the reference batch (RS).

[0103] Table 5 Ratio of slope of self-developed batch (TS) and slope of reference batch (RS)

[0104] RS1 RS2 RS3 RS4 RS5 RS6 TS1 TS1 / RS1 TS1 / RS2 TS1 / RS3 TS1 / RS4 TS1 / RS5 TS1 / RS6 TS2 TS2 / RS1 TS2 / RS2 TS2 / RS3 TS2 / RS4 TS2 / RS5 TS2 / RS6 TS3 TS3 / RS1 TS3 / RS2 TS3 / RS3 TS3 / RS4 TS3 / RS5 TS3 / RS6 TS4 TS4 / RS1 TS4 / RS2 TS4 / RS3 TS4 / RS4 TS4 / RS5 TS4 / RS6 TS5 TS5 / RS1 TS5 / RS2 TS5 / RS3 TS5 / RS4 TS5 / RS5 TS5 / RS6 TS6 TS6 / RS1 TS6 / RS2 TS6 / RS3 TS6 / RS4 TS6 / RS5 TS6 / RS6

[0105] After calculating the T / R slope ratio, the 36 T / R slope ratios were sorted from low to high. Then, the 8th and 29th slope ratios were converted to percentage units (i.e., multiplied by 100), representing the lower and upper limits of the 90% confidence interval, respectively. If both were within the range of 90% to 111%, the first stage test was passed.

[0106] If the first phase of the test fails, 4 more tests (2 reference and 2 self-developed, 6 diffusion cells each) should be added, and 12 slopes should be added for each test batch. For all 18 T / R slopes of each test batch, a total of 324 T / R slope ratios are calculated in the above manner, and then sorted from low to high. If the 110th and 215th slope ratios (converted to percentage units, representing the lower and upper limits of the 90% confidence interval, respectively) are both within the range of 90% to 111%, the second phase of the test is passed.

[0107] The slope ratio of the self-made preparation (TS) and the reference preparation (RS) is shown in Table 6 below:

[0108] Table 6 Comparison results of slope (TS) of self-made preparation and slope (RS) of reference preparation

[0109] TS / RS RS1 RS2 RS3 RS4 RS5 RS6 TS1 102％ 105％ 109％ 98％ 101％ 104％ TS2 109％ 112％ 117％ 105％ 108％ 111％ TS3 107％ 110％ 115％ 103％ 106％ 109％ TS4 111％ 115％ 120％ 107％ 110％ 113％ TS5 93％ 96％ 100％ 89％ 92％ 94％ TS6 96％ 98％ 102％ 92％ 95％ 97％

[0110] Conclusion: The release rate of the self-developed preparation was compared with that of the reference preparation, and the 90% confidence interval was 96%-111%. Within the range of 90%-111%, they were statistically equivalent. Therefore, the in vitro release of the self-developed preparation and the reference preparation was equivalent.

[0111] Example 2

[0112] Membrane treatment: Take the polyethersulfone membrane and soak it in the receiving medium for 45 minutes, take it out and dry it for later use; the receiving medium is selected as pH 7.4 phosphate buffer-ethanol (7:3).

[0113] Place the treated filter membrane on weighing paper, take a 2mm gasket and place it in the middle of the filter membrane, weigh 300mg of fusidic acid cream (reference preparation) into the gasket, scrape it flat and install it in the transdermal instrument, with the filter membrane without the drug layer facing downwards to the receiving pool, which is filled with degassed receiving medium. Run the transdermal instrument according to the set method, take samples at different time points, use HPLC to determine the release amount of fusidic acid in the receiving medium and draw a release curve. The release curve is as follows: Fig.15 , R of the linear equation for release rate 2 It is 0.9974, which meets the acceptance standard (NLT 0.97).

[0114] Among the IVRT parameters, the diffusion cell temperature was 26° C. and the stirring speed was 750 rpm / min. Other IVRT parameters and chromatographic conditions were consistent with those in Example 1.

[0115] Example 3

[0116] Membrane treatment: Take the polyvinylidene fluoride membrane and soak it in the receiving medium for 60 minutes, take it out and dry it for later use; the receiving medium is selected as pH 7.4 phosphate buffer-methanol (7:3).

[0117] Place the treated filter membrane on weighing paper, take a 2mm gasket and place it in the middle of the filter membrane, weigh 300mg of fusidic acid cream (reference preparation) into the gasket, scrape it flat and install it in the transdermal instrument, with the filter membrane without the drug layer facing downwards to the receiving pool, which is filled with degassed receiving medium. Run the transdermal instrument according to the set method, take samples at different time points, use HPLC to determine the release amount of fusidic acid in the receiving medium and draw a release curve. The release curve is as follows: Fig.16 , R of the linear equation for release rate2 It is 0.9988, which meets the acceptance standard (NLT 0.97).

[0118] Among the IVRT parameters, the diffusion cell temperature was 37° C. and the stirring speed was 200 rpm / min. Other IVRT parameters and chromatographic conditions were consistent with those in Example 1.

[0119] Example 4

[0120] Membrane treatment: soak the polytetrafluoroethylene membrane in the receiving medium for 50 minutes, take it out and dry it for later use; the receiving medium is pH 7.4 phosphate buffer-methanol (7:3).

[0121] Place the treated filter membrane on weighing paper, take a 2mm gasket and place it in the middle of the filter membrane, weigh 300mg of fusidic acid cream (reference preparation) into the gasket, scrape it flat and install it in the transdermal instrument. The filter membrane without the drug layer faces downward to the receiving pool, and the receiving pool is filled with degassed receiving medium. Run the transdermal instrument according to the set method, take samples at different time points, use HPLC to determine the release amount of fusidic acid in the receiving medium and draw a release curve. R of the linear equation of release rate 2 It is 0.998, which meets the acceptance standard (NLT 0.97).

[0122] The IVRT parameters and chromatographic conditions were consistent with those in Example 1.

[0123] Comparative Example 1

[0124] Membrane treatment: Soak the nitrocellulose membrane in the receiving medium for at least 30 minutes, take it out and dry it for later use; the receiving medium is pH 7.4 phosphate buffer-methanol (7:3).

[0125] Place the treated filter membrane on weighing paper, take a 2mm gasket and place it in the middle of the filter membrane, weigh 300mg of fusidic acid cream into the gasket, scrape it flat and install it in the transdermal instrument, with the filter membrane without the drug layer facing downwards to the receiving pool, which is filled with degassed receiving medium. Run the transdermal instrument according to the set method, take samples at different time points, use HPLC to determine the release amount of fusidic acid in the receiving medium and draw a release curve. The release curve is as follows: Fig.17 , R of the linear equation for release rate 2 It is 0.9598, which does not meet the acceptance standard (NLT 0.97).

[0126] The IVRT parameters and chromatographic conditions were consistent with those in Example 1.

[0127] Comparative Example 2

[0128] Membrane treatment: Soak the cellulose acetate membrane in the receiving medium for at least 30 minutes, take it out and dry it for later use; the receiving medium is pH 7.4 phosphate buffer.

[0129] Place the treated filter membrane on weighing paper, take a 2mm gasket and place it in the middle of the filter membrane, weigh about 300mg of fusidic acid cream into the gasket, scrape it flat and install it in the transdermal instrument, with the filter membrane without the drug layer facing downwards to the receiving pool, which is filled with degassed receiving medium. Run the transdermal instrument according to the set method, take samples at different time points, use HPLC to determine the release amount of fusidic acid in the receiving medium and draw a release curve. The release curve is as follows: Fig.18 , the cumulative release is 3.99%, which is a low cumulative release.

[0130] The IVRT parameters and chromatographic conditions were consistent with those in Example 1.

Claims

1. An in vitro release analysis method for fusidic acid cream, characterized in that: The steps include: Take the fusidic acid cream sample to be tested and smear it on the filter membrane and place it on the transdermal instrument. The filter membrane without the sample layer faces downward to the receiving pool. Close the loop and add the receiving medium to the receiving pool. Run the transdermal instrument according to the method set. Take the receiving solution in the receiving pool for detection at different times.

2. The in vitro release analysis method of a fusidic acid cream according to claim 1, wherein: The transdermal instrument is a vertical diffusion cell.

3. The in vitro release analysis method of a fusidic acid cream according to claim 1, characterized in that: The filter membrane is a polyethersulfone membrane, a cellulose acetate membrane, a polyvinylidene fluoride membrane or a polytetrafluoroethylene membrane, preferably a cellulose acetate membrane.

4. The in vitro release analysis method of a fusidic acid cream according to claim 1, characterized in that: The receiving medium is a phosphate buffer containing an organic solvent.

5. The in vitro release analysis method of a fusidic acid cream according to claim 4, characterized in that: The receiving medium is a phosphate buffer containing an organic solvent, wherein the organic solvent is selected from methanol or ethanol.

6. The in vitro release analysis method of a fusidic acid cream according to claim 5, characterized in that: The receiving medium is a phosphate buffer containing methanol, the volume ratio of the phosphate buffer to methanol is 7:3, and the pH is 7.

4.

7. The in vitro release analysis method of fusidic acid cream according to any one of claims 1 to 6, characterized in that: The temperature of the transdermal instrument is 26-37°C, preferably 32°C.

8. The in vitro release analysis method of a fusidic acid cream according to claim 7, characterized in that: The rotation speed of the transdermal instrument is 200-750 rpm, preferably 600 rpm.

9. The in vitro release analysis method of fusidic acid cream according to any one of claims 1 to 8, characterized in that: The detection is carried out by high performance liquid chromatography, and the chromatographic column of the high performance liquid chromatography is Waters Symmetry C18, 4.6 mm×15 cm, 3.5 μm.

10. The in vitro release analysis method of a fusidic acid cream according to claim 9, characterized in that: The mobile phase of the high performance liquid chromatography method is methanol-0.05M phosphoric acid-acetonitrile in a volume ratio of 10:40:50.

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