Separation method, detection method, composition of rotigotine or a pharmaceutical salt thereof, pharmaceutical preparation, and use
By optimizing the mobile phase and column conditions using high-performance liquid chromatography, the problem of separating and detecting impurity L in rotigotine was solved, achieving quality control of high-purity rotigotine and improving product safety and detection accuracy.
Patent Information
- Application Number
- CN202310280149.9
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2023-03-22
- Publication Date
- 2025-11-07
- Estimated Expiration
- 2043-03-22
AI Technical Summary
Existing technologies cannot effectively separate and detect impurity L in rotigotine, leading to difficulties in quality control. The European Pharmacopoeia detection method suffers from severe peak tailing.
High-performance liquid chromatography (HPLC) was used, employing an aqueous and organic mobile phase with specific pH values, combined with a chromatographic column packed with octadecyl or octyl-bonded silica gel, to separate and detect rotigotine or its pharmaceutical salts. The gradient elution conditions of the mobile phase were optimized to ensure the effective separation and accurate quantification of substance A.
It achieves efficient separation and detection of substance A in rotigotine or its medicinal salt, improves purity and quality control, ensures the safety and quality of rotigotine products, can effectively separate multiple impurities, has good peak symmetry, high sensitivity, and wide applicability.
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Figure CN116359381B_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The present application relates to the technical field of detection, in particular to a separation method, a detection method, a composition of rotigotine or a pharmaceutical salt thereof, a pharmaceutical preparation and a use of rotigotine or a composition thereof. BACKGROUND
[0002] The English generic name of rotigotine is Rotigotine, the chemical name is (6S)-6-[propyl-[2-(2-thienyl)ethyl]-amino]-5,6,7,8-tetrahydro-1-naphthalenol, the molecular formula is C 19 H 25 NOS, which belongs to non-ergot selective dopamine receptor agonists (D1 / D2 / D3), is used for the treatment of Parkinson's disease and restless leg syndrome, and works by stimulating dopamine receptors in the body and mimicking the neurotransmitter dopamine. The structural formula of rotigotine is as follows:
[0003]
[0004] Impurity research is an extremely important research content in the development process of drugs. According to the existing synthesis process of rotigotine, as shown in the following figure. The applicant infers that there may be impurities with the structure shown as formula L.
[0005]
[0006] Although the European Pharmacopoeia (European Pharmacopoeia 10.0) discloses that impurities B-K may exist in rotigotine (European Pharmacopoeia (European Pharmacopoeia 10.0) 3748-3750 pages).
[0007] However, according to the European Pharmacopoeia, the main peak of rotigotine is seriously tailing, and the content of the impurity shown as formula L in rotigotine cannot be accurately determined, which brings quality risks to rotigotine.
[0008] In patent CN 110726786A, a HPLC separation and analysis method of rotigotine and its important intermediates is provided. In this method, only the separation of impurities RG03 (impurity H described above), RG02 (not the impurity recorded in the European Pharmacopoeia) and rotigotine is described, and other impurities in the European Pharmacopoeia and the impurity shown as formula L are not considered, which cannot effectively control the product quality. SUMMARY
[0009] In order to solve the above problems in the prior art, the application provides a separation method, a detection method, a composition containing rotigotine or a pharmaceutically acceptable salt thereof, a pharmaceutical preparation and an application of substance A in rotigotine or a composition thereof, rotigotine or a pharmaceutically acceptable salt thereof, the separation and detection method has the advantages of good separation degree, symmetrical peak shape, high sensitivity and accurate quantification of substance A, and defects in rotigotine quality control are made up, and the separation and detection method can be used for quality inspection or quality control of rotigotine or a composition thereof, rotigotine or a pharmaceutically acceptable salt thereof.
[0010] The first aspect of the application provides a separation method of substance A in rotigotine or a composition thereof, rotigotine or a pharmaceutically acceptable salt thereof, comprising:
[0011] The rotigotine or a composition thereof, rotigotine or a pharmaceutically acceptable salt thereof is separated by high performance liquid chromatography, a mobile phase in the high performance liquid chromatography comprises an aqueous phase and an organic phase, and the aqueous phase has a pH value of 6.0-8.0.
[0012] The substance A comprises a compound represented by formula L or a salt thereof.
[0013]
[0014] In some embodiments of the application, the separation method is a separation method of substance A in rotigotine or a composition thereof.
[0015] In some embodiments of the application, the separation method is a separation method of substance A in rotigotine raw material.
[0016] In the application, the compound represented by formula L or a salt thereof is impurity L, and in some embodiments of the application, the compound represented by formula L is impurity L.
[0017] In some embodiments of the application, the pharmaceutically acceptable salt of rotigotine is a salt of rotigotine and an inorganic acid or an organic acid, the inorganic acid is selected from one or more of hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid and nitric acid, and the organic acid is selected from one or more of citric acid, fumaric acid, maleic acid, acetic acid, benzoic acid, lactic acid, methanesulfonic acid, naphthalenesulfonic acid and p-toluene sulfonic acid.
[0018] In some specific embodiments of the application, the pharmaceutically acceptable salt of rotigotine is hydrochloride of rotigotine or hydrobromide of rotigotine.
[0019] In some embodiments of the application, the organic phase is acetonitrile or methanol, and preferably acetonitrile.
[0020] In some embodiments of the application, the aqueous phase is a buffer, and preferably an inorganic salt buffer; the inorganic salt is preferably one or more of disodium hydrogen phosphate, dipotassium hydrogen phosphate and diammonium hydrogen phosphate.
[0021] In the present application, the inorganic salt buffer is a mixed solution of inorganic salt and inorganic acid corresponding to the inorganic salt. The inorganic salt buffer is obtained by adjusting the pH value of the inorganic salt solution with inorganic acid corresponding to the inorganic salt.
[0022] In some embodiments of the present application, the pH value of the aqueous phase is 7.4-7.8.
[0023] In some specific embodiments of the present application, the pH value of the aqueous phase is 6.0, 7.0, 7.4, 7.6, 7.8 or 8.0.
[0024] In some embodiments of the present application, the mobile phase is composed of an organic phase and an aqueous phase.
[0025] In some embodiments of the present application, the high performance liquid chromatography separation further comprises separation using a chromatographic column with octadecyl-bonded silica or octyl-bonded silica as the filler.
[0026] In some specific embodiments of the present application, the chromatographic column with octadecyl-bonded silica as the filler is shim-pack VP-ODS, and the specification of the chromatographic column is preferably 4.6 mm x 250 mm, 5 μm.
[0027] In some specific embodiments of the present application, the chromatographic column with octyl-bonded silica as the filler is Agilent XDB C8, and the specification of the chromatographic column is preferably 4.6 mm x 150 mm, 5 μm.
[0028] In some embodiments of the present application, the high performance liquid chromatography separation further comprises the following chromatographic conditions:
[0029] flow rate: 1.0-1.5 ml / min; and / or the separation method further comprises high performance liquid chromatography separation using the following chromatographic conditions:
[0030] column temperature: 25-35℃; and / or
[0031] injection volume: 5-20 μL; and / or
[0032] mobile phase: A: buffer, B: acetonitrile, in volume percentage, gradient elution conditions as follows:
[0033] Time (min) Mobile phase A (%) Mobile phase B (%) 0 90 10 2 90 10 12 50 50 30 15 85 35 15 85 35.1 90 10 45 90 10 .
[0034] In some specific embodiments of the present application, the high performance liquid chromatography separation chromatogram is:
[0035] Chromatographic column: chromatographic column with octadecyl-bonded silica or octyl-bonded silica as filler;
[0036] Column temperature: 25-35℃;
[0037] Injection volume: 5-20μL;
[0038] Mobile phase: A: phosphate buffer, B: acetonitrile, gradient elution with volume percentage as shown in the following table;
[0039] Time (min) Mobile phase A (%) Mobile phase B (%) 0 90 10 2 90 10 12 50 50 30 15 85 35 15 85 35.1 90 10 45 90 10
[0040] Flow rate: 1.0-1.5ml / min.
[0041] In some embodiments of the present application, the injection volume is 5μL, 10μL, 20μL, preferably 10μL.
[0042] In some embodiments of the present application, the substance A is a compound of formula L or a salt thereof.
[0043] In some embodiments of the present application, the substance A is a compound of formula L.
[0044] In some embodiments of the present application, the substance A is a salt of a compound of formula L.
[0045] In some embodiments of the present application, the salt of a compound of formula L is a hydrobromide salt of a compound of formula L.
[0046] In some embodiments of the present application, the substance A further comprises one or more of impurity B or a salt thereof, impurity C or a salt thereof, impurity D or a salt thereof, impurity E, impurity F or a salt thereof, impurity J or a salt thereof, impurity H or a salt thereof, impurity I or a salt thereof, impurity G or a salt thereof, and impurity K, and the structural formulae of the impurity B, the impurity C, the impurity D, the impurity E, the impurity F, the impurity J, the impurity H, the impurity I, the impurity G, and the impurity K are as follows:
[0047]
[0048] In some embodiments of the present application, the substance A further comprises one or more of impurity B, impurity C, impurity D, impurity E, impurity F, impurity J, impurity H, impurity I, impurity G, and impurity K.
[0049] In the present application, impurity 1 is a compound of formula L or a salt thereof; and impurity 2 is one or more of impurity B or a salt thereof, impurity C or a salt thereof, impurity D or a salt thereof, impurity E, impurity F or a salt thereof, impurity J or a salt thereof, impurity H or a salt thereof, impurity I or a salt thereof, impurity G or a salt thereof, and impurity K.
[0050] In some embodiments of the present application, the impurity 1 is the compound of Formula L.
[0051] In some embodiments of the present application, the substance A comprises: impurity 1 and impurity 2.
[0052] In some embodiments of the present application, the substance A is impurity 1 and impurity 2.
[0053] In some embodiments of the present application, the impurity 2 is one or more selected from the group consisting of impurity B, impurity C, impurity D, impurity E, impurity F, impurity J, impurity H, impurity I, impurity G and impurity K.
[0054] In some embodiments of the present application, the impurity 2 is the impurity B, the impurity C, the impurity D, the impurity E, the impurity F, the impurity G, the impurity H, the impurity I, the impurity J and the impurity K.
[0055] In some embodiments of the present application, the substance A is the compound of Formula L, the impurity B, the impurity C, the impurity D, the impurity E, the impurity F, the impurity G, the impurity H, the impurity I, the impurity J and the impurity K.
[0056] In some embodiments of the present application, the substance A is the compound of Formula L, the impurity B, the impurity C, the impurity D, the impurity F, the impurity G, the impurity H, the impurity I and the impurity K.
[0057] The second application of the present application provides a detection method of substance A in rotigaptide or a composition thereof, a pharmaceutically acceptable salt of rotigaptide or a composition thereof, comprising the separation method of the first aspect;
[0058] In some embodiments of the present application, the detection method is a detection method of substance A in rotigaptide or a composition thereof.
[0059] In some embodiments of the present application, the detection method is a detection method of substance A in rotigaptide raw material.
[0060] In some embodiments of the present application, the detection method further comprises: using ultraviolet detection to determine the content of the substance A; and the detection wavelength of the ultraviolet detection is preferably 218-222 nm, and more preferably 220 nm.
[0061] In some embodiments of the present application, the chromatographic conditions are as follows:
[0062] The chromatographic column is a chromatographic column with octadecyl-bonded silica or octyl-bonded silica as the filler;
[0063] Column temperature: 25-35℃;
[0064] Injection volume: 5-20 μL;
[0065] Mobile phase: A: phosphate buffer, B: acetonitrile, gradient elution with volume percentage as shown in the table below;
[0066]
[0067]
[0068] Flow rate: 1.0-1.5 ml / min;
[0069] UV detector detection wavelength: 218-222 nm.
[0070] The third aspect of the present application provides a composition comprising rivastigmine or a pharmaceutically acceptable salt thereof and substance A, wherein the substance A comprises: a compound represented by formula L or a salt thereof.
[0071]
[0072] The content of the compound represented by formula L or a salt thereof is less than or equal to 0.05 wt%; preferably less than or equal to 0.03 wt%; more preferably less than or equal to 0.015 wt%.
[0073] In some embodiments of the present application, the purity of the rivastigmine or a pharmaceutically acceptable salt thereof is greater than or equal to 99.5 wt%, preferably greater than or equal to 99.9 wt%.
[0074] In some embodiments of the present application, the maximum single impurity content in the composition, except for the compound represented by formula L or a salt thereof, is less than or equal to 0.03 wt%; preferably less than or equal to 0.02 wt%; more preferably less than or equal to 0.015 wt%.
[0075] The substance A of the third aspect of the present application is the substance A of the first aspect.
[0076] In some embodiments of the present application, it is composed of rivastigmine or a pharmaceutically acceptable salt thereof and substance A.
[0077] In some embodiments of the present application, the content of the impurity B or a salt thereof, the impurity C or a salt thereof, the impurity D or a salt thereof, the impurity E, the impurity F or a salt thereof, the impurity G or a salt thereof, the impurity H or a salt thereof, the impurity I or a salt thereof, the impurity J or a salt thereof, and the impurity K is less than or equal to 0.03 wt%; preferably less than or equal to 0.02 wt%; more preferably less than or equal to 0.015 wt%.
[0078] In some embodiments of the present application, the content of the impurity B, the impurity C, the impurity D, the impurity E, the impurity F, the impurity G, the impurity H, the impurity I, the impurity J, the impurity K, respectively, is less than or equal to 0.03wt%; preferably less than or equal to 0.02wt%; more preferably less than or equal to 0.015wt%.
[0079] In some embodiments of the present application, the content of the impurity B or its salt in the composition is less than or equal to 0.03wt%; preferably less than or equal to 0.02wt%; more preferably less than or equal to 0.015wt%; further preferably less than or equal to 0.005wt%.
[0080] In some embodiments of the present application, the content of the impurity C or its salt in the composition is less than or equal to 0.03wt%; preferably less than or equal to 0.02wt%; more preferably less than or equal to 0.015wt%; further preferably less than or equal to 0.005wt%.
[0081] In some embodiments of the present application, the content of the impurity D or its salt in the composition is less than or equal to 0.03wt%; preferably less than or equal to 0.02wt%; more preferably less than or equal to 0.015wt%; further preferably less than or equal to 0.004wt%.
[0082] In some embodiments of the present application, the content of the impurity E in the composition is less than or equal to 0.03wt%; preferably less than or equal to 0.02wt%; more preferably less than or equal to 0.015wt%; further preferably less than or equal to 0.006wt%.
[0083] In some embodiments of the present application, the content of the impurity F or its salt in the composition is less than or equal to 0.03wt%; preferably less than or equal to 0.02wt%; more preferably less than or equal to 0.015wt%; further preferably less than or equal to 0.006wt%.
[0084] In some embodiments of the present application, the content of the impurity G or its salt in the composition is less than or equal to 0.03wt%; preferably less than or equal to 0.02wt%; more preferably less than or equal to 0.015wt%; further preferably less than or equal to 0.004wt%.
[0085] In some embodiments of the present application, the content of the impurity H or its salt in the composition is less than or equal to 0.03wt%; preferably less than or equal to 0.02wt%; more preferably less than or equal to 0.015wt%; further preferably less than or equal to 0.005wt%.
[0086] In some embodiments of the present application, the content of impurity I or its salt in the composition is less than or equal to 0.03wt%; preferably less than or equal to 0.02wt%; more preferably less than or equal to 0.015wt%; further preferably less than or equal to 0.004wt%.
[0087] In some embodiments of the present application, the content of impurity J or its salt in the composition is less than or equal to 0.03wt%; preferably less than or equal to 0.02wt%; more preferably less than or equal to 0.015wt%; further preferably less than or equal to 0.006wt%.
[0088] In some embodiments of the present application, the content of impurity K in the composition is less than or equal to 0.03wt%; preferably less than or equal to 0.02wt%; more preferably less than or equal to 0.015wt%; further preferably less than or equal to 0.002wt%.
[0089] The fourth aspect of the present application provides a pharmaceutical preparation comprising the composition of the third aspect, and one or more pharmaceutically acceptable excipients.
[0090] The fifth aspect of the present application provides the use of substance A as an impurity standard or control in the quality inspection or quality control of rotigaptide or its composition, rotigaptide pharmaceutical salt or its composition, or pharmaceutical preparation containing rotigaptide or its pharmaceutical salt; the substance A comprises: a compound represented by formula L or its salt.
[0091] In some embodiments of the present application, the composition is the composition of the third aspect.
[0092] In some embodiments of the present application, the pharmaceutical preparation is the pharmaceutical preparation of the fourth aspect.
[0093] The beneficial effects of the present application are as follows:
[0094] (1) The pH of the mobile phase in the separation and detection method provided by the present application is crucial for the separation of chromatographic peaks. Within the pH range of the mobile phase of the present application, structurally similar compounds can be effectively separated, substance A in rotigotine or its composition, rotigotine pharmaceutical salt or its composition can be effectively separated, and the peak shape can be improved, solving the problem of accurate quantification of substance A in rotigotine sample, and ensuring the effective detection of substance A in rotigotine or its composition, rotigotine pharmaceutical salt or its composition. Substance A includes a compound represented by formula L or a salt thereof, i.e. the present application realizes the effective detection of the compound represented by formula L. Preferably, the present application is aimed at the separation and detection of substance A in rotigotine or its composition, which can realize the accurate quantification of the compound represented by formula L. Further, the rotigotine used in the present application is high-purity rotigotine, which ensures the effective detection of the compound represented by formula L in high-purity rotigotine.
[0095] (3) The separation and detection method of the present application is simple to operate, has a wide range of selectable chromatographic columns, and has a wide range of mobile phase pH, column temperature, flow rate and injection volume, and has good durability.
[0096] (4) The present application provides the use of substance A as an impurity standard or control in the quality inspection or quality control of the composition or pharmaceutical preparation. The content of the compound represented by formula L or its salt in rotigotine or its composition, rotigotine pharmaceutical salt or its composition is low, the purity of the rotigotine pharmaceutical salt is high, the product quality of rotigotine or its composition, rotigotine pharmaceutical salt or its composition is improved, which is conducive to the control and improvement of drug quality and safety.
[0097] Further, the separation and detection method of the present application can separate and detect rotigotine sample and its pharmaceutical salt.
[0098] Further, the present application not only realizes the separation and detection of the compound represented by formula L or its salt, but also realizes the separation of a plurality of impurities (impurities B-K or their salts). The plurality of impurities can achieve baseline separation, the separation degree between each impurity and the pharmaceutical ingredient (rotigotine or its pharmaceutical salt) is large, the specificity is better, the peak shape is symmetrical, each impurity has a large absorption under the detection wavelength of the present application, and the requirement for instrument equipment is low, the method has good durability, good sensitivity, good injection precision and good repeatability.
[0099] (5) Compared with the European Pharmacopoeia, the present application can effectively detect more impurities, has better specificity, higher sensitivity and more symmetrical peak shape, and provides a basis for the development of quality standards for rotigotine or its composition, rotigotine pharmaceutical salt or its composition.
[0100] (6)The prior art (Liu Zhuolin. Research on Related Substances of Rotigotine Bulk Drug [D]. Yantai: Yantai University, 2020) discloses the detection of impurities in rotigotine starting material, intermediate, final product and bulk drug. The chromatographic conditions of the disclosed detection methods are different. In the detection method disclosed in the prior art related to impurity 35, different mobile phase conditions are used than in the present application. Compared with the prior art, the present application has a wider range of chromatographic column, a wider range of pH, column temperature and flow rate, and better method durability. In the prior art, only impurity 35 in the intermediate and final product is controlled as an unknown single impurity, and the single impurity limit is only ≤0.10%. However, the present application can effectively control the compound represented by formula L or its salt in rotigotine or its composition, rotigotine pharmaceutical salt or its composition, has higher detection sensitivity, and the sample purity is also very high. Further, compared with the prior art, the present application can also effectively control the various impurities (impurities B-K) in the European Pharmacopoeia, effectively separate each peak, and each peak has good symmetry. BRIEF DESCRIPTION OF DRAWINGS
[0101] Figure 1 Specificity spectrum of system suitability solution in Example 1;
[0102] Figure 2 Specificity spectrum of system suitability solution in Example 2;
[0103] Figure 3 Specificity spectrum of system suitability solution in Example 3;
[0104] Figure 4 Specificity spectrum of system suitability solution in Example 4;
[0105] Figure 5 Specificity spectrum of system suitability solution 1 in Example 5;
[0106] Figure 6 Specificity spectrum of system suitability solution 2 in Example 5;
[0107] Figure 7 Specificity spectrum of system suitability solution 3 in Example 5;
[0108] Figure 8 Specificity spectrum at a flow rate of 1.5 ml / min and a column temperature of 35℃ in Example 6;
[0109] Figure 9 Specificity spectrum at a flow rate of 1.0 ml / min and a column temperature of 25℃ in Example 6;
[0110] Figure 10 Specificity spectrum at pH 7.4 in Example 7;
[0111] Figure 11 Specificity profile for Example 7 at pH 7.8;
[0112] Figure 12 Specificity profile for Comparative Example 1 System Suitability Solution 1;
[0113] Figure 13 European Pharmacopoeia System Suitability Solution Reference for Comparative Example 1. DETAILED DESCRIPTION
[0114] In order to make the present application more readily understood, reference is made to the following examples, which are intended to be illustrative only and not limiting of the scope of the application.
[0115] The structures of the compound L, impurity B, impurity C, impurity D, impurity E, impurity F, impurity J, impurity H, impurity I, impurity G, impurity K and rotigaptide involved in the embodiments of the present application are shown in the following table:
[0116] Table 1
[0117]
[0118]
[0119] The impurities B to K are all impurities in the European Pharmacopoeia.
[0120] The European Pharmacopoeia referred to in the present application is European Pharmacopoeia 10.0 3758-3750.
[0121] In the embodiments of the present application, the conditions not specified are carried out according to the conventional conditions or the conditions recommended by the manufacturers. The reagents or instruments not specified by the manufacturers are all conventional products available in the market.
[0122] The reference substance of the compound L (impurity L) shown in the formula L, the reference substances of the impurities B to K, rotigaptide and its pharmaceutical salt samples in the embodiments of the present application are all commercially available products.
[0123] The reference substance of the compound L (impurity L) shown in the formula L in the embodiments of the present application is the hydrobromide of the compound L shown in the formula L, which is hereinafter referred to as the reference substance of the compound L.
[0124] High performance liquid chromatograph: Agilent 1100 (Agilent Technologies (China) Co., Ltd.), Shimazdu 2030 (Shimadzu Enterprise Management (China) Co., Ltd.); chromatographic column: shim-pack VP-ODS, 4.6 mm x 250 mm, 5 μm (Shimadzu Enterprise Management (China) Co., Ltd.), Agilent XDB C8, 4.6 mm x 150 mm, 5 μm (Agilent Technologies (China) Co., Ltd.).
[0125] Example 1
[0126] A method for separating and detecting a rofecoxib related substance, comprising the following steps:
[0127] Structure confirmation of compound L: compound L is (S)-6-(n-propyl(2-(thiophen-3-yl)ethyl)amino)-5,6,7,8-tetrahydronaphthalen-1-ol, and the nuclear magnetic resonance spectrum is as shown in the following table.
[0128] Table 2
[0129]
[0130]
[0131] Nuclear magnetic resonance structure number is as follows:
[0132]
[0133] Sample preparation:
[0134] Take 10.37 mg of compound L reference substance and place it in a 10 ml volumetric flask, add diluent (0.5 VOL% phosphoric acid solution-acetonitrile (phosphoric acid solution and acetonitrile in a volume ratio of 85:15)) to dissolve and dilute to the mark, shake well, then take 1.5 ml of the solution to a 100 ml volumetric flask, dilute to the mark with diluent, shake well, and use as a compound L stock solution. Take 10.10 mg of rofecoxib sample and place it in a 10 ml volumetric flask, add 1 ml of compound L stock solution to it, and dissolve and dilute to the mark with the above diluent, as a system suitability solution.
[0135] Chromatographic conditions:
[0136] Agilent 1100 high performance liquid chromatograph was used, and a chromatographic column shim-pack VP-ODS (4.6 mm x 250 mm, 5 μm) with octadecyl bonded silica as the filler was selected; acetonitrile-phosphate buffer (mobile phase: A: phosphate buffer, which is obtained by adjusting 10 mmol / L KH2PO4 solution with phosphoric acid to pH 6.0; B: acetonitrile) was used as the mobile phase, and the following chromatographic conditions were used for separation:
[0137] The flow rate was 1.2 ml / min;
[0138] The column temperature was 30℃;
[0139] The UV detector wavelength was 220 nm;
[0140] The injection volume was 10 μL;
[0141] The gradient elution conditions in the mobile phase, calculated by volume percentage, were as follows:
[0142] Table 3
[0143]
[0144] The chromatographic separation results were shown in Figure 1 The specific information of the chromatographic peaks of rotigotine and compound L was shown in the following table:
[0145] Table 4
[0146]
[0147] Example 2
[0148] A separation and detection method of rotigotine related substances was as follows:
[0149] The system suitability solution in Example 1 was taken, and an Agilent 1100 high performance liquid chromatograph was used, and a chromatographic column Agilent XDB C8 (4.6 mm x 150 mm, 5 μm) with octyl bonded silica gel as the filler was selected; acetonitrile-phosphate buffer (mobile phase: A: phosphate buffer, which was obtained by adjusting 10 mmol / L diammonium hydrogen phosphate solution with phosphoric acid to pH 6.0; B: acetonitrile) was used as the mobile phase, and the following chromatographic conditions were used for separation:
[0150] The flow rate was 1.2 ml / min;
[0151] The column temperature was 30℃;
[0152] The UV detector wavelength was 220 nm;
[0153] The injection volume was 10 μL;
[0154] The gradient elution conditions in the mobile phase, calculated by volume percentage, were as follows:
[0155] Table 5
[0156]
[0157]
[0158] The chromatographic separation results were shown inFigure 2 The specific information of the chromatographic peaks of rotigotine and compound L is shown in the following table:
[0159] Table 6
[0160]
[0161] Example 3
[0162] A method for detecting a rotigotine impurity, the steps of which are as follows:
[0163] The system suitability solution in Example 1 was taken, and an Agilent 1100 high performance liquid chromatograph was used, with an Agilent XDB C8 (4.6 mm x 150 mm, 5 μm) chromatographic column with octyl-bonded silica gel as the filler; acetonitrile-phosphate buffer (mobile phase: A: phosphate buffer, which is obtained by adjusting 10 mmol / L diammonium hydrogen phosphate solution with phosphoric acid to pH 7.0; B: acetonitrile) was used as the mobile phase, and the following chromatographic conditions were used for separation:
[0164] The flow rate was 1.2 ml / min;
[0165] The column temperature was 30°C;
[0166] The wavelength of the ultraviolet detector was 220 nm;
[0167] The injection volume was 10 μL;
[0168] The gradient elution conditions in the mobile phase, calculated as the volume percentage, are as follows:
[0169] Table 7
[0170]
[0171]
[0172] The chromatographic separation results are shown in Figure 3 The specific information of the chromatographic peaks of rotigotine and compound L is shown in the following table:
[0173] Table 8
[0174]
[0175] Example 4
[0176] A method for detecting a rotigotine impurity, the steps of which are as follows:
[0177] Take the system suitability solution in Example 1, use Agilent 1100 high performance liquid chromatograph, select octyl bonded silica gel as the filler of the chromatographic column Agilent XDB C8 (4.6 mm x 150 mm, 5 μm); with acetonitrile-phosphate buffer (A: phosphate buffer, which is obtained by adjusting 10 mmol / L of dihydrogen phosphate solution with phosphoric acid to pH 8.0; B: acetonitrile) as the mobile phase, the following chromatographic conditions are used for separation:
[0178] The flow rate is 1.2 ml / min;
[0179] The column temperature is 30°C;
[0180] The UV detector wavelength is 220 nm;
[0181] The injection volume is 10 μL;
[0182] In the mobile phase, the gradient elution conditions are as follows:
[0183] Table 9
[0184]
[0185] The chromatographic separation results are shown in Figure 4 (the system suitability solution specificity spectrum) as shown, the specific information of the chromatographic peaks of rotigotine and compound L is as follows:
[0186] Table 10
[0187]
[0188] According to Examples 1-4, using octadecyl or octyl bonded silica gel as the filler of the chromatographic column, under the condition of pH 6.0-8.0 of the mobile phase, rotigotine and compound L can be well separated, and the main peak shape is symmetrical, the symmetry factor is greater than 0.5, the symmetry factor of compound L can be 0.54-0.76, and the symmetry factor of rotigotine can be 0.54-0.83.
[0189] Example 5
[0190] A method for detecting a rotigotine impurity, the steps of which are as follows:
[0191] Sample preparation:
[0192] System suitability solution 1: same as the system suitability solution in Example 1.
[0193] System Suitability Solution 2: Weigh appropriate amounts of reference standards of Impurity B, Impurity C, Impurity D, Impurity E, Impurity G, Impurity H, Impurity I, Impurity K and Rotigaptin, dissolve and dilute with diluent (0.5 VOL% Phosphoric acid solution - Acetonitrile (85:15 by volume of Phosphoric acid solution and Acetonitrile)) to make a mixture solution containing about 1.5 μg of Impurity B, 2 μg of Impurity C, 1 μg of Impurity D, 0.3 μg of Impurity E, 2 μg of Impurity G, 1 μg of Impurity H, 1 μg of Impurity I, 1 μg of Impurity K and 1 mg of Rotigaptin per 1 mg.
[0194] System Suitability Solution 3: Weigh appropriate amounts of reference standards of Impurity B, Impurity C, Impurity D, Impurity F, Impurity G, Impurity I and Impurity J, dissolve and dilute with diluent (0.5 VOL% Phosphoric acid solution - Acetonitrile (85:15 by volume of Phosphoric acid solution and Acetonitrile)) to make a mixture solution containing about 1.5 μg of Impurity B, 2 μg of Impurity C, 1 μg of Impurity D, 2 μg of Impurity G, 1 μg of Impurity F, 1 μg of Impurity I and 1 μg of Impurity J per 1 mg.
[0195] Limit of Detection Solution: Dissolve and dilute reference standards of Impurity B, Impurity C, Impurity D, Impurity E, Impurity F, Impurity G, Impurity H, Impurity I, Impurity J, Impurity K, Compound L with diluent (0.5 VOL% Phosphoric acid solution - Acetonitrile (85:15 by volume of Phosphoric acid solution and Acetonitrile)) to the respective limit of detection concentration.
[0196] Test Solution 1: Take an appropriate amount of Rotigaptin sample, dissolve and dilute with diluent (0.5 VOL% Phosphoric acid solution - Acetonitrile (85:15 by volume of Phosphoric acid solution and Acetonitrile)) to make a solution of about 1 mg / ml.
[0197] Test Solution 2: Take an appropriate amount of Rotigaptin hydrochloride sample, dissolve and dilute with diluent (0.5 VOL% Phosphoric acid solution - Acetonitrile (85:15 by volume of Phosphoric acid solution and Acetonitrile)) to make a solution of about 2 mg / ml.
[0198] Test Solution 3: Take an appropriate amount of Rotigaptin hydrobromide sample, dissolve and dilute with diluent (0.5 VOL% Phosphoric acid solution - Acetonitrile (85:15 by volume of Phosphoric acid solution and Acetonitrile)) to make a solution of about 0.5 mg / ml.
[0199] Reference Solution: Weigh appropriate amounts of reference standards of Impurity B, Impurity C, Impurity D, Impurity G, Impurity H, Impurity I and Impurity K, dissolve and dilute with diluent (0.5 VOL% Phosphoric acid solution - Acetonitrile (85:15 by volume of Phosphoric acid solution and Acetonitrile)) to make a mixture solution containing about 1.5 μg of Impurity B, 2 μg of Impurity C, 1 μg of Impurity D, 2 μg of Impurity G, 1 μg of Impurity H, 1 μg of Impurity I and 1 μg of Impurity K per 1 mg.
[0200] Test sample spiked solution: 10 mg of rotigotine sample was weighed, dissolved with the control solution and quantitatively diluted to 10 ml, and 6 samples were prepared in parallel.
[0201] Chromatographic conditions:
[0202] An Agilent 1100 high performance liquid chromatograph was used, and an Agilent XDB C8 (4.6 mm x 150 mm, 5 μm) chromatographic column with octyl-bonded silica gel as the filler was selected. Acetonitrile-phosphate buffer (A: phosphate buffer, which is obtained by adjusting 10 mmol / L of a dihydrogen ammonium phosphate solution with phosphoric acid to pH 7.6; B: acetonitrile) was used as the mobile phase, and the following chromatographic conditions were used for separation:
[0203] The flow rate was 1.2 ml / min;
[0204] The column temperature was 30°C;
[0205] The wavelength of the ultraviolet detector was 220 nm;
[0206] The injection volume was 5-20 μL;
[0207] The gradient elution conditions in the mobile phase, calculated according to the volume percentage, were as follows:
[0208] Table 11
[0209]
[0210] The injection volume of test sample solution 2 was 5 μL, the injection volume of test sample solution 3 was 20 μL, and the injection volume of the rest of the samples was 10 μL.
[0211] The chromatographic separation results of the system suitability solution are shown in Figures 5 to 7 (the specificity chromatograms of system suitability solutions 1-3), and the specific information of the chromatographic peaks of rotigotine and each impurity is shown in the following table. The peak shapes of each impurity and rotigotine are relatively symmetrical (the symmetry factor is greater than 0.5), while the peak shape in the European Pharmacopoeia is severely tailing (such as the comparative example Figure 12 ), and the separation degree between each peak is obviously greater than 1.5. The method can effectively separate the impurities (impurities B-K) and compound L in the European Pharmacopoeia. The separation degree of impurity H (EP-H) and impurity G (EP-G) in the European Pharmacopoeia is not less than 1.5 as the requirement for system suitability, and the method under this example is obviously superior to the requirement of the European Pharmacopoeia.
[0212] Table 12
[0213]
[0214] According to the detection limit of each impurity, as shown in the following table, the worst can reach 0.03wt% of the test sample concentration (1mg / ml), while the method in the European Pharmacopoeia has a large baseline noise (comparative example Figure 13 ), and the limit of neglect is 0.05wt%, and the detection sensitivity of the method of the present application is high.
[0215] Table 13
[0216] Impurity Signal to noise ratio Concentration concentration (μg / ml) Relative to test sample concentration Impurity B 5.6 0.051 0.005 wt% Impurity C 6.6 0.045 0.005 wt% Impurity D 11.1 0.040 0.004 wt% Impurity E 8.4 0.257 0.03 wt% Impurity F 4.0 0.062 0.006 wt% Impurity G 4.0 0.044 0.004 wt% Impurity H 4.4 0.048 0.005 wt% Impurity I 4.4 0.042 0.004 wt% Impurity J 7.6 0.063 0.006 wt% Impurity K 7.0 0.015 0.002 wt% Compound L 6.7 0.042 0.004 wt%
[0217] The relative standard deviation of the peak area of each impurity of the control solution is shown in Table 14, and the RSD is less than 3.0%, indicating that the precision of the method is good. The average spiked recovery rate and the RSD of the spiked recovery rate of each impurity in the 6 test sample spiked solutions are shown in Table 14, and the average spiked recovery rate is between 90% and 108%, and the RSD of the spiked recovery rate is less than 3.0%, indicating that the repeatability and accuracy of the method are high.
[0218] Table 14
[0219]
[0220] In this example, the samples of rotigotine and rotigotine pharmaceutical salt were detected, and according to the detection results of each batch, as shown in the following table, the impurities were all less than 0.05wt%, and the purity was greater than 99.5wt%, indicating that the purity of the samples of rotigotine and rotigotine pharmaceutical salt was very high.
[0221] Table 15
[0222]
[0223] Example 6
[0224] A method for detecting a rotigotine related substance, the steps of which are as follows:
[0225] Sample preparation:
[0226] System suitability solution: take impurities B, C, D, G, H, I, K and rotigotine control each appropriate amount, dissolve and dilute with diluent (0.5VOL% phosphoric acid solution-acetonitrile (phosphoric acid solution and acetonitrile volume ratio 85:15)) to prepare a mixed solution containing about 10μg impurity B, 10μg impurity C, 10μg impurity D, 15μg impurity G, 15μg impurity H, 5μg impurity I, 20μg impurity K and 1mg rotigotine per 1mg.
[0227] Chromatographic conditions: on the basis of Example 5, investigate the temperature and flow rate resistance
[0228] Shimadzu 2030 high performance liquid chromatograph, using octyl bonded silica gel as the filler of the chromatographic column Agilent XDB C8 (4.6mmx150mm, 5μm); using acetonitrile-phosphate buffer (A: phosphate buffer, which is obtained by adjusting 10mmol / L of dihydrogen ammonium phosphate solution with phosphoric acid to pH 7.6; B: acetonitrile) as the mobile phase, the following chromatographic conditions are used for separation:
[0229] The flow rate is: ①1.5ml / min; ②1.0ml / min
[0230] The column temperature is: ①35℃; ②25℃
[0231] The wavelength of the ultraviolet detector is 220nm;
[0232] The injection volume is 10μL;
[0233] In the mobile phase, the gradient elution conditions are as follows, calculated by volume percentage:
[0234] Table 16
[0235]
[0236] After adjusting the flow rate and column temperature, the chromatographic separation results of each component are shown in Figure 8 (the specificity spectrum under the condition of flow rate 1.5ml / min, column temperature 35℃), Figure 9 (the specificity spectrum under the condition of flow rate 1.0ml / min, column temperature 25℃) shown, the chromatographic peak of rofecoxib and each impurity is as shown in the following table, each peak can still be effectively separated. It is proved that the method of the present application has good durability to column temperature and flow rate.
[0237] Table 17
[0238]
[0239]
[0240] Note: Compound L is contained in the rofecoxib reference substance.
[0241] Example 7
[0242] A method for detecting rofecoxib related substances, the steps are as follows:
[0243] Sample preparation:
[0244] System suitability solution: accurately weigh appropriate amounts of the reference substances of impurity B, impurity C, impurity D, impurity K, impurity G, impurity H, impurity I and rotigaptide, dissolve and dilute with diluent (0.5 VOL% phosphoric acid solution-acetonitrile (phosphoric acid solution and acetonitrile in a volume ratio of 85:15)) to prepare a mixed solution containing about 1.5 μg of impurity B, 2 μg of impurity C, 1 μg of impurity D, 2 μg of impurity G, 1 μg of impurity H, 1 μg of impurity I, 1 μg of impurity K and 1 mg of rotigaptide per 1 mg.
[0245] Chromatographic conditions: adjust the pH value of the buffer on the basis of Example 5
[0246] A Shimadzu 2030 high performance liquid chromatograph was used, and an Agilent XDB C8 (4.6 mm x 150 mm, 5 μm) chromatographic column with octyl-bonded silica gel as the filler was selected. Acetonitrile-phosphate buffer (A: phosphate buffer, which is obtained by adjusting the pH of a 10 mmol / L solution of diammonium hydrogen phosphate with phosphoric acid to ① 7.4, ② 7.8; B: acetonitrile) was used as the mobile phase, and the following chromatographic conditions were used for separation:
[0247] The flow rate was 1.2 ml / min;
[0248] The column temperature was 30°C;
[0249] The wavelength of the ultraviolet detector was 220 nm;
[0250] The injection volume was 10 μL;
[0251] The gradient elution conditions in the mobile phase, calculated as the volume percentage, were as follows:
[0252] Table 18
[0253]
[0254]
[0255] After adjusting the pH value of the buffer, the chromatographic separation results of the components are shown in Figure 10 (pH 7.4 specificity spectrum), Figure 11 (pH 7.8 specificity spectrum), and the chromatographic peak information of rotigaptide and each impurity is as follows. The buffer salt is between pH 7.4 and 7.8, and rotigaptide and each impurity peak can still be effectively separated, and the symmetry of each peak is still good (the tailing factor is less than 2.0). It is shown that the separation and detection method of the application has strong pH resistance.
[0256] Table 19 Chromatographic separation results at pH 7.4
[0257] Compound Retention time (min) Minimum resolution Theoretical plates Tailing factor Impurity B 7.449 31.5 33175 1.5 Impurity C 12.588 9.3 94599 1.2 Impurity D 16.541 6.1 108282 1.1 Impurity G 23.655 3.1 184220 1.1 Impurity H 27.291 9.4 169941 1.0 Impurity I 29.777 9.4 202113 1.0 Impurity K 14.036 9.3 142232 1.2 Compound L 17.825 6.1 106821 1.1 Roglitazone 20.033 10.9 179103 1.2
[0258] Table 20 Chromatographic separation results at pH 7.8
[0259] Compound Retention time (min) Minimum resolution Theoretical plates Tailing factor Impurity B 7.966 32.4 37599 1.6 Impurity C 13.229 6.0 107168 1.2 Impurity D 17.311 6.6 114022 1.0 Impurity G 23.957 3.6 185183 1.1 Impurity H 27.923 9.1 167761 0.6 Impurity I 30.365 9.1 206388 0.9 Impurity K 14.162 6.0 142254 1.2 Compound L 18.760 6.6 101665 1.3 Roglitazone 20.761 10.1 267978 1.2
[0260] Note: Compound L is contained in the Rotigotine reference substance.
[0261] Example 8
[0262] A detection method of Rotigotine related substances is as follows:
[0263] Sample preparation:
[0264] System suitability solution: same as the system suitability solution under Example 7
[0265] Chromatographic conditions: adjust the detection wavelength based on Example 5
[0266] A Shimadzu 2030 high performance liquid chromatograph was used, and an Agilent XDB C8 (4.6 mm x 150 mm, 5 μm) chromatographic column with octyl bonded silica as the filler was selected. Acetonitrile-phosphate buffer (mobile phase: A: phosphate buffer, which is a 10 mmol / L diammonium hydrogen phosphate solution adjusted by phosphoric acid to pH 7.6; B: acetonitrile) was used as the mobile phase, and the following chromatographic conditions were used for separation:
[0267] The flow rate was 1.2 ml / min;
[0268] The column temperature was 30°C;
[0269] The UV detector wavelength was 218 nm, 222 nm;
[0270] The injection volume was 10 μL;
[0271] The gradient elution conditions in the mobile phase, calculated as a percentage by volume, were as follows:
[0272] Table 21
[0273]
[0274] At different detection wavelengths, the theoretical plate numbers of Rotigotine and each impurity peak were all high, all greater than 5000, indicating that when the wavelength has a certain change, the response of Rotigotine and each impurity is still good.
[0275] Table 22 Detection results at different wavelengths
[0276]
[0277]
[0278] Note: Compound L is contained in rotigotine reference standard.
[0279] Comparative Example
[0280] A method for detecting rotigotine impurities, referring to the related substances analysis method in the European Pharmacopoeia (EP10.0), comprises the following steps:
[0281] Using the system suitability solution from Example 1, an Agilent 1100 high-performance liquid chromatograph was used with an Agilent XDB C8 column (4.6 mm × 150 mm, 5 μm) packed with octyl-bonded silica gel. Separation was performed under the following chromatographic conditions using 0.03 VOL% trifluoroacetic acid solution (mobile phase A) - 0.02 VOL% trifluoroacetic acid acetonitrile solution (mobile phase B):
[0282] The flow rate was 2.0 ml / min;
[0283] The column temperature is 40℃;
[0284] The wavelength of the ultraviolet detector is 220 nm;
[0285] The injection volume was 10 μL;
[0286] In the mobile phase, the gradient elution conditions, calculated by volume percentage, are as follows:
[0287] Table 23
[0288]
[0289] In an acidic mobile phase (pH 2–3), the system's suitability for the solution ( Figure 12 In the specific spectrum of solution 1 (system suitability), rotigotine (9.410 min) completely encapsulated compound L, and the two could not be effectively separated. Furthermore, the symmetry factor of the rotigotine peak was 0.20, the peak tailing was severe, the theoretical plate number was 2776, and the column efficiency was poor.
[0290] Reference chart of system suitability in the European Pharmacopoeia ( Figure 13 European Pharmacopoeia System Suitability Solution Reference Diagram Figure 13 In the diagram, Impurity B is impurity B, Impurity C is impurity C, Impurity G is impurity G, and Impurity H is impurity H. Impurity G (impurity G) and Impurity H (impurity H) show baseline separation, but with significant baseline noise. Using this method to test the rotigotine test solution in Example 5, no impurities were detected in any batches. However, the method's ability to separate and detect impurities is inferior to that of the method of this invention.
[0291] In summary, the method for analyzing related substances of rotigotine according to the present application has good specificity, symmetrical peak shape and good detection sensitivity. The rotigotine sample detected by the method has high purity, and the purity is greater than 99.5 wt%, and the detection amount of the compound L is very small, less than 0.05 wt%. The applicant finds that under acidic conditions, the peak of rotigotine is easy to tail, when the pH value of the mobile phase is closer to the pKa (acid-base dissociation constant) of rotigotine, the peak shape of rotigotine can be improved, the retention is enhanced, and substances similar in structure to rotigotine can be separated. Generally, because the chromatographic column is not resistant to high pH, the pH of the mobile phase is usually acidic or neutral, and the applicant of the present application selects a neutral to alkaline (pH value is 6.0-8.0, preferably pH value is 7.4-7.8) aqueous phase as the mobile phase A and an organic phase as the mobile phase B, so that the pH value of the mobile phase is close to the pKa of rotigotine. The above-mentioned effects are achieved by using the mobile phase according to the present application and preferably using an alkali-resistant chromatographic column (chromatographic column with octadecyl bonded silica or octyl bonded silica as the filler).
[0292] The above-mentioned is only the preferred embodiment of the present application. It should be pointed out that for those skilled in the art, under the technical inspiration provided by the present application, other equivalent variants and improvements as the common knowledge in the art can also be made, and should also be considered as the protection scope of the present application.
Claims
1. A method for separating material A from rotigotine, a pharmaceutically acceptable salt of rotigotine, characterized in that, Comprising: The rosiglitazone, the rosiglitazone medicinal salt, and the chromatographic column with octyl bonded silica gel as the filler are used for high performance liquid chromatography separation, the mobile phase in the high performance liquid chromatography is: a mobile phase A inorganic salt buffer and a mobile phase B acetonitrile, the pH value of the mobile phase A is 7.4-7.8; the gradient elution conditions are as follows in terms of volume percentage: The substance A comprises: a compound represented by formula L or a salt thereof. The substance A further comprises one or more of impurity D or a salt thereof, impurity E, impurity H or a salt thereof, and impurity K, and the structural formulae of the impurity D, the impurity E, the impurity H, and the impurity K are as follows:
2. The separation method of claim 1, wherein, The inorganic salt is one or more of disodium hydrogen phosphate, dipotassium hydrogen phosphate, diammonium hydrogen phosphate.
3. The separation method according to claim 1 or 2, characterized in that, The separation method further comprises: using the following chromatographic conditions for high performance liquid chromatography separation: flow rate: 1.0-1.5 ml / min; and / or column temperature: 25-35 DEG C; and / or injection volume: 5-20 mu L.
4. A method for detecting material A in rotigotine, a pharmaceutically acceptable salt of rotigotine, characterized in that, The separation method comprises any one of claims 1-3.
5. The detection method according to claim 4, characterized in that, The detection method further comprises: using ultraviolet detection to determine the content of the substance.
6. The detection method according to claim 5, characterized in that, The detection wavelength of the ultraviolet detection is 218-222 nm.
7. The detection method according to claim 6, characterized in that, The detection wavelength of the ultraviolet detection is 220 nm.