A method for determining related substances of cis-tetrahydrocarboline hydrochloride, an intermediate of tadalafil

Abstract

The invention discloses a method for determining related substances of cis-tetrahydrocarboline hydrochloride, an intermediate of tadalafil, which specifically adopts a diode array detector and uses 0.1% formic acid solution-acetonitrile as a mobile phase for gradient elution. Take an appropriate amount of cis-tetrahydrocarboline hydrochloride, add a solvent (water: acetonitrile 1:1) to prepare a solution containing 0.4 mg of cis-tetrahydrocarboline hydrochloride per 1 ml, as a test solution. Take the test solution and inject it into a liquid chromatograph, record the chromatogram, and process the chromatogram according to the area normalization method. The detection method of related substances of cis-tetrahydrocarboline hydrochloride, an intermediate of tadalafil, can quickly and accurately detect the situation of related substances of cis-tetrahydrocarboline hydrochloride. The operation is simple and the sensitivity is high, and the product quality can be well controlled.

Description

Technical Field

[0001] The invention relates to a method for determining related substances of cis-tetrahydrocarboline hydrochloride, an intermediate of tadalafil, and belongs to the field of drug analysis and detection. Background Art

[0002] Tadalafil, chemical name is (6R-12aR)-6-(1,3-benzodioxol-5-yl)-2-methyl-2,3,6,7,12,12a-hexahydropyrazino[1',2'-1,6]-pyrido[3,4-b]indole-1,4-dione, white crystalline powder, poorly soluble in water. Its English common name is Tadalafil, and its trade names include Cialis, Cialis, and Cialis. Its structural formula is as follows:

[0003] .

[0004] Tadalafil is a selective and reversible inhibitor of cyclic guanosine monophosphate (cGMP)-specific phosphodiesterase 5 (PDE5) jointly developed by Exxon and Eli Lilly in the United States. It is a second-generation phosphodiesterase inhibitor that competitively inhibits the binding of cGMP and PDE5, inactivates PDE5, reduces the hydrolysis of cGMP, and increases the cGMP concentration to enhance erectile function.

[0005] Cis-tetrahydrocarboline hydrochloride is a key intermediate in the synthesis of tadalafil. Its chemical name is cis-(1R, 3R)-1,2,3,4-tetrahydro-1-(3,4-methylenedioxyphenyl)-9H-pyrido[3,4-B]indole-3-carboxylic acid methyl ester hydrochloride, and its structural formula is as follows:

[0006] HCl

[0007] The synthetic route of cis-tetrahydrocarboline hydrochloride is as follows:

[0008]

[0009] The impurities that may be produced are as follows:

[0010]

[0011] Impurity 1 (D-tryptophan);

[0012]

[0013] Impurity 2 (piperonal);

[0014]

[0015] Impurity 3 (D-tryptophan methyl ester hydrochloride);

[0016]

[0017] Impurity 4 (hydrolysis impurity);

[0018]

[0019] Impurity 5 (isomer impurity).

[0020] How to quickly and accurately detect the related substances of cis-tetrahydrocarboline hydrochloride is the key to controlling the product quality of tadalafil. Summary of the invention

[0021] Aiming at the problem of quality control of intermediates existing in the synthesis process of tadalafil at this stage, the present invention provides a method for determining related substances of cis-tetrahydrocarboline hydrochloride, an intermediate of tadalafil, and adopts a diode array detector to carry out related substance inspection on cis-tetrahydrocarboline hydrochloride, an intermediate of tadalafil.

[0022] The technical solution of the present invention is as follows:

[0023] A method for determining related substances of cis-tetrahydrocarboline hydrochloride, an intermediate of tadalafil, comprises the following steps:

[0024] (1) Chromatographic conditions: The detector is a diode array detector, the column temperature is 40°C, and the flow rate is 1.0 mL / min. The mobile phase A of the chromatographic conditions is 0.1% formic acid solution; the mobile phase B is acetonitrile, gradient elution; the theoretical plate number is not less than 3000 calculated based on the cis-tetrahydrocarboline hydrochloride peak;

[0025] (2) Preparation of test solution: Take an appropriate amount of cis-tetrahydrocarboline hydrochloride and add a solvent to prepare a solution as the test solution;

[0026] (3) Determination method: Inject the test sample solution into the liquid chromatograph, record the chromatogram, and process the chromatogram by the area normalization method.

[0027] Preferably, the gradient elution conditions in step (1) are:

[0028] .

[0029] Preferably, the liquid chromatography column in step (1) is made of octadecylsilane bonded silica gel.

[0030] Preferably, the solvent in step (2) is water:acetonitrile in a volume ratio of 1:1.

[0031] The method for detecting related substances of cis-tetrahydrocarboline hydrochloride, an intermediate of tadalafil, is suitable for detecting cis-tetrahydrocarboline hydrochloride products and monitoring of reaction liquids thereof, and can effectively control the quality of cis-tetrahydrocarboline hydrochloride.

[0032] The positive effects of the present invention are:

[0033] 1. The present invention uses a diode array detector to detect related substances in the product, and establishes an effective method for quickly and intuitively evaluating the purity of the chromatographic peak: the purity of the chromatographic peak can be intuitively judged, and the position of the interfering peak can be judged, all the spectral information "in flow" can be obtained, and the absorption spectrum of the chromatographic components can be quickly obtained. The advantage of the diode array detector over the general ultraviolet-visible detector is that it can not only perform qualitative analysis based on the retention time of the chromatogram, but also perform qualitative analysis based on the spectral information it provides, which greatly improves the credibility of the qualitative analysis.

[0034] 2. The detection method of the present invention is accurate, simple to operate, has good reproducibility, and high sensitivity. It can fully meet the inspection of relevant substances, ensure product quality, and has strong practicality in work. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 It is the liquid chromatogram of cis-tetrahydrocarboline hydrochloride and related substances. DETAILED DESCRIPTION

[0036] A method for determining related substances of cis-tetrahydrocarboline hydrochloride, an intermediate of tadalafil, comprises the following steps:

[0037] 1. Selection of chromatographic conditions:

[0038] Instrument: Agilent: 1260II, the best column temperature is 40℃, flow rate is 1.0ml / min. The liquid chromatography column uses octadecylsilane bonded silica gel as filler (250mm×4.6mm, 5µm), and various conditions such as methanol-water, acetonitrile-water, methanol-water-trifluoroacetic acid, methanol-acetonitrile-phosphate buffer solution are selected successively. Finally, the mobile phase is determined to be composed of 0.1% formic acid solution-acetonitrile, and linear gradient elution is performed according to the table below; injection volume is 20µl.

[0039]

[0040] Under the chromatographic conditions, the main peak of cis-tetrahydrocarboline hydrochloride has a moderate retention time and a good peak shape.

[0041] 2. Preparation of test solution:

[0042] Take an appropriate amount of cis-tetrahydrocarboline hydrochloride and add water / acetonitrile (1:1) to prepare a 0.4 mg / ml solution as the test solution.

[0043] 3. Determination method: Take 20μl of the test solution and inject it into the liquid chromatograph. Record the chromatogram to twice the retention time of the main peak of cis-tetrahydrocarboline hydrochloride. Process the chromatogram by area normalization method. The total amount of impurities shall not exceed 2%.

[0044] Example 2 Specificity Test

[0045] Accurately weigh appropriate amounts of this product, D-tryptophan, D-tryptophan methyl ester hydrochloride, piperonal, hydrolysis impurities and isomeric impurities, place in the same volumetric flask, add solvent to dissolve and quantitatively dilute to make a mixed solution containing 0.4 mg of cis-tetrahydrocarboline hydrochloride per 1 ml. Accurately measure 20 μl and inject into the liquid chromatograph to record the chromatogram ( Figure 1 ), the results showed that cis-tetrahydrocarboline hydrochloride was well separated from the impurities, and the impurities were well separated from each other.

[0046] The relative retention time of D-tryptophan is 0.24; the relative retention time of piperonal is 0.93; the relative retention time of D-tryptophan methyl ester hydrochloride is 0.34; the relative retention time of hydrolysis impurities is 0.87; and the relative retention time of isomer impurities is 1.05.

[0047] Example 3 Sensitivity measurement

[0048] Accurately weigh 10.44 mg of cis-tetrahydrocarboline hydrochloride, place it in a 25 ml volumetric flask, dissolve it with a solvent and dilute it to the mark, shake it well; accurately measure 2.5 ml of the above solution, place it in a 50 ml volumetric flask, accurately measure 0.2 ml, place it in a 50 ml volumetric flask, dilute it to the mark with a solvent, shake it well, and use it as the detection limit concentration solution. Accurately measure 20 μl and inject it into the liquid chromatograph to produce a signal with a main peak of three times the baseline noise. After testing, the minimum detection amount is 0.0829 μg / ml, which is approximately equivalent to 0.02% of the test solution concentration. The results show that this method has high sensitivity and can fully meet the requirements of the inspection and determination of related substances.

[0049] Example 4 Durability Test

[0050] The intermediate II test solution was used as the durability test solution. The column temperature, flow rate, and trifluoroacetic acid concentration in the mobile phase were investigated for durability, taking the main peak chromatographic purity and the separation degree between the main peak and the adjacent impurity peaks in the test solution as indicators. The results are as follows:

[0051] Table 1 Durability test results (flow rate)

[0052]

[0053] Table 2 Durability test results (column temperature)

[0054]

[0055] Table 3 Durability test results (formic acid concentration)

[0056]

[0057] The above test results show that this method is simple, sensitive, and has good reproducibility, and can be used to detect the quality of cis-tetrahydrocarboline hydrochloride.

Claims

1. A method for determining impurities produced in cis-tetrahydrocarboline hydrochloride, an intermediate of tadalafil, and its synthetic route, characterized in that: The following steps are involved: (1) Chromatographic conditions: The detector is a diode array detector, the column temperature is 40°C, and the flow rate is 1.0 mL / min. The liquid chromatography column is filled with octadecylsilane bonded silica gel, and the specifications are: 250 mm × 4.6 mm, 5 µm. The mobile phase A of the chromatographic conditions is 0.1% formic acid solution; the mobile phase B is acetonitrile, gradient elution; the injection volume is 20 µL; the theoretical plate number is calculated based on the cis-tetrahydrocarboline hydrochloride peak and is not less than 3000; wherein, the gradient elution conditions are: 0min, A-85%, B-15%; 3min, A-85%, B-15%; 21min, A-58%, B-42%; 26min, A-58%, B-42%; 36min, A-5%, B-95%; 40min, A-5%, B-95%; 43min, A-85%, B-15%; 60min, A-85%, B-15%; (2) Preparation of test solution: Take an appropriate amount of cis-tetrahydrocarboline hydrochloride and add water:acetonitrile in a volume ratio of 1:1 to prepare a solution as the test solution; (3) Determination method: inject the test sample solution into the liquid chromatograph, record the chromatogram, and process the chromatogram by the area normalization method; The synthetic route of cis-tetrahydrocarboline hydrochloride is: The impurities produced in the synthetic route are: Impurity 1: D-tryptophan ; Impurity 2: piperonal ; Impurity 3: D-tryptophan methyl ester hydrochloride ; Impurity 4: Hydrolysis impurities ; Impurity 5: Isomer impurity .

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