RU2772608C1 - Method for quantitative determination of papaverine hydrochloride and its related impurities in medicines - Google Patents

Abstract

FIELD: pharmaceutical industry.

SUBSTANCE: group of inventions relates to the field of pharmaceutical chemistry. Methods for the quantitative determination of papaverine hydrochloride and its related impurities in medicines are disclosed, including the preparation of solutions of a standard sample of papaverine hydrochloride at concentrations of 0.1 mg/ml and 0.004 mg/ml, the preparation of 2 mg/ml and 0.1 mg/ml solutions of test samples, while a mixture of acetonitrile and phosphate buffer solution pH 3.0 is used as a solvent in a ratio of 20:80, carrying out chromatography by HPLC 0.1 mg/ml and 0.004 mg/ml solutions of a standard sample of papaverine hydrochloride, 2 mg/ml of the test solution for the quantitative determination of related impurities and 0.1 mg/ml of the test sample solution for the quantitative determination of papaverine hydrochloride in the drug under gradient elution conditions, the quantitative content of papaverine hydrochloride and its related impurities is calculated by the external standard method.

EFFECT: group of inventions provides an increase in the accuracy of simultaneous quantitative determination of the basic substance and related impurities and a reduction in the time of sample preparation and testing.

3 cl, 5 dwg, 7 tbl, 3 ex

Description

The invention relates to the field of pharmaceutical chemistry, in particular to methods for determining the main substance of papaverine hydrochloride and related impurities of papaverine hydrochloride in various dosage forms, namely, it can be used to control the quality of the following drugs: "Papaverine, injection solution 20 mg/ml", "Papaverine, rectal suppositories, 20 mg", "Papaverine, tablets 40 mg", according to the indicators "Quantitative determination" and "Related impurities".

Papaverine hydrochloride (Papaverini hydrochloridum) is a drug belonging to the group of myotropic antispasmodics. It is used in medical practice in the form of a solution for injections, rectal suppositories and tablets for spasms of smooth muscles of the abdominal organs, bronchi, peripheral vessels, cerebral vessels, kidneys; angina pectoris as part of combination therapy, also as an auxiliary drug for premedication. The pharmacological action is based on the ability of papaverine hydrochloride to inhibit the phosphodiesterase enzyme, cause the accumulation of cyclic 3',5'-adenosine monophosphate in the cell and reduce the content of intracellular calcium. As a result, the tone of the smooth muscles of the internal organs and blood vessels decreases in spastic conditions, which leads to the expansion of the arteries and contributes to an increase in blood flow (Handbook of drugs Vidal. https://www.vidal.ru).

For the first time, papaverine was isolated from the milky juice of the immature fruits of the poppy (Papaver somniferum) - opium, where its content reaches 1%. Currently, the substance of papaverine is obtained synthetically. In medical practice, papaverine is used in the form of hydrochloride (Solodukhina LA, Abrosimova LA. Method for obtaining papaverine hydrochloride. Patent of the Russian Federation No. 2647583 C2; 2018. Marinchenko DM. Alkaloids of the isoquinoline group and their use in medicine and pharmacy. In the book: Sikorsky AV, Doronina OK, ed. Actual problems of modern medicine and pharmacy, collection of abstracts of the LXXI International Scientific and Practical Conference of Students and Young Scientists, Minsk: BSMU, 2017, p. 202). According to the chemical structure, papaverine hydrochloride belongs to alkaloids derived from benzylisoquinoline (1-[(3,4-dimethoxyphenyl)methyl]-6,7-dimethoxyisoquinoline hydrochloride).

Related impurities of papaverine include noscapine, thebaine, which are associated alkaloids in the production of papaverine from opium; as well as papaverinol, dihydropapaverine, papaveraldine, tetrahydropapaverine, related to the intermediate products of the synthesis of papaverine hydrochloride (Hifnawy MS, Muhtadi FJ. Analytical profile of papaverine hydrochloride. Analytical Profiles of Drug Substances. 1988:17;367-447. https://doi. org/10.1016/S0099-5428(08)60223-3).

The review of methods for the quantitative determination of alkaloid-containing drugs shows that chromatographic, optical and electrochemical methods, in particular, potentiometry, are currently most widely used (Lizunova GM, Yambulatova EV. Review of methods for the quantitative determination of alkaloid-containing drugs. Young scientist. 2013; (5 ):187-92). Quantitative determination by high performance liquid chromatography (HPLC) in dosage forms containing papaverine hydrochloride is not provided for in the regulatory documentation included in the State Register of Medicines (State Register of Medicines https://grls.rosminzdrav.ru).

An analogue of the claimed invention is a method for assessing related impurities by thin layer chromatography (TLC) and a spectrophotometric method for the quantitative determination of papaverine, described in the Pharmacopoeia article "Papaverine hydrochloride solution 2% for injection" (FS 42-1704-97). The essence of the well-known method based on the TLC technique is the extraction of analytes with chloroform, application to a chromatographic plate, elution in a chamber with a mixture of solvents toluene-ethyl acetate-diethylamine (70:20:10) and viewing in UV light at a wavelength of 254 nm.

The disadvantages of this method are the semi-quantitative assessment of related impurities (determination of compounds directly on the plate by visual comparison of the size of the spots and their color with the corresponding parameters of the spots of standard samples), the influence of a large number of factors on the quality of separation of the mixture components: preliminary saturation of the chamber and the sorbent layer with mobile phase vapor ; starting spot size; distance from the start to the bottom edge of the plate; relative humidity of the air in the laboratory room; average particle diameter and shape; thickness and uniformity of application of the sorbent layer; the presence of microdamages of the layer; type of substance that binds the sorbent; elution rate; solvent volume in the chamber; the presence of impurities in the eluent; convection in the gas phase inside the chamber.

The spectrophotometric method for the quantitative determination of papaverine consists in measuring the optical density of a standard solution of papaverine and the test solution of the drug in a 0.1 M solution of hydrochloric acid on a spectrophotometer at an absorption maximum at a wavelength of 309 nm (Arzamastsev A.P. Pharmaceutical chemistry. - Geotar-med, 2004 ).

The disadvantage of this approach is the insufficient selectivity of the method. Since the spectrophotometric method is based on the absorption of electromagnetic radiation from compounds containing chromophore and auxochromic groups, the spectra of related compounds may overlap, and the results of quantitative determination may give overestimated results.

The prototype of the claimed invention is a method for determining related impurities in the substance of papaverine hydrochloride (FS.2.1.0153.18. Papaverine hydrochloride. Papaverine. State Pharmacopoeia of the Russian Federation. XIV ed. T. 3. M .; 2018), in which the substance of papaverine hydrochloride is dissolved in a mixture solvents of acetonitrile and a phosphate buffer solution pH 3.0 in a ratio of 20:80, then the impurities are determined by high performance liquid chromatography (HPLC), chromatography is carried out on a reverse-phase column filled with an octasilyl silica gel sorbent deactivated with respect to bases (C8), column size 250 × 4.6 mm, particle size 5 μm, under conditions of gradient elution with detection at 238 nm, while a solution of papaverine hydrochloride substance at a concentration of 0.1% relative to the test solution is used as a reference solution.

The disadvantage of the prototype is the inability to use it for the quantitative determination of the main substance in dosage forms: injection, tablets and rectal suppositories.

An urgent task is to develop a sensitive and selective method for the simultaneous quantitative determination of related impurities and the main substance in various dosage forms: injection solution, tablets and rectal suppositories containing papaverine hydrochloride.

The method proposed by the authors makes it possible to increase the accuracy of the quantitative determination of the content of the main substance of papaverine hydrochloride and related impurities of papaverine hydrochloride in drugs in the form of a solution for injections, tablets and rectal suppositories, as well as to simultaneously quantify the main substance of papaverine hydrochloride and related impurities and reduce the time of sample preparation of a standard and test specimens and test.

The problem is solved by variants of the method for the quantitative determination of papaverine hydrochloride and its related impurities in medicines, which includes preparing a solution of a standard sample of papaverine hydrochloride at concentrations of 0.1 mg/ml and 0.004 mg/ml, preparing test solutions at concentrations of 2 mg/ml and 0.1 mg / ml, while a mixture of acetonitrile and phosphate buffer solution pH 3.0 in a ratio of 20:80 is used as a solvent, then HPLC chromatography is carried out with 0.1 mg / ml of papaverine hydrochloride standard sample solution, 0.004 mg / ml of standard sample solution papaverine hydrochloride, 2 mg/ml test solution for the quantitative determination of related impurities and 0.1 mg/ml test solution for the quantitative determination of papaverine hydrochloride in medicinal products in the form of a solution for injection, tablets and rectal suppositories under gradient elution conditions, calculate the quantitative content of papaverine rin hydrochloride and its related impurities by the method of an external standard, while the quantitative content of papaverine hydrochloride (X) in injection solution (1), tablets (2), rectal suppositories (3) is calculated by formulas (1) - (3), respectively:

, мг/мл, (1)X =

, mg/ml, (1)

, мг/табл., (2)X =

, mg/tab., (2)

, мг/супп., (3),X =

, mg/supp., (3),

where

S ₁ - papaverine peak area on the chromatogram of the test solution;

S ₀ - papaverine peak area on the chromatogram of the standard solution;

a ₀ - weight of a standard sample of papaverine hydrochloride, mg;

a ₁ - sample of the drug, mg;

V is the volume of the drug, ml;

P is the content of papaverine hydrochloride in a standard sample of papaverine hydrochloride, %;

G is the average weight of one tablet (one suppository), mg.

The quantitative content of related impurities in injection solution (1), tablets (2), rectal suppositories (3) in percent (X) is calculated by formulas (1-3), respectively:

(1);

(one);

(2);

(2);

(3);

(3);

where _Si is the area of the peak of each of the impurities in the chromatogram of the test solution;

S ₀ - papaverine peak area on the chromatogram of a standard sample solution of papaverine hydrochloride (B);

- навеска стандартного образца папаверина гидрохлорида, мг;

- weight of a standard sample of papaverine hydrochloride, mg;

V ₁ - the volume of the drug, ml;

a ₁ - sample of the drug, mg;

G - average weight of one tablet, mg

L is the declared amount of papaverine hydrochloride in the preparation, mg / ml (mg; mg).

The figure 1 shows the chromatogram of the solution to test the suitability of the chromatographic system (papaverine hydrochloride, noscapine); figure 2 - chromatogram of a 0.1 mg/ml solution of a standard sample of papaverine hydrochloride; figure 3 - chromatogram of the test solution ("Papaverine, solution for injection 20 mg/ml"); figure 4 - chromatogram of the test solution ("Papaverine hydrochloride, tablets, 40 mg"); figure 5 - chromatogram of the test solution (rectal suppositories).

The chromatogram of the chromatographic system suitability test solution shows a satisfactory separating power of the chromatographic system, the separation factor is 4.5.

The method proposed by the authors for the quantitative determination of papaverine hydrochloride and its related impurities in medicines is implemented as follows.

For all dosage forms: solution for injection, tablets and suppositories, test solutions are prepared at a concentration of 2 mg / ml for the determination of related impurities and at a concentration of 0.1 mg / ml for the quantitative determination of papaverine.

Example 1. Quantitative determination of papaverine hydrochloride and its related impurities in the drug "Papaverine, injection solution 20 mg/ml" by the claimed method.

The sample preparation process includes the preparation of a phosphate buffer solution with a pH of 3.0, a solvent, papaverine hydrochloride standard sample solutions and test solutions, solutions for testing the suitability and sensitivity of the chromatographic system.

To prepare a phosphate buffer solution with a pH of 3.0, dissolve 3.40 g of potassium dihydrogen phosphate in 900 ml of water, adjust the pH of the solution potentiometrically to 3.0 using concentrated phosphoric acid, and bring the volume of the solution to 1000.0 ml with water. As a solvent for the preparation of solutions, a mixture of acetonitrile and phosphate buffer solution pH 3.0 is used in a ratio of 20:80.

For the quantitative determination of papaverine hydrochloride in the medicinal product "Papaverine, injection 20 mg/ml", prepare a solution of a standard sample of papaverine hydrochloride at a concentration of 0.1 mg/ml according to the following method: 50.51 mg of a standard sample of papaverine hydrochloride is placed in a volumetric flask with a capacity of 25 ml, dissolve in a solvent and bring the volume of the solution with a solvent to the mark. 5.0 ml of the resulting solution is placed in a volumetric flask with a capacity of 100 ml and the volume is adjusted to the mark with the solvent (solution A).

For the quantitative determination of related impurities in the medicinal product "Papaverine, solution for injection 20 mg / ml", a solution of a standard sample of papaverine hydrochloride is prepared at a concentration of 0.004 mg / ml, which, in relation to the concentration of the main substance in the test solution, corresponds to the maximum expected concentration of any unidentifiable impurity, normalized at the level of 0.2%. To prepare the above solution, 1.0 ml of solution A is placed in a volumetric flask with a capacity of 25 ml and the volume is adjusted to the mark with the solvent (solution B).

To determine related impurities, a solution of the test sample is prepared in duplicate: the volume of the drug, corresponding to about 20 mg of papaverine hydrochloride, is placed in a 10 ml volumetric flask and adjusted to the mark with a solvent.

For the quantitative determination of papaverine hydrochloride, a test sample solution is prepared in triplicate: 1.0 ml of the test solution prepared for the determination of related impurities is placed in a 20 ml volumetric flask and the volume of the solution is adjusted to the mark with a solvent.

A solution for testing the suitability of a chromatographic system is prepared by dissolving 12.015 mg of noscapine (impurity A, CAS 128-62-1) and 12.250 mg of papaverine hydrochloride standard in solvent in a 100 ml volumetric flask and bringing the volume of the solution to the mark with solvent.

To establish the ignore threshold, a solution is prepared to test the sensitivity of the chromatographic system at a concentration of 0.001 mg/ml, which corresponds to 0.05% of the content of the main substance in the test solution. To prepare the above solution, 5.0 ml of solution B of the standard sample of papaverine hydrochloride is placed in a volumetric flask with a capacity of 20 ml and the volume of the solution is adjusted to the mark with a solvent.

The suitability of the chromatographic system for obtaining reliable chromatographic data is checked as follows:

1) on the chromatogram of the solution to test the sensitivity of the chromatographic system, the signal-to-noise ratio (S/N) for the papaverine peak should be at least 10;

2) on the chromatogram of the solution for testing the suitability of the chromatographic system, the resolution (R) between the peaks of papaverine and noscapine must be at least 1.5;

3) on the chromatogram of a standard sample solution of papaverine hydrochloride (solution B), the peak asymmetry factor ( _AS ) of papaverine should be no more than 2.0, and the relative standard deviation of the papaverine peak area should be no more than 5.0% (5 determinations);

4) on the chromatogram of a standard sample solution of papaverine hydrochloride (solution A), the peak asymmetry factor (AS) of _papaverine should be no more than 2.0 and the relative standard deviation of the papaverine peak area should be no more than 5.0% (5 determinations).

The test is carried out under gradient elution conditions. Chromatography conditions are shown in Table 1.

Table 1

Column Size 250 × 4.6 mm, octadecylsilyl silica gel sorbent for chromatography (C18), 5 µm mobile phase A - phosphate buffer solution pH 3.0
B - acetonitrile
B - methanol Elution method gradient program Time, min Ah, about. % B, about. % In about. % 0 - 5 85 5 ten 5 - 12 85 → 60 5 10 → 35 12 - 20 60 5 35 20 - 24 60 → 40 5 → 20 35 → 40 24 - 27 40 twenty 40 27 - 32 40 → 85 20 → 5 40 → 10 32 - 40 85 5 ten Column temperature 35 °С Flow rate 1.0 ml/min Detector spectrophotometric or diode array Wavelength 238 nm Injected sample volume 5 µl

Relative retention time of compounds: papaverine
hydrochloride - 1 (about 20 min); noscapine - about 1.07.

Chromatographic solution for testing the sensitivity of the chromatographic system, solution for testing the suitability of the chromatographic system, solution of a standard sample of papaverine hydrochloride at a concentration of 0.004 mg / ml (solution B), solution of a standard sample of papaverine hydrochloride at a concentration of 0.1 mg / ml (solution A) , test solutions: 2 mg/ml test solution for quantitative determination of related impurities and 0.1 mg/ml test solution for quantitative determination of papaverine hydrochloride. Get 5 chromatograms of a standard sample of papaverine hydrochloride solution A and solution B to assess the reproducibility (relative standard deviation, RSD) values of peak areas. The results are considered reliable, since the requirements of the test "Chromatographic system suitability test" are met. Chromatograms of the solution for checking the suitability of the chromatographic system, the solution of the standard sample of papaverine hydrochloride (solution A), the test solution are shown in Fig. 1-5. The retention time of the main peak of papaverine hydrochloride is 19 minutes.

Do not take into account peaks whose area is less than the area of the main peak in the chromatogram of the solution for checking the sensitivity of the chromatographic system (less than 0.05%), as well as peaks with retention times less than 4 minutes.

To calculate the content of analytes, the external standard method is used.

The content of each of the impurities in the solution for injection in percent (X) is calculated by the formula:

where S _i is the peak area of each of the impurities in the chromatogram of the test solution;

S ₀ - papaverine peak area on the chromatogram of a standard sample solution of papaverine hydrochloride (B);

a ₀ is a weight of a standard sample of papaverine hydrochloride, mg;

V ₁ - the volume of the drug, ml;

L is the declared amount of papaverine hydrochloride in the preparation, mg/ml.

The chromatogram of the test solution revealed two non-ideitified impurities with a retention time of 16.47 (1) and 17.14 (2) min (Fig. 3).

The content of an unidentified impurity (1) is calculated by the formula:

The average value of the content of an unidentified impurity (1) is X=0.011%

The content of an unidentified impurity (2) is calculated by the formula:

= 0,038%The average value of the content of an unidentified impurity (2) is

= 0.038%

The average peak areas were used in the calculations.

The sum of impurities is calculated by arithmetic addition and is 0.049%.

The results of the quantitative determination of related impurities in the test solution of the drug "Papaverine hydrochloride, solution for injection" are shown in Table 2.

table 2

Determined value Standard sample of papaverine hydrochloride Test solution, content
unidentified impurity impurity 1 admixture 2 Peak area 105.3
104.8
104.6
105.1
104.8
Average value ( S ₀ ):
104.9 5.79
5.79
Mean:
5.79 19.55
19.59
Mean:
19.57 5.84
5.83
Mean:
5.84 19.60
19.65
Mean:
19.63 Content of unidentified impurity, % 0.011
0.011 0.038
0.038 Average content of unidentified impurity, %

0,011

0.011

0.038

Thus, in the test solution of the drug "Papaverine hydrochloride, solution for injection" two unidentified related impurities of papaverine hydrochloride were found and calculated according to the formula proposed by the authors - 0.011% and 0.038%, the total impurities amounted to 0.049%.

The permissible content of related impurities is:

- any unidentified impurity - no more than 0.2%;

- the amount of impurities - no more than 1.0%.

Quantitative determination of the content of papaverine hydrochloride (X) in an injection solution is calculated by the formula:

, мг/мл,X =

, mg/ml,

where

S ₁ - papaverine peak area on the chromatogram of the test solution;

S ₀ - papaverine peak area on the standard sample chromatogram;

a ₀ - weight of a standard sample of papaverine hydrochloride, mg;

V is the volume of the drug, ml;

P is the content of papaverine hydrochloride in a standard sample of papaverine hydrochloride, %;

where X ₁ , X ₂ , X ₃ - the content of papaverine hydrochloride in the test solution of the drug; the average peak areas were used in the calculations.

The results of the quantitative determination of papaverine hydrochloride in the test solution of the drug "Papaverine hydrochloride, solution for injection" are shown in Table 3.

Table 3

Determined value Standard sample of papaverine hydrochloride Test solution, content of papaverine hydrochloride Peak area 2527 2496
2493 2537 2531 2488
2489 2527 2529 2478
2481 Mean: 2530.2 The content of papaverine hydrochloride, mg / ml 19.899
19.851
19.779 Average content of papaverine hydrochloride, mg/ml 19.83 Relative standard deviation (RSD), % 0.30

The content of papaverine hydrochloride in the test solution of the drug "Papaverine hydrochloride, solution for injection" was 19.83 mg/ml (RSD = 0.30%).

Thus, example 1 confirms the possibility of using the claimed method for the quantitative determination of papaverine hydrochloride and its related impurities in the drug "Papaverine, injection solution 20 mg/ml".

Example 2. Quantitative determination of papaverine hydrochloride and its related impurities in the drug "Papaverine hydrochloride, tablets" by the claimed method.

The preparation of a phosphate buffer solution with a pH of 3.0, a solvent, solutions of a standard sample of papaverine hydrochloride in two concentrations: 0.1 mg / ml and 0.004 mg / ml, solutions for testing the suitability and sensitivity of the chromatographic system is carried out in the same way as described in example 1.

To determine related impurities, prepare a test solution with a concentration of 2 mg / ml: 330.890 mg of an accurate weight of powder of crushed tablets, corresponding to about 40 mg of papaverine hydrochloride, is placed in a 20 ml volumetric flask, 10 ml of solvent is added, sonicated for 10 minutes, cooled to room temperature, bring the volume of the solution to the mark with the same solvent and filter through a membrane filter with a pore size of not more than 0.45 μm.

For the quantitative determination of papaverine hydrochloride in the drug "Papaverine hydrochloride, tablets", a test solution is prepared with a concentration of 0.1 mg / ml: 1.0 ml of the test solution prepared for the determination of related impurities is placed in a 20 ml volumetric flask and the volume of the solution is adjusted solvent to the mark.

Next, the suitability of the chromatographic system is checked in the same way as in example 1.

Chromatograph under the conditions given in table 1, the test solution for the quantitative determination of papaverine hydrochloride (at a concentration of 0.1 mg/ml) and related impurities (at a concentration of 2 mg/ml). The chromatogram of the test sample solution is shown in Fig. 4. The retention time of the main peak of papaverine hydrochloride is 19 minutes.

Do not take into account peaks whose area is less than the area of the main peak in the chromatogram of the solution to check the sensitivity of the chromatographic system (less than 0.05%).

The content of each of the impurities in tablets as a percentage (X) is calculated by the formula:

where S _i is the peak area of each of the impurities in the chromatogram of the test solution;

S ₀ - papaverine peak area on the chromatogram of a papaverine standard sample solution (B);

a ₀ is a weight of a standard sample of papaverine hydrochloride, mg;

a _i - weighed powder of crushed tablets, mg;

L is the declared amount of papaverine hydrochloride in one tablet, mg;

G is the average weight of one tablet, mg.

The chromatogram of the test solution revealed two unidentified impurities with a retention time of 17.09 (1) and 21.66 (2) min. The chromatogram of the test sample solution is shown in Fig. four.

The average value of the content of an unidentified impurity (1) is X=0.018%

The content of an unidentified impurity (2) is calculated by the formula:

The content of unidentified impurity (1) is calculated by the formula:

= 0,028%The average value of the content of an unidentified impurity (2) is

= 0.028%

The average values of the peak areas were used in the calculations.

The sum of impurities is calculated by arithmetic addition and is 0.046%.

The results of the quantitative determination of related impurities in the test solution of the drug "Papaverine hydrochloride, tablets" are shown in Table 4.

Table 4

Determined value Standard sample of papaverine hydrochloride Test solution, content
unidentified impurity admixture 1 admixture 2 Peak area 105.3
104.8
104.6
105.1
104.8
Average value ( S ₀ ):
104.9 8.72
8.70
Mean:
8.71 13.53
13.51
Mean:
13.52 8.87
8.90
Mean:
8.88 13.81
13.78
Mean:
13.80 Content of unidentified impurity, % 0.018
0.018 0.028
0.028 Average content of unidentified impurity, %

0,018

0.018

0.028

In the sample of the drug "Papaverine hydrochloride, tablets" two unidentified related impurities of papaverine hydrochloride were found in the amount of 0.018% and 0.028%, the amount of impurities was 0.046%.

The rate of content of related impurities is given in example 1.

Quantitative determination of the content of papaverine hydrochloride (X) in tablets is calculated by the formula:

, мг/табл.,X =

, mg/tab.,

where

S ₁ - papaverine peak area on the chromatogram of the test solution;

S ₀ - papaverine peak area on the standard sample chromatogram;

a ₀ - weight of a standard sample of papaverine hydrochloride, mg;

a ₁ - sample of the drug, mg;

P is the content of papaverine hydrochloride in a standard sample of papaverine hydrochloride, %;

G is the average weight of one tablet, mg.

where X ₁ , X ₂ , X ₃ - the content of papaverine hydrochloride in the test solution of the drug; the average peak areas were used in the calculations.

The results of the quantitative determination of papaverine hydrochloride in the test solution of the drug "Papaverine hydrochloride, tablets" are shown in Table 5.

Table 5

Determined value Standard sample of papaverine hydrochloride Test sample, content of papaverine hydrochloride Peak area 2527 2274
2277 2519 2517 2468
2464 2527 2529 2364
2365 Mean: 2523.8 The content of papaverine hydrochloride, mg / ml 38.49
39.30
38.69 Average content of papaverine hydrochloride, mg/ml 38.83 Relative standard deviation (RSD), % 1.09

The content of papaverine hydrochloride in the test solution of the drug "Papaverine hydrochloride, tablets" was 38.83 mg/ml (RSD = 1.09%).

Thus, example 2 confirms the possibility of using the claimed method for the quantitative determination of papaverine hydrochloride and its related impurities in the drug "Papaverine, 40 mg tablets".

Example 3. Quantitative determination of papaverine hydrochloride and its related impurities in the drug "Papaverine hydrochloride, rectal suppositories" by the claimed method

The preparation of a phosphate buffer solution with a pH of 3.0, a solvent, solutions of a standard sample of papaverine hydrochloride in two concentrations: 0.1 mg / ml and 0.004 mg / ml, solutions for testing the suitability and sensitivity of the chromatographic system is carried out in the same way as described in example 1.

To determine related impurities, prepare a test solution at a concentration of 2 mg / ml: 3125.00 mg of the mass of crushed suppositories, corresponding to 50 mg of papaverine hydrochloride, are placed in a 50 ml conical flask, 20 ml of phosphate buffer solution pH 3.0 is added, heated in water bath until the base is melted and shaken thoroughly for 3 minutes while heating. The contents of the flask are cooled to room temperature, filtered into a volumetric flask with a capacity of 25 ml and the volume of the solution is adjusted to the mark with phosphate buffer solution pH 3.0.

For the quantitative determination of papaverine hydrochloride in the drug "Papaverine hydrochloride, rectal suppositories" prepare a test solution of 0.1 mg / ml: 1.0 ml of the test solution prepared to determine related impurities is placed in a volumetric flask with a capacity of 20 ml and the volume of the solution is adjusted with a solvent to the mark.

Next, the suitability of the chromatographic system is checked in the same way as the test given in example 1.

Chromatograph the test solution to determine related impurities and the test solution to quantify papaverine hydrochloride. The chromatogram of the test solution is presented in Fig.5. The retention time of the main peak of papaverine hydrochloride is 19 minutes.

Do not take into account peaks whose area is less than the area of the main peak in the chromatogram of the solution for checking the sensitivity of the chromatographic system (less than 0.05%), as well as peaks with retention times less than 4 min.

The content of each of the impurities in the medicinal product in percent (X) is calculated by the formula:

where _Si is the area of the peak of each of the impurities in the chromatogram of the test solution;

S ₀ - papaverine peak area on the chromatogram of a papaverine standard sample solution (B);

- навеска препарата, мг;

- weight of the drug, mg;

- навеска стандартного образца папаверина гидрохлорида, мг;

- weight of a standard sample of papaverine hydrochloride, mg;

G is the average weight of one suppository, mg;

L is the declared amount of papaverine hydrochloride in one suppository, mg;

The chromatogram of the test solution revealed two unidentified impurities with a retention time of 16.70 (1) and 17.41 (2) min.

The content of unidentified impurity (1) is calculated by the formula:

= 0,007%The average value of the content of an unidentified impurity (1) is

= 0.007%

The content of an unidentified impurity (2) is calculated by the formula:

= 0,007%The average value of the content of an unidentified impurity (2) is

= 0.007%

The average values of the peak areas were used in the calculations.

The amount of impurities is calculated by arithmetic addition and is 0.014%.

The rate of content of related impurities is given in example 1, paragraph "Calculation".

The results of the quantitative determination of related impurities in the test solution of the drug "Papaverine hydrochloride, rectal suppositories, 20 mg" are shown in Table 6.

Table 6

Determined value Standard sample of papaverine hydrochloride Test solution, content
unidentified impurity admixture 1 admixture 2 Peak area 105.3
104.8
104.6
105.1
104.8
Average value ( S ₀ ):
104.9 3.62
3.72
Mean:
3.67 3.39
3.42
Mean:
3.41 3.89
3.78
Mean:
3.83 3.44
3.41
Mean:
3.43 Content of unidentified impurity, % 0.018
0.018 0.028
0.028 Average content of unidentified impurity, %

0,018

0.018

0.028

In the test solution of the drug "Papaverine hydrochloride, rectal suppositories, 20 mg", two unidentified related impurities of papaverine hydrochloride were found in the amount of 0.007% and 0.007%, the amount of impurities was 0.014%.

The allowable content of related impurities is given in example 1.

Quantitative determination of the content of papaverine hydrochloride (X) in rectal suppositories is calculated by the formula:

, мг/супп.,X =

, mg / supp.,

where

S ₁ - papaverine peak area on the chromatogram of the test solution;

S ₀ - papaverine peak area on the chromatogram of the standard solution;

a ₀ - weight of a standard sample of papaverine hydrochloride, mg;

a ₁ - sample of the drug, mg;

P is the content of papaverine hydrochloride in a standard sample of papaverine hydrochloride, %;

G is the average weight of one suppository, mg.

where X ₁ , X ₂ , X ₃ - the content of papaverine hydrochloride in the test sample of the medicinal product; the average peak areas were used in the calculations.

The results of the quantitative determination of papaverine hydrochloride in the test solution of the drug "Papaverine hydrochloride, rectal suppositories, 20 mg" are shown in Table 7.

Table 7

Determined value Standard sample of papaverine hydrochloride Test sample, content of papaverine hydrochloride Peak area 2533 2330
2327 2521 2531 2326
2332 2531 2522 2336
2337 Mean: 2527.6 The content of papaverine hydrochloride, mg / ml 18.59
18.59
18.66 Average content of papaverine hydrochloride, mg/ml 18.61 Relative standard deviation (RSD), % 0.25

The content of papaverine hydrochloride in the sample of the drug "Papaverine hydrochloride, rectal suppositories, 20 mg" was 18.61 mg/ml (RSD = 0.25%).

Thus, example 3 confirms the possibility of using the claimed method for the quantitative determination of papaverine hydrochloride and its related impurities in the drug "Papaverine, rectal suppositories, 20 mg".

The presented examples do not limit the scope of the claims of the present invention and serve for the purpose of illustration.

The method proposed by the authors makes it possible to increase the accuracy of the quantitative determination of the content of related compounds and the main substance of papaverine hydrochloride in drugs in the form of a solution for injections, tablets and rectal suppositories by using the external standard method when calculating the concentration of the analyte according to chromatographic data obtained by HPLC; simultaneously carry out a quantitative determination of the main substance of papaverine hydrochloride and its related impurities by creating a stage of sample preparation of a standard sample of papaverine hydrochloride to determine papaverine hydrochloride at a concentration of 0.1 mg / ml and to determine related impurities of papaverine hydrochloride at a concentration of 0.004 mg / ml, as well as subjects solutions for determining the concentration of papaverine hydrochloride 0.1 mg / ml and for the determination of related impurities of papaverine hydrochloride at a concentration of 2 mg / ml; reduce sample preparation time by using one method to standardize claimed medicinal products in terms of "Related Impurities" and "Quantitative Determination".

Claims (52)

1. A method for the quantitative determination of papaverine hydrochloride and its related impurities in the drug "Papaverine, injection solution 20 mg/ml", including the preparation of solutions of a standard sample of papaverine hydrochloride at concentrations of 0.1 mg/ml and 0.004 mg/ml, preparation of 2 mg /ml and 0.1 mg/ml solutions of the test samples, while a mixture of acetonitrile and phosphate buffer solution pH 3.0 in a ratio of 20:80 is used as a solvent, HPLC chromatography is carried out 0.1 mg/ml and 0.004 mg/ml solutions of the standard sample of papaverine hydrochloride, 2 mg/ml of the test solution for the quantitative determination of related impurities and 0.1 mg/ml of the test sample solution for the quantitative determination of papaverine hydrochloride in the drug "Papaverine, solution for injection 20 mg/ml" under gradient elution conditions , the quantitative content of papaverine hydrochloride and its related impurities is calculated by the external standard method, while the quantitative content of papaverine hydrochloride in mg / ml (X) is calculated by the formula:

where S ₁ - papaverine peak area on the chromatogram of the test solution; S ₀ - papaverine peak area on the chromatogram of the standard solution; a ₀ - weight of a standard sample of papaverine hydrochloride, mg; V is the volume of the preparation, ml; P is the content of papaverine hydrochloride in a standard sample of papaverine hydrochloride, %; the quantitative content of related impurities in percent (X) in the solution for injection is calculated by the formula:

where S _i is the area of the peak of each of the impurities on the chromatogram of the test solution; S ₀ is the papaverine peak area on the chromatogram of a papaverine hydrochloride standard sample solution (B); a ₀ is a weight of a standard sample of papaverine hydrochloride, mg; V ₁ - the volume of the drug, ml; L is the declared amount of papaverine hydrochloride in the preparation, mg/ml. 2. Method for the quantitative determination of papaverine hydrochloride and its related impurities in the drug "Papaverine hydrochloride, tablets, 40 mg", including the preparation of solutions of a standard sample of papaverine hydrochloride at concentrations of 0.1 mg/ml and 0.004 mg/ml, preparation of 2 mg/ml and 0.1 mg / ml solutions of the test samples, while a mixture of acetonitrile and phosphate buffer solution pH 3.0 in a ratio of 20:80 is used as a solvent, HPLC chromatography is carried out 0.1 mg / ml and 0.004 mg / ml solutions of a standard papaverine hydrochloride sample, 2 mg/ml test solution for the quantitative determination of related impurities and 0.1 mg/ml test sample solution for the quantitative determination of papaverine hydrochloride in the medicinal product "Papaverine hydrochloride, tablets" under gradient elution conditions, the quantitative content of papaverine hydrochloride and its related impurities are calculated by the external standard method, while the amount venous content of papaverine hydrochloride in mg / tab. (X) is calculated by the formula:

where S ₁ - papaverine peak area on the chromatogram of the test solution; S ₀ - papaverine peak area on the chromatogram of the standard solution; a ₀ - weight of a standard sample of papaverine hydrochloride, mg; a ₁ - sample of the drug, mg; P is the content of papaverine hydrochloride in a standard sample of papaverine hydrochloride, %; G is the average weight of one tablet, mg; the quantitative content of related impurities in the injection solution is calculated by the formula:

where S _i is the area of the peak of each of the impurities on the chromatogram of the test solution; S ₀ is the papaverine peak area on the chromatogram of a papaverine hydrochloride standard sample solution (B); a ₀ is a weight of a standard sample of papaverine hydrochloride, mg; a ₁ - sample of the drug, mg; V ₁ - the volume of the drug, ml; L is the declared amount of papaverine hydrochloride in the preparation, mg. 3. Method for the quantitative determination of papaverine hydrochloride and its related impurities in the drug "rectal suppositories", including the preparation of solutions of a standard sample of papaverine hydrochloride at concentrations of 0.1 mg/ml and 0.004 mg/ml, preparation of 2 mg/ml and 0.1 mg / ml of solutions of the test samples, while a mixture of acetonitrile and phosphate buffer solution pH 3.0 in a ratio of 20:80 is used as a solvent, HPLC chromatography is carried out 0.1 mg / ml and 0.004 mg / ml solutions of a standard sample of papaverine hydrochloride, 2 mg / ml of the test solution for the quantitative determination of related impurities and 0.1 mg / ml of the test sample solution for the quantitative determination of papaverine hydrochloride in the drug "rectal suppositories" under gradient elution conditions, the quantitative content of papaverine hydrochloride and its related impurities is calculated by the method of an external standard, while the quantitative content of papaver ina hydrochloride in percent (X) is calculated by the formula:

where S ₁ - papaverine peak area on the chromatogram of the test solution; S ₀ - papaverine peak area on the chromatogram of the standard solution; a ₀ - weight of a standard sample of papaverine hydrochloride, mg; a ₁ - sample of the drug, mg; P is the content of papaverine hydrochloride in a standard sample of papaverine hydrochloride, %; G is the average weight of one tablet (one suppository), mg; the quantitative content of related impurities in the injection solution is calculated by the formula:

where S _i is the area of the peak of each of the impurities on the chromatogram of the test solution; S ₀ is the papaverine peak area on the chromatogram of a papaverine hydrochloride standard sample solution (B); a ₀ is a weight of a standard sample of papaverine hydrochloride, mg; a ₁ - sample of the drug, mg; V ₁ - the volume of the drug, ml; L is the declared amount of papaverine hydrochloride in the preparation, mg / ml (mg; mg).

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