RU2338189C1 - Method of detecting quantitative content of methylaminoantipyrine admixture in multi-component medications of anti-fever, analgetic, anti-cold acton - Google Patents

Abstract

FIELD: chemistry.

SUBSTANCE: invention relates to analytical chemistry, namely, to method of detecting 4-methylaminoantipyrine (4-MAAP) admixture in multi-component medications of anti-fever, analgetic, anti-cold action by method of inverse-phase high performance liquid chromatography (HPLC) with ultra-violet (UV) detector. In analysis of tablets "Penalgin N" and "Pentalgin "ФС" solution of tested preparations and control solution, which represents 50 times diluted tested solution, stabilised with sodium sulfite, are successively chromatised on liquid chromatograph in regime of linear gradient of acetonitryl concentration in mobile phase (mixture of acetonitryl with phosphate buffer solution), area of 4-MAAP peak is determined at two different moments of time after beginning of analysis, by formula area of 4-MAAP peak at the moment of sample dissolution is calculated, and basing on this area and on area of analgin peak in control solution, initial content of 4-MAAP is calculated.

EFFECT: creation of efficient method of detecting admixture in multi-component medications of anti-fever, analgetic, anti-cold action.

4 tbl, 5 dwg

Description

The invention relates to high-performance liquid chromatography (HPLC), and in particular to methods for determining impurities in multicomponent drugs with antipyretic, analgesic, anti-cold action, and can be used in the practice of control and analytical laboratories of pharmaceutical enterprises and the pharmacy network. The method belongs to the mass.

The composition of the tablets “Pentalgin N” and “Pentalgin FS” includes analgin (0.300 g), caffeine (0.050 g) and phenobarbital (0.010 g) as active substances. In addition, the preparations contain 0.008 g of codeine and 0.100 g of naproxen (Pentalgin N) and 0.008 g of codeine phosphate and 0.300 g of paracetamol (Pentalgin FS) and excipients. In connection with the content of relatively unstable analgin in tablets, the presence of decomposition products of this substance is also possible in them. To ensure the quality of drugs and their safety for consumers, it is necessary to control the content of these impurities during the production and storage of tablets. In this regard, the problem of developing reliable methods for their quantitative determination is very relevant. This task is also complicated by the instability of dipyrone, which decomposes in the test solution immediately after its preparation.

The closest to the claimed invention according to the task, technical solution and the results achieved is a method for determining the quantitative content of impurities in analginum and its solutions for injection by HPLC (A.A. Tulaganov, A.V. Mikhalev, L.M. Osokina, A. P. Arzamastsev. Quantitative determination of the content of impurities in analginum and its solutions for injection by HPLC. Chem.-Pharm. Journal. 1992. - Issue 4. - P.76 - 77), in which the analysis is carried out by reverse-phase HPLC. phase chromatographic column and ultraviolet detectors torus, using as a mobile phase a mixture of methanol with a phosphate buffer solution in isocratic mode, using an external standard for the quantitative determination of impurities in samples.

The disadvantage of the described method is the low accuracy. So, the authors of the prototype indicated that at an impurity concentration of 4-methylaminoantipyrine (4-MAAP) from 0.1 to 6%, the relative standard deviation is 6%. The reason for this is the decomposition process of dipyrone in the test solution, therefore, the result obtained is the sum of the initial content of 4-MAAP in dipyrone and its amount formed in the solution after dissolution of the analyzed sample.

The objective of the present invention is to develop a method for determining the quantitative content of 4-MAAP tablets "Pentalgin N" and "Pentalgin FS" without taking into account the impurities formed in the test solution.

The problem is achieved using the proposed method, which consists in the fact that the analysis is carried out by HPLC with a reverse phase chromatographic column and an ultraviolet detector using an external standard, characterized in that when analyzing the solution of the test preparations containing analgin, paracetamol, caffeine, phenobarbital and codeine phosphate or analgin, naproxen, caffeine, phenobarbital and codeine and a comparison solution, which is a 50 times diluted test solution, stabilized sulfite ohm sodium, sequentially chromatographed on a liquid chromatograph in the linear gradient gradient mode, the concentration of acetonitrile in the mobile phase during the analysis using a mixture of acetonitrile with phosphate buffer solution as the mobile phase, determine the peak area of 4-MAAP at two different times, then calculate the area using the formula this impurity at the initial time and according to the formula calculate the content of 4-MAAP in the analyzed tablets.

The decomposition of dipyrone begins immediately when a sample of a water-organic solvent is added to the sample. This is confirmed by the dependence on time from the moment of dissolution of the content of 4-MAAP in the tested solutions of tablets "Pentalgin N" (figure 1). Since it takes at least 3 minutes to obtain one chromatogram (sample dissolution, solution filtration, chromatographic process), it is practically impossible to obtain the value of the impurity area at the initial time. However, in this work, we established the possibility of theoretical calculation of this value corresponding to the content of 4-MAAP in the initial (dry) sample. This possibility is also based on the linear nature of the dependence of the concentration of 4-MAAP in the test solution of tablets on time from the moment of dissolution. This linear relationship is confirmed by the data presented in table 1. The linear correlation coefficient calculated using the least squares method is close to 1 (0.99). Using this dependence, the peak area value of 4-MAAP was calculated at the time of dissolution of the sample. The meaning of this calculation explains figure 2. The content of 4-MAAP in the analyzed tablets was calculated relative to the peak area of dipyrone in the comparison solution. The comparison solution was prepared by diluting the test solution 50 times, that is, this solution contains 2% analgin from the initial concentration. To stabilize analgin in the test solution and in the comparison solution, sodium sulfite at a concentration of 5.0 mg / ml was added to them. This reagent only slows down the formation of 4-MAAP, but the peak area of dipyrone during the period of measurements (10-30 minutes after dissolution of the sample) remains almost constant. MAAP-4 concentration was calculated as the product of the peak area ratio MAAP-4 in the test solution and the comparison solution dipyrone in response to _none dipyrone coefficient K with respect to 4-MAAP. The need to introduce _none K due to different sensitivity used in the diode-array detector to 4 MAAP and dipyrone. The latter is explained by the difference in the molar repayment coefficients of these substances. _None K in this study was determined from chromatograms of the solution containing 4-MAAP and analgin in the same concentration.

The method is as follows: the solution of the test drug and the comparison solution are analyzed on a liquid chromatograph with an ultraviolet (UV) detector and a reversed-phase chromatographic column, characterized in that when analyzing the solution of the test preparations containing analgin, paracetamol, caffeine, phenobarbital and codeine phosphate or analgin, naproxen, caffeine, phenobarbital and codeine, and a comparison solution, which is a 50 times diluted test solution stabilized with sodium sulfite, The concentration of acetonitrile in the mobile phase is chromatographed in a linear gradient mode during the analysis using a mixture of acetonitrile with a phosphate buffer solution as the mobile phase, the peak areas of 4-MAAP are determined at two different time points, then the area of this impurity is calculated using the formula at the initial time and the formula calculates the content of 4-MAAP in the analyzed tablets

The method is illustrated by the following examples.

Preparation of a 4-MAAP solution of a certain concentration.

About 0.0600 g of dipyrone (accurately weighed) was placed in a 100 ml volumetric flask, 20 ml of 1M HCl was added, adjusted to the mark with water, stirred and heated in a water bath at 60 ° C for 2 hours. The concentration of the solution was calculated by the equation of the proposed reaction and confirmed by HPLC. According to the stoichiometry of the reaction, 0.060 g of dipyrone corresponds to 0.037 g of 4-MAAP.

Determination of the response coefficient of dipyrone with respect to 4-MAAP.

About 0.0600 g of dipyrone (accurate weighed) and about 0.5 g of sodium sulfite were placed in a 100 ml volumetric flask, 40 ml of water was added and mixed until the substances dissolved. The resulting solution was adjusted to the mark with water and mixed (solution A). About 0.5 g of sodium sulfite was placed in a 100 ml volumetric flask, 40 ml of water was added and stirred until the substance dissolved. 2.0 ml of a 4-MAAP solution of a certain concentration was added, the resulting solution was brought to the mark with water, mixed and filtered through a membrane filter with a pore size of 0.45 μm (solution B). 20 μl of solution B was chromatographed on a liquid chromatograph using a reverse phase chromatography column in a gradient mode. The composition of the PF during the analysis was changed according to the following program:

Time min PF A, vol.% PF V, vol.% 0 one hundred 0 10 0 100 (80) eleven 0 100 (80) 12 one hundred 0 fifteen one hundred 0

As mobile phases (PF), solutions containing acetonitrile, a 0.025 M solution of potassium phosphate monosubstituted and water were used (volume ratio 10:25:65 for PF A and 60:25:15 for PF B). For PF B, the values used in the analysis of the Pentalgin FS tablets are shown in parentheses. Detection was carried out at 244 nm.

Prepared least 3 chromatogram peak area was calculated dipyrone and 4 K calculated MAAP and _none by the formula:

where S _en and S _4MAAP are the peak areas of _dipyrone and 4-MAAP, respectively, in mV · s;

C _4MAAP and C _an are the concentrations of 4-MAAP and analgin, respectively, in mg / ml.

The data for the calculation of K _off are presented in Table 2. According to these data

K _off = 0.660.

Determination of the metrological characteristics of the method for determining 4-MAAP in tablets "Pentalgin FS".

The correctness of the methodology was evaluated by the results of the analysis of model solutions containing all active and all auxiliary substances of the drug, according to the method of "entered - found". Pounded tablets were used as a placebo. To prepare model solutions, 0.16 g of placebo (1/5 tablet), 0.5 g of Na ₂ SO ₃ , 10 ml of H ₂ O were placed in 100 ml volumetric flasks, stirred for 2 min, 30 ml of CH _{3 was} added CN, stirred for 2 min, a certain amount of 4-MAAP solution was added, adjusted to the Н ₂ O mark, mixed and filtered through a hydrophilic membrane filter with a pore size of 0.45 μm. The added amounts of 4-MAAP covered the range from 25 to 150% of the MPC of the impurity in tablets (2% of the content of analgin). A total of 17 solutions were prepared and analyzed. In parallel, a comparison solution was prepared, for which 0.16 g of placebo (1/5 tablet), 0.5 g of Na ₂ SO ₃ , 10 ml of H ₂ O were placed in a 100 ml volumetric flask, stirred for 2 min, 30 was added ml of CH ₃ CN, stirred for 2 min, adjusted to the mark with water and stirred. 2.0 ml of the resulting solution was placed in a 100 ml volumetric flask, to which 0.5 g of Na ₂ SO ₃ , 10 ml of H ₂ O and 5 ml of CH ₃ CN were also added, stirred, adjusted to the mark with water and filtered through a hydrophilic membrane filter with a pore size of 0.45 microns. The resulting solutions were analyzed under the conditions described in the section "Determination of the response coefficient of dipyrone with respect to 4-MAAP". Received 3 chromatograms of each solution and calculated the peak areas of 4-MAAP on the 1st and 3rd chromatograms (time difference 32 min). The peak area of 4-MAAP was calculated at the initial moment of time S ₀ according to the formula:

where S _p is the peak area of 4-MAAP in the last chromatogram;

S ₁ is the peak area of 4-MAAP in the first chromatogram;

Δt is the time difference between the 1st and 3rd chromatograms (32 min);

t is the time since the dissolution of the sample (42 min).

We found MAAP-4 content of the treated samples _etc. With a percentage of the content of dipyrone formula:

where S ₀ - peak area MAAP-4 at the initial time;

S _a - dipyrone peak area in the chromatogram of the reference solution;

With _a - the concentration of dipyrone in the comparison solution as a percentage of the original (2.0 V / V%).

From the obtained values of C _etc. subtracted values found with a similar analysis of placebo solution without additives MAAP-4 solution. The obtained amounts of 4-MAAP were compared with those introduced and the relative error values were calculated. The results of the calculation performed using Excel are shown in Table 3. They show that there is no systematic error in the determination, since the average value of the relative error e _{r cf is} small and less than the confidence interval Δe _{r of the} values of the relative error.

Determination of the content of 4-MAAP in tablets "Pentalgin N" and "Pentalgin FS".

The analysis was carried out similarly to that described in the section "Determination of metrological characteristics", but without adding a solution of 4-MAAP. A portion of the crushed tablets was 0.160 g for Pentalgin FS and 0.145 g for Pentalgin N. The results are presented in tables 1 and 4, the chromatograms of the tested solutions in figure 3 and 4, the chromatograms of the comparison solutions in figure 5.

The results of the determination of 4-MAAP tablets "Pentalgin N"
Table 1 Lab.work: 8-1103 t min 10 26 42 58 74 By _off 0.660 S _ans , mV · s 276876 271105 266680 273029 267758 S _ol , mV · s 104537 117521 132871 147835 163576 C _ol ,% (α = 0.05, n = 3) 0.452 ± 0.009 Lab work 9-1203 S _ans , mV · s 297893 285011 281888 286298 285393 S _ol , mV · s 106239 120039 135575 148466 163574 C _ol ,% (α = 0.05, n = 3) 0.429 ± 0.012 Lab work 10-1203 S _ans , mV · s 274999 270255 269818 273876 272113 S _ol , mV · s 108955 121933 137514 149956 165705 C _ol ,% (α = 0.05, n = 3) 0.478 ± 0.009 22072005 Series t min 10 28 46 64 82 S _ol , mV · s 113838 130561 146802 166009 185732 _Ave C,% (α = 0,05, n = 5) 0.418 ± 0.005 35102005 Series S _ol , mV · s 123081 135576 154853 172974 189951 C _ol ,% (α = 0.05, n = 5) 0.459 ± 0.007 471105 Series S _ol , mV · s 112197 126807 144354 158,498 172105 C _ol ,% (a = 0.05, n = 5) 0.423 ± 0.003

Designations: S _en - peak area of dipyrone in the test solution;

S _ans is the peak area of dipyrone in the comparison solution;

S _ave - peak area MAAP-4;

C _ave - MAAP-4 concentration, calculated by the formula (1).

Data for calculating the response coefficient of dipyrone with respect to 4-MAAP

table 2 with an., mg / ml S _en , mV · s c _4MAAP , mg / ml S _4MAAP , mV · s 0.012 320128 0.0108 433026 318617 432066 317328 434158 S av, mV * s 318691 433083 For an _off / 4MAAP 0.660

Metrological characteristics of the method for determining 4-MAAP in tablets "Pentalgin FS" obtained in the analysis of model solutions

Table 3 S _max, % s ² _max ε _cf ,% ε _max ,% e _{r cf} ,% e _{r max} ,% Δe _r ,% 0,078 0.006 9.2 17.3 - 0,044 -3.64 0.673

Legend:

S _max - the maximum value of the standard deviation;

S ² _max - the maximum value of the variance;

ε _cf - the average value of the relative confidence interval;

ε _max - the maximum value of the relative confidence interval;

_cf. e _r - average relative error;

e _{r max} - the maximum value of the relative error;

Δе _r - confidence interval of values of relative error

The results of the determination of 4-MAAP in tablets "Pentalgin FS" using different solvents. (P = 0.95; n = 3)

Claims (1)

The method for determining the impurity of 4-methylaminoantipyrine (4-MAAP) in multicomponent drugs antipyretic, analgesic, anti-cold action by reverse phase HPLC with an ultraviolet (UV) detector using an external standard, characterized in that when analyzing the solution of the tested drugs containing analgin, paracetamol, caffeine, phenobarbital and codeine phosphate or analgin, naproxen, caffeine, phenobarbital and codeine, and a comparison solution, which is a test solution diluted 50 times a thief stabilized with sodium sulfite is sequentially chromatographed on a liquid chromatograph in the linear gradient gradient concentration of acetonitrile in the mobile phase during the analysis using a mixture of acetonitrile with phosphate buffer solution as the mobile phase, the peak area of 4-MAAP is determined at two different points in time after the start of analysis , calculate according to the formula the peak area of 4-MAAP at the moment of dissolution of the sample and calculate the initial content of 4-MAAP according to the formula:

where C _a is the concentration of dipyrone in the comparison solution as a percentage of the original; S _a - dipyrone peak area in the chromatogram of the reference solution; S ₀ - peak area of 4-MAAP at the initial time, calculated by the formula:

where S _p is the peak area of 4-MAAP in the last chromatogram; S ₁ is the peak area of 4-MAAP in the first chromatogram; Δt is the time difference between the chromatograms; t is the time since the dissolution of the sample.

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