CN111323502A - Detection method for unknown impurities in valsartan preparation - Google Patents
Detection method for unknown impurities in valsartan preparation Download PDFInfo
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- CN111323502A CN111323502A CN201911312121.9A CN201911312121A CN111323502A CN 111323502 A CN111323502 A CN 111323502A CN 201911312121 A CN201911312121 A CN 201911312121A CN 111323502 A CN111323502 A CN 111323502A
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- G—PHYSICS
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
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Abstract
The invention belongs to the technical field of drug impurity detection, and particularly relates to a detection method for an unknown impurity in a valsartan preparation. The invention discloses a method for detecting unknown impurities in a valsartan preparation, which comprises the following steps: s1, crushing valsartan, and screening out powder by using a 200-230-mesh sieve; s2, taking 0.3g of the powder obtained in the step S1, adding 0.6-1.2mL of a solvent containing at least one of N, N-dimethylformamide, isopropanol and water, placing the mixture into a 20mL headspace bottle, sealing the headspace bottle, and detecting by using HS-GC/MS, wherein the unknown impurities are diethylnitrosamine and N-methyl-N-nitrosomethylamine.
Description
Technical Field
The invention belongs to the technical field of drug impurity detection, and particularly relates to a detection method for an unknown impurity in a valsartan preparation.
Background
At present, the detection method of nitrosamine impurities mainly comprises a gas chromatography-mass spectrometry method, a liquid chromatography-mass spectrometry method and a gas chromatography-thermal energy analyzer method, is mainly applied to the industries of food, drinking water and cosmetics, and has few research reports on the detection method of nitrosamine impurities in medicines at home and abroad. In addition, in the conventional detection method, a relatively complex pretreatment is usually required to be performed on the sample, and the sample matrix interferes with the analysis, so that the requirements on repeatability, accuracy, linearity, sensitivity and the like are often difficult to meet at the same time.
Disclosure of Invention
In order to solve the above technical problems, a first aspect of the present invention provides a method for detecting an unknown impurity in a valsartan preparation, comprising the steps of:
s1, crushing valsartan, and screening out powder by using a 200-230-mesh sieve;
s2, taking 0.3g of the powder obtained in the step S1, adding 0.6-1.2mL of a solvent containing at least one of N, N-dimethylformamide, isopropanol and water, placing the mixture into a 20mL headspace bottle, sealing, and detecting by using HS-GC/MS.
Wherein the unknown impurities are diethylnitrosamine and N-methyl-N-nitrosomethylamine.
As a preferable technical solution, the step S2 includes: taking 0.3g of the powder obtained in the step S1, adding 0.6-1.2mL of a mixed solvent comprising N, N-dimethylformamide and isopropanol, placing the mixture into a 20mL headspace bottle, sealing, and detecting by using HS-GC/MS.
As a preferable technical scheme, the volume ratio of the N, N-dimethylformamide to the isopropanol is 1: (2-4).
As a preferred technical solution, the HS test conditions are: headspace heating box temperature: 140 ℃; quantitative ring temperature: 150 ℃; transmission line temperature: 150 ℃; equilibrium temperature: 70-80 ℃; sample introduction time: 1.0 min; equilibrium retention time: 15 min; filling pressure: 15 psi.
As a preferred technical solution, the HS test conditions are: headspace heating box temperature: 140 ℃; quantitative ring temperature: 150 ℃; transmission line temperature: 150 ℃; equilibrium temperature: 75 ℃; sample introduction time: 1.0 min; equilibrium retention time: 15 min; filling pressure: 15 psi.
As a preferred technical scheme, the stationary phase in the gas chromatography is as follows: 5-7% of cyanopropyl phenyl and 93-95% of methyl polysiloxane.
As a preferred technical scheme, the stationary phase in the gas chromatography is as follows: cyanopropyl phenyl 6% and methyl polysiloxane 94%.
As a preferred technical solution, the gas chromatography conditions are:
sample inlet temperature: 240 ℃; flow rate: 0.8-1.2 mL/min;
shunting mode: the split ratio is 5: 1;
temperature programming: the initial temperature is 70 deg.C, maintained for 4min, increased to 100 deg.C at 2-4 deg.C/min, maintained for 1min, increased to 240 deg.C at 8-12 deg.C/min, and maintained for 3.5 min.
As a preferred technical solution, the test conditions of the mass spectrum are as follows: type of ion source: EI; scanning mode: MRM; electron energy: 40-60 eV; filament emission current: 60-80 muA; ion source temperature: 220 ℃; quadrupole temperature: 150 ℃; transmission line temperature: 230 ℃ to 230 ℃.
As a preferred technical solution, in the mass spectrometry detection: and (3) quantifying ions: N-methyl-N-nitrosomethylamine: m/z 74, diethylnitrosamine: and m/z is 102.
Has the advantages that: the detection method disclosed by the invention has high efficiency, accuracy, repeatability and sensitivity for detecting the diethylnitrosamine and the N-methyl-N-nitrosomethylamine in the valsartan.
Detailed Description
For purposes of the following detailed description, it is to be understood that the invention may assume various alternative variations and step sequences, except where expressly specified to the contrary. Moreover, other than in any operating examples, or where otherwise indicated, all numbers expressing, for example, quantities of ingredients used in the specification and claims are to be understood as being modified in all instances by the term "about". Accordingly, unless indicated to the contrary, the numerical parameters set forth in the following specification and attached claims are approximations that may vary depending upon the desired properties to be obtained by the present invention. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be construed in light of the number of reported significant digits and by applying ordinary rounding techniques.
Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the invention are approximations, the numerical values set forth in the specific examples are reported as precisely as possible. Any numerical value, however, inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements.
When a range of values is disclosed herein, the range is considered to be continuous and includes both the minimum and maximum values of the range, as well as each value between such minimum and maximum values. Further, when a range refers to an integer, each integer between the minimum and maximum values of the range is included. Further, when multiple range-describing features or characteristics are provided, the ranges may be combined. In other words, unless otherwise indicated, all ranges disclosed herein are to be understood to encompass any and all subranges subsumed therein. For example, a stated range from "1 to 10" should be considered to include any and all subranges between the minimum value of 1 and the maximum value of 10. Exemplary subranges of the range 1 to 10 include, but are not limited to, 1 to 6.1, 3.5 to 7.8, 5.5 to 10, and the like.
In order to solve the above problems, the present invention provides a method for detecting unknown impurities in a valsartan formulation, comprising the steps of:
s1, crushing valsartan, and screening out powder by using a 200-230-mesh sieve;
s2, taking 0.3g of the powder obtained in the step S1, adding 0.6-1.2mL of a solvent containing at least one of N, N-dimethylformamide, isopropanol and water, placing the mixture into a 20mL headspace bottle, sealing, and detecting by using HS-GC/MS.
Wherein the unknown impurities are diethylnitrosamine and N-methyl-N-nitrosomethylamine.
Step S1
The valsartan, also called as valsartan, ambroxol hydrochloride impurity D, N-pentanoyl-N- [ [2'- (1H-tetrazole-5-yl) [1,1' -biphenyl ] -4-yl ] methyl ] -L-valine is an angiotensin II receptor antagonist antihypertensive drug, and the drug has the effects of blocking an I (AT1) receptor of angiotensin II, increasing the blood plasma level of the angiotensin II, stimulating an unblocked AT2 receptor and simultaneously countering an AT1 receptor, so that the effects of expanding blood vessels and reducing blood pressure are achieved.
The applicant finds that when screening valsartan by using a 200-230-mesh sieve, powder particles have a certain particle size and have a proper diffusion speed in a solvent; a certain amount of powder is added into the headspace bottle, so that the sample injector can be ensured to contain a proper target compound, and the interference caused by other impurities is reduced.
Step S2
As a preferred embodiment, the step S2 includes: taking 0.3g of the powder obtained in the step S1, adding 0.6-1.2mL of a mixed solvent comprising N, N-dimethylformamide and isopropanol, placing the mixture into a 20mL headspace bottle, sealing, and detecting by using HS-GC/MS.
As a more preferable embodiment, the step S2 includes: 0.3g of the powder obtained in the step S1 was taken, 0.9mL of a mixed solvent containing N, N-dimethylformamide and isopropanol was added, the mixture was sealed in a 20mL headspace bottle, and detection was performed by HS-GC/MS.
The volume ratio of the N, N-dimethylformamide to the isopropanol is 1: (2-4).
The term "HS-GC/MS" refers to headspace gas chromatography-mass spectrometry.
As a preferred embodiment, the HS test conditions are: headspace heating box temperature: 140 ℃; quantitative ring temperature: 150 ℃; transmission line temperature: 150 ℃; equilibrium temperature: 70-80 ℃; sample introduction time: 1.0 min; equilibrium retention time: 15 min; filling pressure: 15 psi.
As a more preferred embodiment, the HS test conditions are: headspace heating box temperature: 140 ℃; quantitative ring temperature: 150 ℃; transmission line temperature: 150 ℃; equilibrium temperature: 75 ℃; sample introduction time: 1.0 min; equilibrium retention time: 15 min; filling pressure: 15 psi.
The applicant finds that in addition to the main active ingredient valsartan, the valsartan preparation also contains various auxiliary ingredients such as microcrystalline cellulose, crospovidone, povidone, sodium dodecyl sulfate, magnesium stearate, gelatin, sodium dodecyl sulfate, iron oxide and titanium dioxide, so that in the extraction process, the dissolution and extraction of the target ingredient are interfered through various mechanisms such as physical and chemical mechanisms, and the accuracy of the detection method is influenced.
The pharmacopoeia of 2000 th edition of China already adopts headspace gas chromatography to analyze residual organic solvents in medicines, but a more perfect headspace technology has a plurality of problems, and one of the main problems is that no proper sample solvent selection rule exists at present.
The inventor of the application finds that when 0.3g of the obtained powder is added with 0.6-1.2mL of a mixed solvent of N, N-dimethylformamide and isopropanol, the equilibrium temperature is controlled to be 70-80 ℃, and the equilibrium holding time is 15min, the recovery rate, the accuracy and the like of the detection method can be improved. Probably because the headspace system is in dynamic equilibrium, the solution formed by N, N-dimethylformamide and isopropanol influences the molecular mobility of diethylnitrosamine and N-methyl-N-nitrosomethylamine out of the drug and into the solvent; when the headspace equilibrium temperature is lower than 70 ℃, the molecular moving speed is slow, and the headspace system is difficult to achieve dynamic equilibrium. The volume ratio of the N, N-dimethylformamide to the isopropanol is 1: and (2-4), the dissolution of the component to be detected in the medicine is promoted, and the hydrophilic hydroxyl and the hydrophobic alkyl of the isopropanol make up the deficiency of the polar hydrophobic solvent of the N, N-dimethylformamide, so that the gas-liquid two phases can quickly reach a diffusion balance stable state.
Meanwhile, when the balance temperature is too high and the balance time is too long, some volatile components with unstable properties in the system undergo side reactions such as thermal decomposition, oxidation and the like, so that the pressure of the headspace gas is too high, and the airtightness is reduced; on the other hand, when the equilibration temperature is too low and the equilibration time is too short, it is difficult for the target compound to completely diffuse or volatilize, resulting in too low a concentration thereof, which also reduces the accuracy of the detection method. When the amount of N, N-dimethylformamide is large, strong intermolecular forces firmly adsorb the component to be detected in the solvent, and the gas-liquid two phases are difficult to reach diffusion balance. Under the complex conditions of numerous influencing factors and unpredictable results, the applicant is careful to research the parameters.
As a preferred embodiment, the stationary phase in the gas chromatography is: 5-7% of cyanopropyl phenyl and 93-95% of methyl polysiloxane.
As a more preferred embodiment, the stationary phase in the gas chromatography is: cyanopropyl phenyl 6% and methyl polysiloxane 94%.
The specification of a chromatographic column in the gas chromatography is DB-624(30m × 250 μm × 1.4.4 μm).
Polysiloxanes are currently used as stationary phases, and standard polysiloxanes are formed by linking a plurality of individual siloxanes, each silicon atom being linked to two functional groups, the type and number of which determine the nature of the stationary phase of the chromatographic column. When other substituents are substituted for methyl, the number of said substituents is indicated by a percentage.
As a preferred embodiment, the gas chromatography conditions are:
sample inlet temperature: 240 ℃; flow rate: 0.8-1.2 mL/min;
shunting mode: the split ratio is 5: 1;
temperature programming: starting temperature is 70 deg.C, maintaining for 4min, raising to 100 deg.C at 2-4 deg.C/min, maintaining for 1min, raising to 240 deg.C at 8-12 deg.C/min, and maintaining for 3.5 min;
carrier gas: high purity helium (99.999%).
As a more preferred embodiment, the gas chromatography conditions are:
sample inlet temperature: 240 ℃; flow rate: 1.0 mL/min;
shunting mode: the split ratio is 5: 1;
temperature programming: the initial temperature is 70 deg.C, maintained for 4min, increased to 100 deg.C at 3 deg.C/min, maintained for 1min, increased to 240 deg.C at 10 deg.C/min, and maintained for 3.5 min;
carrier gas: high purity helium (99.999%).
The applicant has found that when a programmed temperature increase is employed: the initial temperature is 70 ℃, the temperature is kept for 4min, the temperature is increased to 100 ℃ at the speed of 2-4 ℃/min, the temperature is kept for 1min, the temperature is increased to 240 ℃ at the speed of 8-12 ℃/min, the temperature is kept for 3.5min, and the sensitivity of the detection method can be improved. The guessing reason is probably because the proper programmed temperature rise is selected, the separation speed and the separation degree of the sample to be detected are favorably improved, the diffusion of impurities in the detector is reduced, the interference of impurity peaks is inhibited, sharp chromatographic peaks are obtained, the phenomenon of overloading of a chromatographic column can be effectively avoided under the condition of proper split ratio, the generation of tailing peaks is inhibited, the separation effect is further improved, and the sensitivity of the detection method is improved.
The applicant has also found that when the stationary phase composition of the chromatographic column comprises cyanopropylbenzene and methylpolysiloxane, in particular cyanopropylphenyl 5-7% and methylpolysiloxane 93-95%, the accuracy, repeatability and limit of quantitation of the detection method can be further improved. The reason for guessing is: the cyanopropyl phenyl grafted polysiloxane has dispersed side chain benzene rings to produce delocalized conjugated large pi bond, and through strict control of temperature raising rate, fixed phase and the component to be measured produce different induced dipole force and dispersion force, so as to produce excellent separation effect, especially the separation between N-nitroso-N-methyl-4-amino butyric acid and diethyl nitrosamine, and thus the detection may be performed accurately.
As a preferred embodiment, the test conditions of the mass spectrum are: type of ion source: EI; scanning mode: MRM; electron energy: 40-60 eV; filament emission current: 60-80 muA; ion source temperature: 220 ℃; quadrupole temperature: 150 ℃; transmission line temperature: 230 ℃;
in the mass spectrometric detection: and (3) quantifying ions: N-methyl-N-nitrosomethylamine: m/z 74, diethylnitrosamine: and m/z is 102.
As a more preferred embodiment, the test conditions of the mass spectrum are: type of ion source: EI; scanning mode: MRM; electron energy: 50 eV; filament emission current: 70 muA; ion source temperature: 220 ℃; quadrupole temperature: 150 ℃; transmission line temperature: 230 ℃; and (3) quantifying ions: N-methyl-N-nitrosomethylamine: m/z 74, diethylnitrosamine: 102 m/z; solvent retardation: and 4 min.
The inventor adopts the electron impact ion source (EI) in this application, utilizes the electron of certain energy direct action to the sample molecule, makes its ionization, and efficient, helps the mass spectrometer obtain high sensitivity and high resolution. Through the mutual synergistic effect of the parameters such as the ion source temperature, the quadrupole rod temperature, the transmission line temperature and the like, on one hand, the signal-to-noise ratio of the equipment is improved, the influence of noise is reduced, and the signal has higher stability; on the other hand, the ionization efficiency is improved, and under the condition of proper mass-to-charge ratio, the interference on quantitative ions is minimized, and qualitative and quantitative analysis can be more accurately carried out.
In particular, the applicant has also found that when the electron energy: 50 eV; filament emission current: 60-80 μ A, the accuracy of the detection method can be further improved, presumably because: electrons generated by the filament enter the ionization chamber through the slits on the two sides of the ionization chamber and collide with the substance to be detected in the ionization chamber, and when the energy of the electrons is 50 eV; when the filament emission current is 60-80 muA, the characteristic fragment ion peak with good signal intensity can be obtained, and the matching of the characteristic peak and the target compound is convenient.
When ionization energy of 70eV is used, because the ionization potential of the organic compound in the component to be detected is larger than that of the organic compound in the component to be detected, a plurality of molecular ions are further cracked to form generalized fragment ions, and the molecular ions and the fragment ions of various substances have an overlapping phenomenon, so that spectrogram analysis is difficult.
By searching NIST 2011 online mass spectrum database, checking mass spectrum matching degree and referring to related literature data, finding out characteristic peaks of diethylnitrosamine and N-methyl-N-nitrosomethylamine, and calculating the respective relative contents of the diethylnitrosamine and the N-methyl-N-nitrosomethylamine by adopting a peak area normalization method.
As a preferred embodiment, the detection method further includes: preparing a series of standard solutions of unknown impurities for quantification of the unknown impurities.
The standard solution can be an internal standard solution or an external standard solution.
In the application, the preparation method of the series of standard solutions of the diethyl nitrosamine and the N-methyl-N-nitrosomethylamine comprises the following steps:
taking diethyl nitrosamine and N-methyl-N-nitrosomethylamine, dissolving and diluting to 100mL by using a solvent (the volume ratio of N, N-dimethylformamide to isopropanol is 1: 3), and shaking up to obtain a mixed standard stock solution with the concentrations of both the diethyl nitrosamine and the N-methyl-N-nitrosomethylamine being 100 mu g/mL.
The mixed standard stock solution was diluted with a solvent (volume ratio of N, N-dimethylformamide to isopropanol of 1: 3) to a standard curve of 0.05, 0.1, 0.2, 0.3, 0.4. mu.g/mL series of concentrations.
The diethyl nitrosamine is NDEA for short, and the N-methyl-N-nitrosomethylamine is NDMA for short.
The present invention will be specifically described below by way of examples. It should be noted that the following examples are only for illustrating the present invention and should not be construed as limiting the scope of the present invention, and that the insubstantial modifications and adaptations of the present invention by those skilled in the art based on the above disclosure are still within the scope of the present invention.
In addition, the starting materials used are all commercially available, unless otherwise specified.
Examples
Example 1
A method for detecting an unknown impurity in a valsartan formulation, comprising the steps of:
s1, crushing valsartan, and screening out powder by using a 200-mesh sieve;
s2, taking 0.3g of the powder obtained in the step S1, adding 0.9mL of solvent, placing the mixture into a 20mL headspace bottle, sealing, and detecting by using HS-GC/MS.
Wherein the unknown impurities are diethylnitrosamine and N-methyl-N-nitrosomethylamine.
In step S2, the solvent is a mixed solvent of N, N-dimethylformamide and isopropanol, and the volume ratio of N, N-dimethylformamide to isopropanol is 1: 3.
the HS test conditions are as follows: headspace heating box temperature: 140 ℃; quantitative ring temperature: 150 ℃; transmission line temperature: 150 ℃; equilibrium temperature: 75 ℃; sample introduction time: 1.0 min; equilibrium retention time: 15 min; filling pressure: 15 psi.
The stationary phase in the gas chromatography comprises cyanopropylphenyl 6% and methyl polysiloxane 94%, and the specification is DB-624(30m × 250 μm × 1.4.4 μm).
The gas chromatography conditions were:
sample inlet temperature: 240 ℃; flow rate: 1.0 mL/min;
shunting mode: the split ratio is 5: 1;
temperature programming: the initial temperature is 70 deg.C, maintained for 4min, increased to 100 deg.C at 3 deg.C/min, maintained for 1min, increased to 240 deg.C at 10 deg.C/min, and maintained for 3.5 min;
carrier gas: high purity helium (99.999%).
The test conditions of the mass spectrum are as follows: type of ion source: EI; scanning mode: MRM; electron energy: 50 eV; filament emission current: 70 muA; ion source temperature: 220 ℃; quadrupole temperature: 150 ℃; transmission line temperature: 230 ℃; and (3) quantifying ions: N-methyl-N-nitrosomethylamine: m/z 74, diethylnitrosamine: 102 m/z; solvent retardation: and 4 min.
Preparing the series of standard solutions of the diethyl nitrosamine and the N-methyl-N-nitrosomethylamine for quantitative analysis, wherein the preparation method of the series of standard solutions of the diethyl nitrosamine and the N-methyl-N-nitrosomethylamine comprises the following steps:
10.0mg of each of diethylnitrosamine and N-methyl-N-nitrosomethylamine is taken, dissolved and diluted to 100mL by a solvent (the volume ratio of N, N-dimethylformamide to isopropanol is 1: 3), and shaken up to be used as a mixed standard stock solution of the diethylnitrosamine and the N-methyl-N-nitrosomethylamine, wherein the concentration is 100 mug/mL.
The mixed standard stock solution was diluted with a solvent (volume ratio of N, N-dimethylformamide to isopropanol of 1: 3) to a standard curve of 0.05, 0.1, 0.2, 0.3, 0.4. mu.g/mL series of concentrations.
Example 2
A method for detecting an unknown impurity in a valsartan formulation, comprising the steps of:
s1, crushing valsartan, and screening out powder by using a 230-mesh sieve;
s2, taking 0.3g of the powder obtained in the step S1, adding 0.6mL of solvent, placing the mixture into a 20mL headspace bottle, sealing, and detecting by using HS-GC/MS.
Wherein the unknown impurities are diethylnitrosamine and N-methyl-N-nitrosomethylamine.
In step S2, the solvent is a mixed solvent of N, N-dimethylformamide and isopropanol, and the volume ratio of N, N-dimethylformamide to isopropanol is 1: 3.
the HS test conditions and gas chromatography conditions were the same as in example 1.
The test conditions of the mass spectrum are as follows: type of ion source: EI; scanning mode: MRM; electron energy: 50 eV; filament emission current: 60 muA; ion source temperature: 220 ℃; quadrupole temperature: 150 ℃; transmission line temperature: 230 ℃; and (3) quantifying ions: N-methyl-N-nitrosomethylamine: m/z 74, diethylnitrosamine: 102 m/z; solvent retardation: and 4 min.
Example 3
A method for detecting an unknown impurity in a valsartan formulation, comprising the steps of:
s1, crushing valsartan, and screening out powder by using a 200-mesh sieve;
s2, taking 0.3g of the powder obtained in the step S1, adding 1.2mL of solvent, placing the mixture into a 20mL headspace bottle, sealing, and detecting by using HS-GC/MS.
Wherein the unknown impurities are diethylnitrosamine and N-methyl-N-nitrosomethylamine.
In step S2, the solvent is a mixed solvent of N, N-dimethylformamide and isopropanol, and the volume ratio of N, N-dimethylformamide to isopropanol is 1: 3.
the HS test conditions and gas chromatography conditions were the same as in example 1.
The test conditions of the mass spectrum are as follows: type of ion source: EI; scanning mode: MRM; electron energy: 50 eV; filament emission current: 80 muA; ion source temperature: 220 ℃; quadrupole temperature: 150 ℃; transmission line temperature: 230 ℃; and (3) quantifying ions: N-methyl-N-nitrosomethylamine: m/z 74, diethylnitrosamine: 102 m/z; solvent retardation: and 4 min.
Example 4
A method for detecting an unknown impurity in a valsartan formulation, comprising the steps of:
s1, crushing valsartan, and screening out powder by using a 200-mesh sieve;
s2, taking 0.3g of the powder obtained in the step S1, adding 0.9mL of solvent, placing the mixture into a 20mL headspace bottle, sealing, and detecting by using HS-GC/MS.
Wherein the unknown impurities are diethylnitrosamine and N-methyl-N-nitrosomethylamine.
In step S2, the solvent is a mixed solvent of N, N-dimethylformamide and isopropanol, and the volume ratio of N, N-dimethylformamide to isopropanol is 1: 6.
the HS test conditions, gas chromatography conditions, mass spectrometry test conditions were the same as in example 1.
Example 5
A method for detecting an unknown impurity in a valsartan formulation, comprising the steps of:
s1, crushing valsartan, and screening out powder by using a 200-mesh sieve;
s2, taking 0.3g of the powder obtained in the step S1, adding 0.9mL of solvent, placing the mixture into a 20mL headspace bottle, sealing, and detecting by using HS-GC/MS.
Wherein the unknown impurities are diethylnitrosamine and N-methyl-N-nitrosomethylamine.
In step S2, the solvent is a mixed solvent of N, N-dimethylformamide and isopropanol, and the volume ratio of N, N-dimethylformamide to isopropanol is 1: 0.5.
the HS test conditions, gas chromatography conditions, mass spectrometry test conditions were the same as in example 1.
Example 6
The specific steps of a method for detecting unknown impurities in a valsartan preparation are the same as those in example 1, and the difference is that the HS test conditions are as follows: headspace heating box temperature: 140 ℃; quantitative ring temperature: 150 ℃; transmission line temperature: 150 ℃; equilibrium temperature: 50 ℃; sample introduction time: 1.0 min; equilibrium retention time: 15 min; filling pressure: 15 psi.
Example 7
The specific steps of a method for detecting unknown impurities in a valsartan preparation are the same as those in example 1, and the difference is that the stationary phase in gas chromatography is as follows: cyanopropylphenyl 14%, methylpolysiloxane 86%, available from dengxisai scientific instruments ltd, cat # 053164.
Example 8
The specific steps of a method for detecting unknown impurities in a valsartan preparation are the same as those in example 1, and the difference is that the stationary phase in gas chromatography is as follows: cyanopropyl 25% -phenyl 25% -methyl polysiloxane 50%, model CP-Sil 43CB, brand Agilent.
Example 9
The specific steps of a method for detecting unknown impurities in a valsartan preparation are the same as those in example 1, and the difference is that the gas chromatography conditions are as follows:
sample inlet temperature: 240 ℃; flow rate: 1.0 mL/min;
shunting mode: the split ratio is 5: 1;
temperature programming: the initial temperature is 70 deg.C, maintained for 4min, increased to 100 deg.C at 8 deg.C/min, maintained for 1min, increased to 240 deg.C at 10 deg.C/min, and maintained for 3.5 min.
Example 10
The specific steps of a method for detecting unknown impurities in a valsartan preparation are the same as those in example 1, and the difference is that the gas chromatography conditions are as follows:
sample inlet temperature: 240 ℃; flow rate: 1.0 mL/min;
shunting mode: the split ratio is 5: 1;
temperature programming: the initial temperature is 70 deg.C, maintained for 4min, increased to 100 deg.C at1 deg.C/min, maintained for 1min, increased to 240 deg.C at 10 deg.C/min, and maintained for 3.5 min.
Example 11
The specific steps of a method for detecting unknown impurities in a valsartan preparation are the same as those in example 1, and the difference is that the mass spectrum testing conditions are as follows: type of ion source: EI; scanning mode: MRM; electron energy: 70 eV; filament emission current: 20 muA; ion source temperature: 220 ℃; quadrupole temperature: 150 ℃; transmission line temperature: 230 ℃; and (3) quantifying ions: N-methyl-N-nitrosomethylamine: m/z 74, diethylnitrosamine: 102 m/z; solvent retardation: and 4 min.
Example 12
The specific steps of a method for detecting unknown impurities in a valsartan preparation are the same as those in example 1, and the difference is that the mass spectrum testing conditions are as follows: type of ion source: EI; scanning mode: MRM; electron energy: 50 eV; filament emission current: 20 muA; ion source temperature: 220 ℃; quadrupole temperature: 150 ℃; transmission line temperature: 230 ℃; and (3) quantifying ions: N-methyl-N-nitrosomethylamine: m/z 74, diethylnitrosamine: 102 m/z; solvent retardation: and 4 min.
Example 13
The specific steps of a method for detecting unknown impurities in a valsartan preparation are the same as those in example 1, and the difference is that the mass spectrum testing conditions are as follows: type of ion source: EI; scanning mode: MRM; electron energy: 50 eV; filament emission current: 100 muA; ion source temperature: 220 ℃; quadrupole temperature: 150 ℃; transmission line temperature: 230 ℃; and (3) quantifying ions: N-methyl-N-nitrosomethylamine: m/z 74, diethylnitrosamine: 102 m/z; solvent retardation: and 4 min.
Example 14
A method for detecting an unknown impurity in a valsartan formulation, comprising the steps of:
s1, crushing valsartan, and screening out powder by using a 200-mesh sieve;
s2, taking 0.3g of the powder obtained in the step S1, adding 0.9mL of solvent, placing the mixture into a 20mL headspace bottle, sealing, and detecting by using HS-GC/MS.
Wherein the unknown impurities are diethylnitrosamine and N-methyl-N-nitrosomethylamine.
In step S2, the solvent is a mixed solvent of N, N-dimethylformamide and ethyl acetate, and the volume ratio of N, N-dimethylformamide to ethyl acetate is 1: 3.
the HS test conditions, gas chromatography conditions, mass spectrometry test conditions were the same as in example 1.
Performance testing
The methodology of the detection method described in the examples was validated;
in the TIC spectrum: the peak position of the diethyl nitrosamine is 12.8, and the peak position of the N-methyl-N-nitrosomethylamine is 9.8.
The relevant solutions of examples 1-3, 6-14 were formulated as follows:
(1) preparing a blank control solution
Measuring 2ml of solvent (the volume ratio of N, N-dimethylformamide to isopropanol is 1: 3) and placing the solvent in the headspace bottle to serve as the blank control solution;
(2) preparing mother solution of sample
Crushing valsartan, and sieving powder with a 200-mesh sieve; to 0.3g of the obtained powder, 0.9mL of a solvent (volume ratio of N, N-dimethylformamide to isopropanol: 1: 3) was added.
(3) Preparing a sample solution
2mL of the mother solution is sampled and put into a 10mL volumetric flask, and the volume is determined by using a solvent (the volume ratio of N, N-dimethylformamide to isopropanol is 1: 3).
(4) Preparing an accurate solution
2mL of the mother solution is sampled and put into a 10mL volumetric flask, 20 mu L of the mixed standard stock solution is added, and the volume is fixed by using a solvent (the volume ratio of N, N-dimethylformamide to isopropanol is 1: 3). A total of 6 accurate solutions were prepared in parallel in the same way. Diethylnitrosamine and N-methyl-N-nitrosomethylamine were added at a standard concentration of 0.2. mu.g/mL.
(5) Preparing quantitative limiting solution
2mL of the mother solution is sampled and put into a 10mL volumetric flask, 10 mu L of the mixed standard stock solution is added, and the volume is fixed by using a solvent (the volume ratio of N, N-dimethylformamide to isopropanol is 1: 3). A total of 6 quantitation limit solutions were prepared in parallel with the same method. Diethylnitrosamine and N-methyl-N-nitrosomethylamine were added at a standard concentration of 0.1. mu.g/mL.
(6) Preparing a repetitive solution
2mL of the mother solution is sampled and put into a 10mL volumetric flask, 20 mu L of the mixed standard stock solution is added, and the volume is fixed by using a solvent (the volume ratio of N, N-dimethylformamide to isopropanol is 1: 3). A total of 6 replicate solutions were prepared in parallel. Diethylnitrosamine and N-methyl-N-nitrosomethylamine were added at a standard concentration of 0.2. mu.g/mL.
(7) Mixing standard stock solutions
Preparing the series of standard solutions of the diethyl nitrosamine and the N-methyl-N-nitrosomethylamine for quantitative analysis, wherein the preparation method of the series of standard solutions of the diethyl nitrosamine and the N-methyl-N-nitrosomethylamine comprises the following steps:
10.0mg of each of diethylnitrosamine and N-methyl-N-nitrosomethylamine is taken, dissolved and diluted to 100mL by a solvent (the volume ratio of N, N-dimethylformamide to isopropanol is 1: 3), and shaken up to be used as a mixed standard stock solution of the diethylnitrosamine and the N-methyl-N-nitrosomethylamine, wherein the concentration is 100 mug/mL.
The solutions of examples 4 and 5 were prepared as above, except that: the solvent in example 4 was N, N-dimethylformamide and isopropanol in a volume ratio of 1: 6, the solvent in example 5 is N, N-dimethylformamide and isopropanol in a volume ratio of 1: 0.5 of a mixed solvent.
The test results are shown in the following table.
Table 1 example 1 standard solution linear dependence
TABLE 2 accuracy, reproducibility, quantitative limiting concentration measurements of the examples
Example 12 shows no obvious qualitative peak of mass spectrum, and the irregular drifting of the base line of examples 11 and 13 influences the qualitative.
The above description is only a preferred embodiment of the present invention, and is not intended to limit the present invention in other forms, and any person skilled in the art may modify or change the technical content of the above disclosure into equivalent embodiments with equivalent changes, but all those simple modifications, equivalent changes and modifications made to the above embodiments according to the technical spirit of the present invention still belong to the protection scope of the present invention.
Claims (10)
1. A method for detecting unknown impurities in a valsartan preparation, which is characterized by comprising the following steps:
s1, crushing valsartan, and screening out powder by using a 200-230-mesh sieve;
s2, taking 0.3g of the powder obtained in the step S1, adding 0.6-1.2mL of a solvent containing at least one of N, N-dimethylformamide, isopropanol and water, sealing the mixture in a 20mL headspace bottle, detecting by using HS-GC/MS,
wherein the unknown impurities are diethylnitrosamine and N-methyl-N-nitrosomethylamine.
2. The detection method according to claim 1, wherein the step S2 includes: taking 0.3g of the powder obtained in the step S1, adding 0.6-1.2mL of a mixed solvent comprising N, N-dimethylformamide and isopropanol, placing the mixture into a 20mL headspace bottle, sealing, and detecting by using HS-GC/MS.
3. The detection method according to claim 2, wherein the volume ratio of N, N-dimethylformamide to isopropanol is 1: (2-4).
4. The detection method according to claim 1, wherein the HS test conditions are: headspace heating box temperature: 140 ℃; quantitative ring temperature: 150 ℃; transmission line temperature: 150 ℃; equilibrium temperature: 70-80 ℃; sample introduction time: 1.0 min; equilibrium retention time: 15 min; filling pressure: 15 psi.
5. The detection method according to claim 4, wherein the HS test conditions are: headspace heating box temperature: 140 ℃; quantitative ring temperature: 150 ℃; transmission line temperature: 150 ℃; equilibrium temperature: 75 ℃; sample introduction time: 1.0 min; equilibrium retention time: 15 min; filling pressure: 15 psi.
6. The detection method according to claim 1, wherein the stationary phase in the gas chromatograph is: 5-7% of cyanopropyl phenyl and 93-95% of methyl polysiloxane.
7. The detection method according to claim 6, wherein the stationary phase in the gas chromatograph is: cyanopropyl phenyl 6% and methyl polysiloxane 94%.
8. The detection method according to claim 7, wherein the gas chromatography conditions are:
sample inlet temperature: 240 ℃; flow rate: 0.8-1.2 mL/min;
shunting mode: the split ratio is 5: 1;
temperature programming: the initial temperature is 70 deg.C, maintained for 4min, increased to 100 deg.C at 2-4 deg.C/min, maintained for 1min, increased to 240 deg.C at 8-12 deg.C/min, and maintained for 3.5 min.
9. The detection method of any one of claims 1 to 8, wherein the test conditions of the mass spectrometry are: type of ion source: EI; scanning mode: MRM; electron energy: 40-60 eV; filament emission current: 60-80 muA; ion source temperature: 220 ℃; quadrupole temperature: 150 ℃; transmission line temperature: 230 ℃ to 230 ℃.
10. The detection method of any one of claims 1 to 8, wherein in the mass spectrometric detection: and (3) quantifying ions: N-methyl-N-nitrosomethylamine: m/z 74, diethylnitrosamine: and m/z is 102.
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