CN116008430A - Gas chromatography method for determining ethanol and propylene glycol in minoxidil external solution - Google Patents
Gas chromatography method for determining ethanol and propylene glycol in minoxidil external solution Download PDFInfo
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- DNIAPMSPPWPWGF-UHFFFAOYSA-N Propylene glycol Chemical compound CC(O)CO DNIAPMSPPWPWGF-UHFFFAOYSA-N 0.000 title claims abstract description 270
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 title claims abstract description 161
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- 238000004587 chromatography analysis Methods 0.000 claims description 2
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- 229910052739 hydrogen Inorganic materials 0.000 claims description 2
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- 238000010813 internal standard method Methods 0.000 abstract description 6
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 abstract 2
- 230000005526 G1 to G0 transition Effects 0.000 abstract 1
- 239000002202 Polyethylene glycol Substances 0.000 abstract 1
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Abstract
The invention provides a gas chromatography method for determining ethanol and propylene glycol in minoxidil external solution, which adopts an HP-INNOWAX capillary column with polyethylene glycol as a stationary phase, uses nitrogen as carrier gas, calculates by peak area according to an internal standard method, and can accurately determine the content of ethanol and propylene glycol. The method adopts a 50:1 split ratio and direct sample injection mode, and has good specificity, linearity, accuracy and precision for measuring the content of ethanol and propylene glycol in the minoxidil external solution.
Description
Technical Field
The invention belongs to the technical field of analysis and detection, and relates to a gas chromatography method for simultaneously measuring ethanol and propylene glycol in minoxidil external solution.
Background
Minoxidil, chemical name: 6- (1-piperidinyl) -2, 4-pyrimidinediamine-3-oxide. The structural formula is as follows:
the molecular formula: c (C) 9 H 15 N 5 O, a colorless to yellowish clear liquid. Minoxidil is an effective topical drug capable of promoting hair growth, the specific mechanism is unknown, and current research suggests that minoxidil can directly stimulate proliferation and differentiation of hair follicle epithelial cells; promoting angiogenesis, increasing local blood flow; opening potassium ion channels; hair follicles are transformed from telogen to anagen.
At present, the ethanol and the propylene glycol in the minoxidil external solution are measured by a gas chromatography method separately, so that the preparation is troublesome and the steps are complex. The ethanol is measured by gas chromatography, the sample preparation process needs to be diluted in multiple steps, the temperature of the sample to be measured needs to be constant to 20 ℃ before the sample injection measurement, the operation steps are complicated, the sample injection is performed by adopting a headspace split-flow sample injection mode, and the repeatability is not as good as that of the direct sample injection. Propylene glycol is a solution of about 10mg of 1, 3-butanediol in 1ml of absolute ethanol as an internal standard solution, which is relatively troublesome to prepare, as measured by gas chromatography. In order to better and more effectively control the product quality of the minoxidil external solution, a gas chromatography method for simultaneously measuring ethanol and propylene glycol in the minoxidil external solution needs to be developed.
Disclosure of Invention
Aiming at the defects of the prior art, the invention aims to provide a gas chromatography method for simultaneously measuring ethanol and propylene glycol in minoxidil external solution, which can accurately and simultaneously measure the content of ethanol and propylene glycol according to an internal standard method and calculate the peak area, and can better control the product quality of the minoxidil external solution.
In order to achieve the aim of the invention, the invention adopts the following technical scheme:
a gas chromatographic method for determining ethanol and propylene glycol in minoxidil external solution, comprising the steps of:
(1) Preparing a solution:
a. a diluent: methanol;
b. internal standard solution: precisely measuring 30ml of n-propanol as an ethanol internal standard, precisely measuring 50ml of n-butanol as a propylene glycol internal standard, placing the n-butanol and the propylene glycol internal standard into the same 200ml measuring flask, diluting to a scale with methanol, and shaking uniformly to obtain an internal standard solution;
c. control solution: precisely measuring 1ml of propylene glycol and 0.5ml of ethanol, placing in the same 50ml measuring flask, adding 4ml of internal standard solution, diluting to scale with methanol, shaking, precisely measuring 1ml of the solution, placing in a 20ml measuring flask, diluting to scale with methanol, shaking, and taking as reference solution;
c. test solution:
2% w/v assay of ethanol test solution: precisely transferring 1ml of 2% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking;
2% w/v assay propylene glycol test solution: precisely transferring 5ml of 2% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking;
5% w/v assay ethanol and propylene glycol test solutions: precisely transferring 2ml of 5% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking;
(2) And (3) detection:
after the system suitability test passed, the analytical determination was started:
a. sample solution to be tested is sampled in sequence;
b. in the verification process, each sequence table finally needs 1 needle of reference substance solution, and RSD (reactive power detector) of peak area ratio of ethanol and propylene glycol of the reference substance solution to the peak area ratio of propylene glycol to the respective internal standard substances of the reference substance solution and the system applicability test 6 needles of reference substance solution is not more than 5.0%;
the chromatographic conditions were as follows:
chromatographic column: HP-INNOWAX,30 mX0.32 mm,0.25 μm or DB-624,30 mX0.53 mm,3 μm; a detector: FID hydrogen flame ionization detector;
flow rate: 1.9-2.1 ml/min;
detector temperature: 245-255 ℃;
sample inlet temperature: 215-225 ℃;
sample injection mode: directly sampling;
split ratio: 50:1, a step of;
sample injection amount: 1 μl;
column temperature: the initial temperature is 40 ℃, maintained for 5 minutes, and the temperature is raised to 220 ℃ at a rate of 15 ℃ per minute, and maintained for 4 minutes;
sample injection mode: and (5) directly sampling.
Further, the system applicability test:
balancing the chromatographic column to a base line before running the sequence, and repeatedly injecting a blank solution until the system is stable and has no interference;
continuously feeding 6 needles of reference substance solution, wherein the theoretical plate number is not lower than 5000 according to ethanol and propylene glycol; the relative standard deviation RSD of the peak area ratios of ethanol and propylene glycol to the respective internal standard should be no more than 5.0%;
in the reference substance solution graph, the separation degree of adjacent peaks is more than or equal to 1.5.
Preferably, in the preparation process of the reference substance solution in the step (1), propylene glycol is firstly moved in, and then ethanol is moved in.
Preferably, the flow rate of the detection method in the step 2 is 2.0ml/min; the detector temperature was 250 ℃; the sample inlet temperature was 220 ℃.
The invention provides a gas chromatography method for simultaneously measuring ethanol and propylene glycol in minoxidil external solution, which is calculated according to peak area by an internal standard method, can simultaneously and accurately measure the content of ethanol and propylene glycol, and can better control the product quality of the minoxidil external solution.
Drawings
FIG. 1 is a graph showing the linear relationship of ethanol content;
FIG. 2 is a graph showing the linear relationship of propylene glycol content;
FIG. 3 GC diagram of control solution;
FIG. 4 GC diagram of the sample solution.
Detailed Description
In order to make the objects, technical solutions and technical effects of the embodiments of the present invention more clear, the technical solutions in the embodiments of the present invention will be clearly and completely described below, and it is obvious that the described embodiments are some embodiments of the present invention. All embodiments obtained by a person of ordinary skill in the art without making any inventive effort in connection with the embodiments of the present invention are intended to fall within the scope of the present invention.
In the description of the present invention, various instruments and materials are described as follows:
1. material
Minoxidil reference (China food and drug inspection institute, lot number 100238-201702; china food and drug inspection institute, lot number 100238-202203);
minoxidil drug substance (Fu Le Mahong Kai (Dalian) medical Co., ltd., FP0240A21002; fu Le Mahong Kai (Dalian) medical Co., FP0240A21013; tianjin pharmaceutical research institute medical industry Co., ltd., 2105025; tianjin pharmaceutical research institute medical industry Co., ltd., 2010046; tianjin pharmaceutical research institute medical industry Co., 2103011);
reference formulation (JOHNSON & JOHNSON consemer INC,3200CP;JOHNSON&JOHNSON CONSUMER INC,0331CP;JOHNSON CONSUMER INC,0102CP;JOHNSON CONSUMER INC,0331CP;JOHNSON CONSUMER INC,1181CP;JOHNSON CONSUMER INC,0902CP);
2. main instrument
Agilent 7890B-GC gas chromatograph
Examples: methodological verification
1. Specialization of
Blank interference, crude drug interference and positioning separation of each component:
blank solution: methanol
Ethanol positioning solution: precisely measuring 1ml of ethanol, placing into a 200ml measuring flask, diluting to the scale with methanol, and shaking.
Propylene glycol positioning solution: precisely measuring 2ml of propylene glycol, placing into a 200ml measuring flask, diluting to the scale with methanol, and shaking.
N-propanol positioning solution: precisely weighing 1ml of n-propanol, placing in a 50ml measuring flask, diluting to the scale with methanol, and shaking.
N-butanol positioning solution: precisely measuring 2ml of n-butanol, placing in a 50ml measuring flask, diluting to the scale with methanol, and shaking.
API solution: accurately weighing 103.73mg of minoxidil, placing in 50ml measuring flask, adding appropriate amount of methanol, dissolving with ultrasound, cooling, diluting with methanol to scale, and shaking.
Internal standard solution: precisely weighing 30ml of n-propanol and 50ml of n-butanol, placing in the same 20ml measuring flask, diluting to scale with methanol, and shaking.
Control solution: precisely measuring 1ml of propylene glycol and 0.5ml of ethanol, placing in a 50ml measuring flask, adding 4ml of internal standard solution, diluting to scale with methanol, shaking, precisely measuring 1ml, placing in a 20ml measuring flask, diluting to scale with methanol, and shaking.
Precisely measuring 1 μl of each of the blank solution, each component positioning solution and API solution, injecting into a gas chromatograph, recording a chromatogram, and examining the method specificity.
Ethanol and propylene glycol content verification-specificity
2. Precision of
Repeatability (experimenter A)
Blank solution: methanol
Control solution: reference "1, proprietary" under control solution.
2% w/v test solution (assay ethanol): precisely transferring 1ml of 2% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking. (parallel preparation of 6 parts)
2% w/v test solution (propylene glycol assay): precisely transferring 5ml of 2% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking. (parallel preparation of 6 parts)
5% w/v test solution (assay ethanol and propylene glycol): precisely transferring 2ml of 5% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking. (parallel preparation of 6 parts)
Precisely measuring 1 μl of the reference solution and each sample solution, injecting into gas chromatograph, measuring, recording chromatogram, and calculating ethanol and propylene glycol content and relative standard deviation of 6 sample solutions according to internal standard method.
The results of the repeatability test are shown in Table 1.
TABLE 1 results of ethanol and propylene glycol content verification-precision-repeatability experiments
The results show that:
6 parts of 2% minoxidil external solution has the ethanol content of 59.1% -61.4% (v/v) of the sample solution, meets the acceptable standard of 55% -65% (v/v), and has the Relative Standard Deviation (RSD) of 1.5% or less than 10.0%; meets the requirements.
6 parts of 2% minoxidil external solution, the propylene glycol content of the sample solution is between 17.9% and 20.8% (v/v), the propylene glycol content accords with the acceptable standard of 15% to 25% (v/v), and the Relative Standard Deviation (RSD) is 5.5% and is not more than 10.0%; meets the requirements.
6 parts of 5% minoxidil external solution, the ethanol content of the test sample solution is between 29.0% and 30.4% (v/v), 25% to 35% (v/v) of the acceptable standard is met, and the Relative Standard Deviation (RSD) is 1.8% and is not more than 10.0%; meets the requirements.
6 parts of 5% minoxidil external solution, the propylene glycol content of the test sample solution is between 46.0% and 53.0% (v/v), the propylene glycol content meets the acceptable standard of 45% to 55% (v/v), and the Relative Standard Deviation (RSD) is 5.4% or less than 10.0%; meets the requirements.
In summary, the method has better repeatability.
Intermediate precision (experimenter B)
Blank solution: methanol
Internal standard solution: precisely weighing 30ml of n-propanol and 50ml of n-butanol, placing in the same 200ml measuring flask, diluting to scale with methanol, and shaking.
Control solution: precisely measuring 1ml of propylene glycol and 0.5ml of ethanol, placing in the same 50ml measuring flask, adding 4ml of internal standard solution, diluting to scale with methanol, shaking, precisely measuring 1ml, placing in 20ml measuring flask, diluting to scale with methanol, and shaking.
2% w/v test solution (assay ethanol): precisely transferring 1ml of 2% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking. 6 parts were prepared in parallel.
2% w/v test solution (propylene glycol assay): precisely transferring 5ml of 2% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking. 6 parts were prepared in parallel.
5% w/v test solution (assay ethanol and propylene glycol): precisely transferring 2ml of 5% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking. 6 parts were prepared in parallel.
Precisely measuring 1 μl of each of the reference solution and the test solution, injecting into a weather chromatograph, measuring according to a method, recording a chromatogram, calculating the content and relative standard deviation of ethanol and propylene glycol in 6 parts of the test solution according to an internal standard method, and simultaneously comparing the difference value of the average value of the ethanol and propylene glycol of the two test persons and the relative standard deviation of the ethanol and propylene glycol content of the 12 parts of the test solution.
The repeatability results of test person B are shown in Table 2.
TABLE 2 results of ethanol and propylene glycol content verification-precision-intermediate precision test
The results show that:
6 parts of 2% minoxidil external solution, the ethanol content of the sample solution is between 58.4% and 59.0% (v/v), which accords with the acceptable standard of 55% to 65% (v/v), and the Relative Standard Deviation (RSD) is 0.5% and is not more than 10.0%; meets the requirements.
6 parts of 2% minoxidil external solution, the propylene glycol content of the sample solution is between 17.4% and 19.9% (v/v), the propylene glycol content accords with the acceptable standard of 15% to 25% (v/v), and the Relative Standard Deviation (RSD) is 4.6% and is not more than 10.0%; meets the requirements.
6 parts of 5% minoxidil external solution have the ethanol content of 29.6-34.2% (v/v) of the sample solution, accord with the acceptable standard of 25-35% (v/v), and have the Relative Standard Deviation (RSD) of 5.9% or less than 10.0%; meets the requirements.
6 parts of 5% minoxidil external solution, the propylene glycol content of the test sample solution is between 48.0% and 49.5% (v/v), the propylene glycol content meets the acceptable standard of 45% to 55% (v/v), and the Relative Standard Deviation (RSD) is 1.3% or less than 10.0%; meets the requirements.
To sum up, the precision results of experimenters A and B are shown in Table 3.
TABLE 3 comparison of ethanol and propylene glycol content verification-precision-intermediate precision test results
The results show that:
(1) The relative standard deviation (RSD%) of the ethanol content of 6 parts of the test sample of the 2% minoxidil external solution of two testers is 1.5% and 0.5%, and the relative standard deviation (RSD%) of the propylene glycol content is 5.5% and 4.6%, respectively, and is not more than 10.0%; meets the requirement of repeatability;
(2) The relative standard deviation (RSD%) of the ethanol content of 6 parts of the 5% minoxidil external solution for two testers is 1.8% and 5.9% respectively, and the relative standard deviation (RSD%) of the propylene glycol content is 5.4% and 1.3% respectively, and is not more than 10.0%; meets the requirement of repeatability;
(3) The average value of the ethanol content of 6 parts of 2% minoxidil external solution for two testers is 61% and 59%, the difference is 2% and is not more than 5%, and the total RSD of 12 parts of the test is 2.1% and is not more than 10.0%; the average value of the propylene glycol content is 19 percent respectively; 19, the difference is 0 and not more than 5%, and the total RSD of 12 samples to be tested is 4.9% and not more than 10.0%; meets the requirement of intermediate precision.
(4) The average value of the ethanol content of 6 parts of 5% minoxidil external solution for two testers is 30% and 31%, the difference is 1% and is not more than 5%, and the total RSD of 12 parts of the 5% minoxidil external solution for two testers is 4.3% and is not more than 10.0%; the average value of the propylene glycol content is 48 percent respectively; 49%, the difference is 1%, not more than 5%, and the total RSD of 12 samples is 3.8%, not more than 10.0%; meets the requirement of intermediate precision.
In summary, the method has better intermediate precision.
3. Linearity and range
Blank solution: methanol
Internal standard solution: 30ml of n-propanol (as an ethanol internal standard) and 50ml of n-butanol (as a propylene glycol internal standard) are precisely measured, placed in the same 200ml measuring flask, diluted to scale with methanol and shaken well.
Internal standard stock solution: precisely measuring 4ml of internal standard solution, placing in a 50ml measuring flask, diluting to the scale with methanol, and shaking.
Adding standard stock solution into the reference substance: precisely measuring 2ml of propylene glycol and 1ml of ethanol, placing into a 200ml measuring flask, diluting to scale with methanol, and shaking.
Lin-200% solution: precisely measuring 4ml of reference substance, adding 4ml of standard stock solution and 1ml of internal standard stock solution, placing into the same 20ml measuring flask, diluting to scale with methanol, and shaking.
Lin-130% solution: precisely measuring 2.6ml of reference substance, 2.6ml of internal standard stock solution, placing into the same 20ml measuring flask, diluting to scale with methanol, and shaking.
Lin-100% solution: precisely measuring 2ml of reference substance, adding 2ml of standard stock solution and 1ml of internal standard stock solution, placing into the same 20ml measuring flask, diluting to scale with methanol, and shaking.
Lin-80% solution: precisely measuring 1.6ml of reference substance, 1ml of internal standard stock solution, placing into the same 20ml measuring flask, diluting to scale with methanol, and shaking.
Lin-70% solution: precisely measuring 1.4ml of reference substance, 1ml of internal standard stock solution, placing into the same 20ml measuring flask, diluting to scale with methanol, and shaking.
1 μl of each of the above linear solutions was precisely measured, poured into a chromatograph, and the chromatogram was recorded. And (3) taking the concentration as an abscissa and the peak area ratio of the object to be detected to the internal standard as an ordinate, and performing a linear regression equation.
The linear results are shown in Table 4.
TABLE 4 ethanol and propylene glycol content validation-Linear results
Conclusion: the ethanol concentration of the test sample is in the range of 0.0349% v/v to 0.0998% v/v, and the linear regression equation: y= 11.7576x-0.0006, r=1.000, the relative standard deviation of each concentration correction factor is 0.2%, less than 5.0%, and the concentration and peak area ratio have good linear relationship; the propylene glycol concentration of the test sample is in the range of 0.0698% v/v to 0.1995% v/v, and the linear regression equation: y= 7.2878x-0.0045, r=1.000, the relative standard deviation of each concentration correction factor is 0.4%, less than 5.0%, and there is a good linear relationship between concentration and peak area ratio.
4. Accuracy of
Blank solution: blank solutions under the index "3 linear and range" term.
Internal standard solution: index "3 linear and range" term internal standard solution.
Control solution: index "3 linear versus range" control solutions.
Control stock solution: index "3 linear and range" controls were stock solutions labeled.
Minoxidil stock solution: 200.49mg of minoxidil raw material medicine is precisely weighed, placed in a 100ml measuring flask, 8ml of internal standard solution is precisely added, a proper amount of methanol is added, ultrasonic dissolution is carried out, methanol is used for dilution to scale, and shaking is carried out uniformly.
Accuracy solution
Recovery rate of 5% w/v minoxidil topical solution 70%: precisely measuring 1ml of minoxidil stock solution, placing into a 20ml measuring flask, precisely adding 1.4ml of reference stock solution, diluting to scale with methanol, and shaking. (parallel preparation of 3 parts)
Recovery of 5% w/v minoxidil topical solution 100%: precisely measuring 1ml of minoxidil stock solution, placing into a 20ml measuring flask, precisely adding 2ml of reference stock solution, diluting to scale with methanol, and shaking. (parallel preparation of 3 parts)
Recovery of 5% w/v minoxidil topical solution 130%: 1ml of minoxidil stock solution is precisely measured, placed in a 20ml measuring flask, 2.6ml of reference stock solution is precisely added, diluted to scale with methanol, and shaken well. (parallel preparation of 3 parts)
Precisely measuring 1 μl of each of the above accurate solutions, injecting into a gas chromatograph, measuring according to a law, recording a chromatogram, calculating the recovery rate and average recovery rate (n=3) of ethanol and propylene glycol in the recovery solution at each concentration according to an internal standard method, and calculating the recovery rate average value and relative standard deviation of 9 parts of the recovery solution.
The recovery rate test results are shown in Table 5.
TABLE 5 results of ethanol and propylene glycol content verification-recovery test
Conclusion:
the average recovery rate of the ethanol at each concentration of 70%, 100% and 130% (n=3) is 97.1%, 97.5% and 97.0%, respectively, and the average recovery rate is in the range of 90-110%; the average recovery rate at 3 concentrations (n=9) is 97.2%, the relative standard deviation is 0.3% and less than 10.0% in the range of 90% -110%; all meet the requirements.
The average recovery rate of the propylene glycol at each concentration of 70%, 100% and 130% (n=3) is 97.2%, 97.9% and 97.6%, respectively, and the average recovery rate is in the range of 90-110%; the average recovery rate at 3 concentrations (n=9) is 97.6%, the relative standard deviation is 0.4% and less than 10.0% in the range of 90% -110%; all meet the requirements.
In summary, the accuracy of the method is better.
5. Durability of
Solution stability
Control solution: reference "1, proprietary" under control solution.
2% w/v test solution (assay ethanol): the "reproducibility" term is indexed to 2% w/v test solution (assay ethanol).
2% w/v test solution (propylene glycol assay): the "reproducibility" term is indexed to 2% w/v test solution (propylene glycol determination).
5% w/v test solution (assay ethanol and propylene glycol): the 5% w/v test solution under the index "reproducibility" term (determination of ethanol and propylene glycol).
The control solution and the test solution are respectively packaged in parallel into multiple bottles, and are placed under the room temperature condition, 1 μl of the control solution is precisely measured and injected into the gas chromatograph respectively in 0, 8.5, 16 and 24 hours, and 1 μl of the test solution is precisely measured and injected into the gas chromatograph respectively in 0, 10 and 20 hours, and the chromatograms are recorded. RSD was calculated for each time point for the peak area ratio of ethanol and propylene glycol to the respective internal standard.
The solution stability results are shown in Table 6.
TABLE 6 ethanol and propylene glycol content verification-solution stability results
The results show that: the RSD of the peak area ratio of the ethanol and the propylene glycol to the respective internal standard substances measured at each time point of 24 hours of standing at room temperature is less than 10.0%; the RSD of the peak area ratio of the ethanol and the propylene glycol to the internal standard substances measured at each time point is smaller than 10.0% when the sample solution is placed at room temperature for 20 hours, so that the stability of the reference substance solution is good within 24 hours when the reference substance solution is placed at room temperature, and the stability of the sample solution is good within 20 hours when the reference substance solution is placed at room temperature.
Chromatographic conditions
Blank solution: methanol.
Internal standard solution: 30ml of n-propanol (as an ethanol internal standard) and 50ml of n-butanol (as a propylene glycol internal standard) are precisely measured, placed in the same 200ml measuring flask, diluted to scale with methanol, and shaken well.
Control solution: precisely measuring 1ml of propylene glycol and 0.5ml of ethanol, placing into the same 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into 20ml measuring flask, diluting with methanol to scale, shaking
5% w/v test solution (assay ethanol and propylene glycol): precisely transferring 2ml of 5% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking. (parallel preparation of 2 parts)
Precisely measuring 1 μl of blank solution, control solution and sample solution, respectively at detector temperature + -5deg.C, sample inlet temperature + -5deg.C, carrier gas flow rate + -10%, carrier gas flow rate + -5%, chromatographic column replacement of DB-624,30m 0.53mm, and 3 μm, and recording chromatogram.
The content results were compared with the repeatability results (normal condition results) under each condition. The results are shown in Table 7.
TABLE 7 ethanol and propylene glycol content verification-durability chromatographic condition variation results
The results show that: the chromatographic conditions were varied as follows: the temperature of the sample inlet (220 ℃ plus or minus 5 ℃), the temperature of the detector (250 nm plus or minus 5 ℃), the chromatographic column replacement of DB-624,30m 0.53mm,3 μm, the content of ethanol and propylene glycol both meet the regulation, and the content difference value is within plus or minus 10% compared with the normal condition; when the carrier gas flow rate is changed as follows (2.0 ml/min +/-0.2 ml/min), the propylene glycol content is 56% and 56% respectively, and the carrier gas flow rate is reduced to (2.0 ml/min +/-0.1 ml/min) after exceeding the specified range (the specified range is 45% -55% (v/v)), the ethanol content and the propylene glycol content are both in accordance with the specification, and the content difference value between the carrier gas flow rate and the carrier gas flow rate under normal conditions is +/-10%.
In summary, the chromatographic conditions were varied as follows: the method has better durability when the temperature of the sample inlet (220+/-5 ℃), the temperature of the detector (250 nm+/-5 ℃), the chromatographic column is replaced by DB-624,30m is 0.53mm,3 mu m and the flow rate of carrier gas (2.0 ml/min+/-0.1 ml/min) are changed.
Claims (6)
1. A gas chromatographic method for simultaneously determining ethanol and propylene glycol in minoxidil external solution, which is characterized by comprising the following steps:
(1) Preparing a solution:
a. a diluent: methanol;
b. internal standard solution: precisely measuring 30ml of n-propanol as an ethanol internal standard, precisely measuring 50ml of n-butanol as a propylene glycol internal standard, placing the n-butanol and the propylene glycol internal standard into the same 200ml measuring flask, diluting to a scale with methanol, and shaking uniformly to obtain an internal standard solution;
c. control solution: precisely measuring 1ml of propylene glycol and 0.5ml of ethanol, placing in the same 50ml measuring flask, adding 4ml of internal standard solution, diluting to scale with methanol, shaking, precisely measuring 1ml of the solution, placing in a 20ml measuring flask, diluting to scale with methanol, shaking, and taking as reference solution;
d. test solution:
2% w/v assay of ethanol test solution: precisely transferring 1ml of 2% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking;
2% w/v assay propylene glycol test solution: precisely transferring 5ml of 2% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking;
5% w/v assay ethanol and propylene glycol test solutions: precisely transferring 2ml of 5% w/v minoxidil external solution, placing into a 50ml measuring flask, adding 4ml of internal standard solution, diluting with methanol to scale, shaking, precisely measuring 1ml, placing into a 20ml measuring flask, diluting with methanol to scale, and shaking;
(2) And (3) detection:
after the system suitability test passed, the analytical determination was started:
a. sample solution to be tested is sampled in sequence;
b. in the verification process, each sequence table finally needs 1 needle of reference substance solution, and RSD (reactive power detector) of peak area ratio of ethanol and propylene glycol of the reference substance solution to the peak area ratio of propylene glycol to the respective internal standard substances of the reference substance solution and the system applicability test 6 needles of reference substance solution is not more than 5.0%;
the chromatographic conditions were as follows:
chromatographic column: HP-INNOWAX,30 m.times.0.32 mm,0.25 μm; or DB-624,30m 0.53mm,3 μm; a detector: FID hydrogen flame ionization detector;
flow rate: 1.9-2.1 ml/min;
detector temperature: 245-255 ℃;
sample inlet temperature: 215-225 DEG C
Split ratio: 50:1;
sample injection amount: 1 μl;
column temperature: the initial temperature is 40 ℃, maintained for 5 minutes, and the temperature is raised to 220 ℃ at a rate of 15 ℃ per minute, and maintained for 4 minutes;
sample injection mode: and (5) directly sampling.
2. The method for simultaneous determination of ethanol and propylene glycol in a minoxidil topical solution according to claim 1, wherein the system applicability test:
balancing the chromatographic column to a base line before running the sequence, and repeatedly injecting a blank solution until the system is stable and has no interference;
continuously feeding 6 needles of reference substance solution, wherein the theoretical plate number is not lower than 5000 according to ethanol and propylene glycol; the relative standard deviation RSD of the peak area ratios of ethanol and propylene glycol to the respective internal standard should be no more than 5.0%;
in the reference substance solution graph, the separation degree of adjacent peaks is more than or equal to 1.5.
3. The method for simultaneously determining ethanol and propylene glycol in minoxidil topical solution according to claim 1, wherein propylene glycol is first transferred into the control solution in step c, and then transferred into the measuring flask.
4. The method according to claim 1, wherein the carrier gas flow rate of the detection method in the step (2) is 2.0ml/min.
5. The method according to claim 1, wherein the detector temperature of the detection method in the step (2) is 250 ℃.
6. The method for simultaneously determining ethanol and propylene glycol in minoxidil external solution according to claim 1, wherein the sample inlet temperature of the detection method in the step (2) is 220 ℃.
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