CN115469038A - Method for extracting and detecting insoluble drug in jam and insoluble drug mixture - Google Patents
Method for extracting and detecting insoluble drug in jam and insoluble drug mixture Download PDFInfo
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- 239000003814 drug Substances 0.000 title claims abstract description 63
- 229940079593 drug Drugs 0.000 title claims abstract description 52
- 239000000203 mixture Substances 0.000 title claims abstract description 44
- 238000000034 method Methods 0.000 title claims abstract description 40
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims abstract description 100
- 239000003085 diluting agent Substances 0.000 claims abstract description 58
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 52
- 238000000605 extraction Methods 0.000 claims abstract description 41
- 238000001514 detection method Methods 0.000 claims abstract description 37
- 239000012085 test solution Substances 0.000 claims abstract description 21
- 238000004128 high performance liquid chromatography Methods 0.000 claims abstract description 19
- 239000002994 raw material Substances 0.000 claims abstract description 4
- KGFYHTZWPPHNLQ-AWEZNQCLSA-N rivaroxaban Chemical compound S1C(Cl)=CC=C1C(=O)NC[C@@H]1OC(=O)N(C=2C=CC(=CC=2)N2C(COCC2)=O)C1 KGFYHTZWPPHNLQ-AWEZNQCLSA-N 0.000 claims description 73
- 229960001148 rivaroxaban Drugs 0.000 claims description 73
- 239000000843 powder Substances 0.000 claims description 29
- 239000000243 solution Substances 0.000 claims description 22
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 22
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 claims description 12
- 239000012088 reference solution Substances 0.000 claims description 11
- 239000007788 liquid Substances 0.000 claims description 10
- 238000009210 therapy by ultrasound Methods 0.000 claims description 7
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 6
- 229910000147 aluminium phosphate Inorganic materials 0.000 claims description 6
- 238000005119 centrifugation Methods 0.000 claims description 6
- 239000006228 supernatant Substances 0.000 claims description 4
- 239000000945 filler Substances 0.000 claims description 3
- 239000011259 mixed solution Substances 0.000 claims description 3
- YTJSFYQNRXLOIC-UHFFFAOYSA-N octadecylsilane Chemical compound CCCCCCCCCCCCCCCCCC[SiH3] YTJSFYQNRXLOIC-UHFFFAOYSA-N 0.000 claims description 3
- 239000000377 silicon dioxide Substances 0.000 claims description 3
- 239000003643 water by type Substances 0.000 claims description 3
- 238000002525 ultrasonication Methods 0.000 claims 1
- 230000000694 effects Effects 0.000 abstract description 6
- 238000004587 chromatography analysis Methods 0.000 abstract description 3
- 230000000052 comparative effect Effects 0.000 description 23
- 238000012360 testing method Methods 0.000 description 17
- 239000001814 pectin Substances 0.000 description 7
- 235000010987 pectin Nutrition 0.000 description 7
- 229920001277 pectin Polymers 0.000 description 7
- 238000010200 validation analysis Methods 0.000 description 7
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 6
- 239000013558 reference substance Substances 0.000 description 6
- 239000000523 sample Substances 0.000 description 6
- 239000000126 substance Substances 0.000 description 5
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- 206010047249 Venous thrombosis Diseases 0.000 description 3
- 238000007865 diluting Methods 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- 206010051055 Deep vein thrombosis Diseases 0.000 description 2
- 239000007864 aqueous solution Substances 0.000 description 2
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- 229910001220 stainless steel Inorganic materials 0.000 description 2
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- 238000013517 stratification Methods 0.000 description 2
- 238000005303 weighing Methods 0.000 description 2
- 235000021446 Apple puree Nutrition 0.000 description 1
- 208000010378 Pulmonary Embolism Diseases 0.000 description 1
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- 238000002474 experimental method Methods 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- 238000000227 grinding Methods 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 210000003127 knee Anatomy 0.000 description 1
- 239000002075 main ingredient Substances 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 239000004570 mortar (masonry) Substances 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 239000002547 new drug Substances 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 238000011002 quantification Methods 0.000 description 1
- 238000004366 reverse phase liquid chromatography Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 239000007921 spray Substances 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 238000001195 ultra high performance liquid chromatography Methods 0.000 description 1
- 238000002604 ultrasonography Methods 0.000 description 1
- 239000011345 viscous material Substances 0.000 description 1
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- G—PHYSICS
- G01—MEASURING; TESTING
- 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
- G01N30/02—Column chromatography
-
- G—PHYSICS
- G01—MEASURING; TESTING
- 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
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
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- General Health & Medical Sciences (AREA)
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Abstract
The invention discloses a method for extracting and detecting insoluble medicines in a jam and insoluble medicine mixture. The extraction method comprises the following steps: adding a diluent to the mixture of jam and insoluble drug to obtain an extraction system; the diluent is prepared from the following components in a volume ratio of 1:1 to 1:5 of acetonitrile and ethanol. The detection method comprises obtaining an extraction system by the above extraction method, and performing high performance liquid chromatography detection to obtain the content of the insoluble drug as the raw material of the test solution. The diluent with a specific proportion can reduce the wrapping effect of jam on the insoluble medicine, has a better dissolving effect on the insoluble medicine, and can completely extract the insoluble medicine in the mixture; and the detection by the high performance liquid chromatography meets the peak detection requirement of the chromatography, the detection of the main components is accurate and reliable, and the detection accuracy of the insoluble drug in the jam and the insoluble drug mixture is greatly improved.
Description
Technical Field
The invention relates to the technical field of drug analysis, in particular to a method for extracting and detecting an insoluble drug in a mixture of jam and the insoluble drug.
Background
There are many types of drugs and administration methods disclosed in the prior art, and some patients who lose the ability to swallow the whole tablet cannot take the tablet with good difficulty, and in order to achieve smooth administration, the tablet is usually crushed into powder and mixed with a fluid jam such as apple jam. Wherein, the rivaroxaban tablet is a new drug developed by Bayer medicines, and the trade name is as follows: bairuituo
Mainly for hip or knee jointsPrevention of venous thrombosis following replacement; treating adult deep vein thrombosis, and reducing acute deep vein thrombosis recurrence and pulmonary embolism risk. In the administration option, since some patients in such conditions lose the ability to swallow whole tablets, it is desirable to administer rivaroxaban in a form in which the tablets are crushed and mixed with a jam such as apple jam.
Rivaroxaban is a main component in rivaroxaban tablets, and in order to ensure the accuracy of the dosage of the main component in the mixing process of rivaroxaban tablet powder and jam, the main component in the rivaroxaban tablet powder and jam mixture needs to be extracted and measured. The conventional method for determining the content of rivaroxaban in rivaroxaban tablets disclosed in the prior art comprises the following steps: dissolving and extracting rivaroxaban in rivaroxaban tablet powder by using acetonitrile or methanol and an aqueous solution thereof as a diluent, preparing a test solution, and determining the content of rivaroxaban by adopting a high performance liquid chromatography.
The diluent disclosed above can realize the extraction of rivaroxaban in rivaroxaban tablets; however, for the special administration mode of taking the mixture of the jam and the rivaroxaban tablet powder, the conventionally adopted diluent is not suitable for extracting rivaroxaban from the mixture of the jam and the rivaroxaban tablet powder, because the jam is thick, the rivaroxaban tablet powder is completely wrapped, the dissolution and extraction of acetonitrile or methanol and an aqueous solution thereof on rivaroxaban are prevented, and the phenomenon of layering of the jam and the solution exists, so that the content of the rivaroxaban component cannot be accurately determined.
Disclosure of Invention
Therefore, the technical problem to be solved by the invention is to overcome the defect of incomplete extraction of rivaroxaban in a mixture of insoluble medicines such as jam and rivaroxaban tablet powder in the prior art, and thus provide the method for extracting the insoluble medicine from the jam and insoluble medicine mixture to solve the problem.
The invention also aims to solve the other technical problem of overcoming the defect of inaccurate rivaroxaban content measurement in the jam caused by incomplete extraction of the rivaroxaban and other insoluble medicines in the prior art, and thus provides a method for detecting the insoluble medicines in the jam and the insoluble medicine mixture.
The technical scheme of the application is as follows:
a method for extracting insoluble drug from jam and insoluble drug mixture comprises the following steps: adding a diluent to the mixture of jam and insoluble drug to obtain an extraction system; the diluent is prepared from the following components in a volume ratio of 1:1 to 1:5 of acetonitrile and ethanol.
The slightly soluble medicine is rivaroxaban tablet powder.
The concentration of the rivaroxaban tablet powder in an extraction system is 0.01-0.03 mg/ml, and the volume ratio of the diluent to the extraction system is at least 240:500.
also comprises the step of adding water to the mixture of the jam and the insoluble drug and washing; preferably, the rinsing step is performed before the diluent is added.
Also comprises the step of ultrasound; preferably, the diluent is added in one or more times, and the ultrasonic step is arranged after the diluent is added for the first time; more preferably, the ultrasonic frequency is 40KHz, the ultrasonic power is 500w, the power adjusting range is 80-100%, and the ultrasonic time is 5-30 minutes.
Also comprises the step of fixing the volume; preferably, the step of fixing volume is arranged after the ultrasonic treatment, and water and/or diluent are added during the fixing volume.
The extraction system at least contains rivaroxaban tablet powder and other insoluble medicines, jam and diluent, and when the extraction system comprises the steps of washing with water and/or adding water to a constant volume, the extraction system contains rivaroxaban tablet powder and other insoluble medicines, jam, diluent and water.
Further comprising the step of centrifuging the extraction system to obtain a supernatant; preferably, the centrifugation step is set to be 12000-15000 r/min after constant volume and 5-15 min of centrifugation time.
A detection method of an insoluble drug in a jam and insoluble drug mixture is characterized in that an extraction system is obtained by adopting the extraction method of the insoluble drug in the jam and insoluble drug mixture, and the extraction system is used as a raw material of a test solution to be subjected to high performance liquid chromatography detection to obtain the content of the insoluble drug.
The detection process of the high performance liquid chromatography comprises the following steps: preparing a test solution and a reference solution of rivaroxaban tablets, and respectively detecting and analyzing the test solution and the reference solution by adopting a high performance liquid chromatograph;
the chromatographic conditions of the high performance liquid chromatography detection are as follows: the chromatographic column with octadecylsilane chemically bonded silica as filler has mobile phase of mixed solution of 0.01mol/L phosphoric acid solution and acetonitrile at 40-50 deg.c and flow rate of 0.4-0.6 ml/min.
At least one of the following conditions is satisfied:
the chromatographic column is a C18 column; preferably, the chromatographic column is Agilent XDB-C18 or Waters BEH C18;
the detection wavelength of the high performance liquid chromatograph is 250nm;
the sample volume of the test solution and the reference solution is 1-3 mul.
The technical scheme of the invention has the following advantages:
according to the method for extracting the insoluble drug from the jam and the insoluble drug mixture, a diluent (acetonitrile: ethanol is 1:1 to 1:5v/v ], on the first hand, the specific proportion of the diluent can enable pectin in the jam to become flocculent and separate out, so that the wrapping effect of the jam on insoluble drugs is reduced, the component interference of the pectin on the insoluble drugs is reduced, and the solution fluidity is good; in the second aspect, the diluent with a specific proportion has a better dissolving effect on the insoluble medicine, and can completely extract the insoluble medicine in the mixture; in the third aspect, the slightly-soluble drug extract extracted by the diluent with a specific proportion meets the peak requirement of chromatography when being detected by high performance liquid chromatography, the detection of the main component is accurate and reliable, and the detection accuracy of the slightly-soluble drug in the jam and the mixture of the slightly-soluble drugs is greatly improved.
The concentration of the rivaroxaban tablet powder in an extraction system is 0.01-0.03 mg/ml, and the volume ratio of the diluent to the extraction system is at least 240:500 to fully flocculate the pectin with sufficient diluent, eliminate interference and fully extract the main component rivaroxaban.
Preferably, the step of washing is carried out by diluting with water before the diluent is added, so that the mixture in the container can be sufficiently transferred into a measuring flask, and meanwhile, the jam mixed with rivaroxaban tablet powder is smoothly taken to the gastrointestinal tract through water when a person simulates normal eating.
Preferably, the method further comprises a step of carrying out ultrasonic treatment on the extraction system, and preferably, the ultrasonic treatment step is arranged after the diluent is added for the first time so as to completely and quickly extract the main component; preferably, the ultrasonic frequency is 40KHz, the ultrasonic power is 500w, the power adjusting range is 80-100%, and the ultrasonic time is 5-30 minutes.
And preferably, adding water and/or a diluent for constant volume after ultrasonic treatment, ensuring that the constant volume is more accurate, and obtaining a rivaroxaban extraction system suitable for high performance liquid chromatography detection under the condition of meeting the minimum addition of the diluent.
The method also comprises the step of centrifuging the extraction system after constant volume to separate the floccule from the extraction solution to obtain supernatant, preferably, the centrifugation speed is 12000-15000 r/min, and the centrifugation time is 5-15 min.
According to the method for detecting the insoluble drug in the jam and insoluble drug mixture, the extraction system is obtained by adopting the extraction method and is used as a raw material of a test solution to be detected for high performance liquid chromatography detection, the content of the main component of the insoluble drug is accurately and reliably detected, and the detection accuracy of the insoluble drug in the jam and insoluble drug mixture is greatly improved.
Preferably, the high performance liquid chromatography detection process is as follows: preparing a test solution and a reference solution of rivaroxaban tablets, and respectively detecting and analyzing the test solution and the reference solution by adopting a high performance liquid chromatograph; the chromatographic conditions of the high performance liquid chromatography detection are as follows: the chromatographic column with octadecylsilane chemically bonded silica as filler has mobile phase of mixed solution of 0.01mol/L phosphoric acid solution and acetonitrile at 40-50 deg.c and flow rate of 0.4-0.6 ml/min. The chromatographic conditions are selected, so that the detected rivaroxaban content has no obvious difference (less than or equal to 2.0%) from the actual content, and the precision is high.
The chromatographic column is a C18 column; preferably, the chromatography column is Agilent XDB-C18 or Waters BEH C18. The method realizes effective separation of rivaroxaban main components and various interferents by a reversed phase chromatography method, and is convenient for accurate quantification.
The detection wavelength of the high performance liquid chromatograph is 250nm, and the wavelength is the maximum absorption wavelength of rivaroxaban, so that the detection sensitivity and accuracy are facilitated.
The sample amount of the test solution and the reference solution is 1-3 mul, and the method is suitable for the tolerance of the ultra-high performance liquid chromatography column.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.
FIG. 1 is a chromatogram of a test solution of example 2;
FIG. 2 is a partial chromatogram of a test solution of comparative example 4;
FIG. 3a is a chromatogram of a solution of a methodological validation control; FIG. 3b is a chromatogram of a blank solution of a methodological validation adjuvant; FIG. 3c is a chromatogram of a methodically validated test sample solution;
FIG. 4 is a methodological validation linear chromatogram;
a：0.010mg/ml；b：0.015mg/ml；c：0.020mg/ml；d：0.025mg/ml；e：0.030mg/ml；
FIG. 5 is a methodology validation accuracy chromatogram;
a: a blank sample of auxiliary materials; b to d:3 low concentration samples at a concentration of 0.01mg/ml (corresponding to 50% of the detected concentration); e to g:3 medium concentration samples with a concentration of 0.02mg/ml (corresponding to the detection concentration of 100%); h to j:3 high concentration samples with a concentration of 0.03mg/ml (corresponding to a detection concentration of 150%);
FIG. 6 is a chromatogram for validating precision of sample injection by methodology;
continuously feeding a sample a to a first needle by using the same reference substance solution; b: a second needle; c: a third needle; d: a fourth needle; e, a fifth needle; f: a sixth needle;
FIG. 7 is a chromatogram of a methodology verifying repeatability tests;
a to f: one control sample is continuously injected for 6 times, g, h: and continuously injecting another reference substance for 2 times, wherein i-n: 6 parts of test solution;
FIG. 8 is a chromatogram for verifying the stability of a control solution by methodology;
same control solution at room temperature a:0 hour; b:1 hour c:2 hours; d:4 hours; e:6 hours; f:8 hours; g:10 hours; h: injecting samples for 12 hours;
FIG. 9 is a chromatogram for methodology verifying the stability of a test sample solution;
the same test solution was mixed at room temperature a:0 hour; b:1 hour c:2 hours; d:4 hours; e:6 hours; f:8 hours; g:10 hours; h: samples were injected for 12 hours.
Detailed Description
Example 1
Reagent: acetonitrile, chromatographically pure, 99.8% concentration; ethanol, analytically pure, with the concentration of 95% -100%; apple jam: the information is shown in table 1. Rivaroxaban tablets: the content of the main component is 10 mg/tablet, and the weight of each tablet is about 90mg. The rivaroxaban tablet has three specifications of rivaroxaban content of 10 mg/tablet, 15 mg/tablet and 20 mg/tablet as main components, and the weight of each tablet is about 90mg. The mixed jam of the rivaroxaban tablet powder of the three specifications can be 70ml (about 80 g). Therefore, for such special administration mode that the jam is mixed with the rivaroxaban tablet powder for administration, the mass ratio of the rivaroxaban tablet powder to the jam is usually 9:8000, rivaroxaban main ingredient: the jam mass ratio is 1:8000 to 1:4000.
the method for detecting rivaroxaban in the mixture of the jam and the rivaroxaban tablet powder comprises the following steps:
step one, extracting rivaroxaban from a mixture of jam and rivaroxaban tablet powder
(1) Preparing rivaroxaban tablet powder and apple jam mixture: weighing 80g (about 70 ml) of apple jam in a proper container 1, taking 1 rivaroxaban tablet (specification of 10 mg) in the container, crushing or grinding in a stainless steel mortar, completely transferring to the weighed apple jam, taking a stainless steel spoon, fully stirring, and fully mixing the rivaroxaban tablet powder and the apple jam to obtain a mixture of the rivaroxaban tablet powder and the apple jam.
(2) Preparing a test solution: the entire mixture of rivaroxaban tablet powder and applesauce was transferred to a 500ml measuring flask, 65ml of water was taken to wash the container thoroughly, the solution was collected in the same 500ml measuring flask, and the washing was repeated 1 more times with 65ml of water and collected in the same 500ml measuring flask. About 200ml of diluent 1 (acetonitrile: ethanol = 1) is added into the measuring flask, ultrasonic treatment is carried out for 15min, the measuring flask is continuously diluted to a scale with the diluent 1, volume fixing and shaking are carried out, and the concentration of the rivaroxaban in the extraction system is 0.02mg/ml. Taking a proper amount of mixed liquid in the measuring flask, placing the mixed liquid in a 1.5ml centrifuge tube, centrifuging for 10min at 15000rpm, and taking supernatant as a test solution.
Total amount of diluent 1 was about 300ml, volume ratio of acetonitrile to ethanol 1:1.
TABLE 1 jam information used in the experiment
(3) Preparation of control solutions: taking about 10mg of a reference substance, precisely weighing, placing in a 500ml measuring flask, respectively adding 100ml of acetonitrile and ethanol, performing ultrasonic treatment for 15min, adding 200ml of water, diluting with ethanol to a scale, and shaking up to obtain a rivaroxaban reference substance solution of 0.02mg/ml.
Step two, detecting the rivaroxaban content in the jam by high performance liquid chromatography
Respectively sucking 1 μ l of each of the reference solution and the sample solution, injecting into high performance liquid chromatograph, and reading data.
Chromatographic conditions of a high performance liquid chromatograph:
and (3) chromatographic column: c18,2.1 mm. Times.50mm, 1.7 μm;
mobile phase: 0.01mol/L phosphoric acid solution-acetonitrile (75;
the preparation method of the 0.01mol/L phosphoric acid solution comprises the following steps: taking 11.5g or 6.7ml of 85% phosphoric acid, diluting to 10.0L with water, and measuring the pH value;
sample introduction amount: 1. Mu.l, flow rate: 0.5ml/min;
column temperature: 45 ℃;
detection wavelength: UV 250nm.
Example 2
This example differs from example 1 in the proportion of diluent and the other parameters and procedures are the same. Diluent 2 in this example was acetonitrile: ethanol = (1: adding 100ml acetonitrile and 100ml ethanol, carrying out ultrasonic vibration treatment, and then fixing the volume by using 100ml ethanol.
The test spectrum is shown in FIG. 2.
Example 3
This example differs from example 1 in the proportion of diluent and the other parameters and procedures are the same. Diluent 3 in this example was acetonitrile: ethanol = (1: 50ml of acetonitrile and 100ml of ethanol are added, ultrasonic shaking treatment is carried out, and then 150ml of ethanol is used for constant volume.
Example 4
This example differs from example 2 in the amount of diluent 2 added and the amount of water added, and the other parameters and steps are the same. In this example, about 240ml of diluent 2 (acetonitrile: 95% ethanol =1: 2v/v) was sonicated for 15min, diluted to the mark with water, and shaken up.
Example 5
This example differs from example 2 in the order of addition of water and diluent 2 in that the mixture of rivaroxaban flakes and applesauce was transferred to a 500ml measuring flask, 65ml of water was taken to wash the container thoroughly, and the solution was collected in the same 500ml measuring flask. To this flask was added about 300ml of diluent 2 (acetonitrile: ethanol = 12 v/v), sonicated for 15min, diluted to the mark with water, shaken well.
Comparative example 1
This comparative example differs from example 1 in the proportion of diluent, diluent 4 in this comparative example being acetonitrile: water = (2. The diluent adding process comprises the following steps: 100ml of acetonitrile and 100ml of water are added firstly, and the volume is determined by 100ml of acetonitrile after ultrasonic shaking treatment.
Comparative example 2
This comparative example differs from example 1 in the composition of the diluent, and diluent 5 in this comparative example is all acetonitrile. The adding process of the diluent 5 is as follows: 200ml of acetonitrile is added firstly, the volume is determined by the acetonitrile after ultrasonic shaking treatment, and the total volume of the acetonitrile is about 300ml.
Comparative example 3
This comparative example differs from example 1 in the composition of the diluent, diluent 6 in this comparative example being (acetonitrile: ethanol =2 1v/v. The adding process of the diluent 6 is as follows: 100ml acetonitrile and 100ml ethanol, and after ultrasonic shaking treatment, 100ml acetonitrile is used for constant volume.
Comparative example 4
This comparative example differs from example 1 in the composition of the diluent, and the diluent 7 in this comparative example is all ethanol. The diluent adding process comprises the following steps: adding 200ml ethanol, carrying out ultrasonic vibration treatment, and then using the ethanol to fix the volume, wherein the total volume of the ethanol is about 300ml.
Comparison of results
The measurement results of examples 1 to 5 and comparative examples 1 to 4 are shown in Table 2.
Comparative example 1 has a sticky substance such as pectin and the like, and the rivaroxaban tablet powder is layered in a volumetric flask, and cannot be completely extracted when mixed in the sticky substance. Comparative example 1 demonstrates that the diluent used 66.6% acetonitrile in water, there was a delamination phenomenon, rivaroxaban could not be detached from the jam, and only 62.97% of the main component could be detected.
Comparative example 2 has a sticky substance such as pectin and the like, and is layered in a volumetric flask, and a part of rivaroxaban tablet powder is mixed in the sticky substance and cannot be completely extracted. Comparative example 2 demonstrates that rivaroxaban could not be completely extracted using acetonitrile as the diluent, and that stratification was still present, only 82.82% of the major component could be detected.
Comparative example 3 pectin and other viscous substances were less and no significant demixing occurred, but extraction was still incomplete. Comparative example 3 proves that the diluent can improve the extraction effect of the main component by combining acetonitrile with a small amount of ethanol, and the detection rate is 93.57%.
Comparative example 4 has no obvious stratification, good solution mobile phase and floccule precipitation, but has solvent effect with the mobile phase under the liquid chromatography condition, and has poor chromatographic peak shape, which affects the determination accuracy and the service life of the chromatographic column, as shown in fig. 2. Comparative example 4 proves that complete use of ethanol can achieve flocculent separation of pectin in the jam, but the peak shape of chromatographic peak is poor, which is not beneficial to liquid chromatography detection.
In examples 1 to 5, no obvious layering phenomenon occurs, the mixture in the measuring flask has good mobile phase, floccules are separated out from the content, and the main component is completely extracted. Rivaroxaban reaches the detection rate of more than 98%, and the detection accuracy is high.
Therefore, as can be seen from table 2, when the total volume of acetonitrile, ethanol (1 to 1.
TABLE 2 comparison of the apple jam dusting rivaroxaban tablet test examples and comparative examples
Method repeatability
The same apple jam spray test was conducted on rivaroxaban tablets of suzhou neutralization and bayer medicine simultaneously using the method of example 2, with the following measurements:
TABLE 3 comparison of test contents of the dummy and reference formulations
Remarking: the content is within 5 percent, and the difference is judged to be not obvious.
As can be seen from the results in Table 3, when 6 portions of each of the two samples were subjected to the apple jam sprinkling test, the detected contents were substantially the same and were not significantly different from the actual contents of the samples (. Ltoreq.2.0%), indicating that both samples could be completely extracted from the apple jam and accurately measured.
Methodology validation data
Methodological validation included specificity, linearity and range, accuracy (recovery), precision, and solution stability tests of the methods of detecting rivaroxaban in the jam and rivaroxaban tablet powder mixtures of the present application.
Wherein the specificity test is as follows: and respectively carrying out high performance liquid chromatography tests on the reference substance solution, the auxiliary material blank solution and the test solution. The linear and range tests were: the high performance liquid chromatography test is carried out on rivaroxaban test sample solutions of 0.010mg/ml, 0.015mg/ml, 0.020mg/ml, 0.025mg/ml and 0.030mg/ml respectively, and a standard curve is drawn to obtain a correlation coefficient.
The accuracy (recovery rate) is that high performance liquid chromatography tests are respectively carried out on a blank auxiliary material sample and 3 low-concentration samples with the concentration of 0.01mg/ml (equivalent to 50 percent of detection concentration), 3 medium-concentration samples with the concentration of 0.02mg/ml (equivalent to 100 percent of detection concentration), 3 high-concentration samples with the concentration of 0.03mg/ml (equivalent to 150 percent of detection concentration), and three concentration recovery rate values and an average recovery rate are calculated.
The precision test comprises continuously injecting 6 reference solutions, calculating rivaroxaban peak area and retention time RSD, and performing high performance liquid chromatography on 6 identical samples to calculate content relative standard deviation.
The solution stability test is as follows: and respectively injecting samples at 0, 1, 2, 4, 6, 8, 10 and 12-hour time nodes of the reference substance solution and the test substance solution which are placed at room temperature within 12 hours, and testing the stability.
TABLE 4 detection methodology validation results for rivaroxaban in apple puree and rivaroxaban tablet powder mixture
As can be seen from Table 4, the method for detecting the insoluble drug in the jam and the insoluble drug mixture has good specificity, and rivaroxaban has good linearity, good accuracy, good precision and good stability within the concentration range of 0.01-0.03 mg/ml.
It should be understood that the above examples are only for clarity of illustration and are not intended to limit the embodiments. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. And are neither required nor exhaustive of all embodiments. And obvious variations or modifications derived therefrom are intended to be within the scope of the invention.
Claims (10)
1. A method for extracting insoluble medicines from a mixture of jam and the insoluble medicines is characterized by comprising the following steps: adding a diluent into a mixture of the jam and the insoluble drug to obtain an extraction system; the diluent is a mixture of 1:1 to 1:5 of acetonitrile and ethanol.
2. The extraction method according to claim 1, wherein the poorly soluble drug is rivaroxaban tablet powder.
3. The extraction method as claimed in claim 2, wherein the concentration of rivaroxaban tablet powder in the extraction system is 0.01mg/ml to 0.03mg/ml, and the volume ratio of the diluent to the extraction system is at least 240:500.
4. the extraction method according to any one of claims 1 to 3, further comprising the step of adding water to the mixture of the jam and the poorly soluble drug; preferably, the rinsing step is provided before the diluent is added.
5. The extraction method according to any one of claims 1 to 4, further comprising a step of ultrasonication; preferably, the diluent is added in one or more portions, and the ultrasonic step is arranged after the diluent is added for the first time; more preferably, the ultrasonic frequency is 40KHz, the ultrasonic power is 500w, the power adjusting range is 80-100%, and the ultrasonic time is 5-30 minutes.
6. The extraction method according to claim 5, further comprising a step of fixing the volume; preferably, the step of fixing volume is arranged after the ultrasonic treatment, and water and/or diluent are added during the fixing volume.
7. The extraction method according to claim 6, further comprising the step of centrifuging the extraction system to obtain a supernatant; preferably, the centrifugation step is set to be 12000-15000 r/min after constant volume and 5-15 min of centrifugation time.
8. A method for detecting insoluble drugs in a mixture of jam and insoluble drugs is characterized in that an extraction system is obtained by adopting the method for extracting the insoluble drugs in the mixture of the jam and the insoluble drugs according to any one of claims 1 to 7, and the extraction system is used as a raw material of a test solution to carry out high performance liquid chromatography detection to obtain the content of the insoluble drugs.
9. The detection method according to claim 8, wherein the high performance liquid chromatography detection process comprises: preparing a test solution and a reference solution of rivaroxaban tablets, and respectively detecting and analyzing the test solution and the reference solution by adopting a high performance liquid chromatograph;
the chromatographic conditions of the high performance liquid chromatography detection are as follows: a chromatographic column using octadecylsilane chemically bonded silica as a filler, wherein a mobile phase is a mixed solution of 0.01mol/L phosphoric acid solution and acetonitrile, the column temperature is 40-50 ℃, and the flow rate is 0.4-0.6 ml/min.
10. The detection method according to claim 9, wherein at least one of the following conditions is satisfied:
the chromatographic column is a C18 column; preferably, the chromatographic column is Agilent XDB-C18 or Waters BEH C18;
the detection wavelength of the high performance liquid chromatograph is 250nm;
the sample amount of the test solution and the reference solution is 1-3 mul.
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