CN116840378A - Method for detecting content of monohydrate of ropivacaine/meloxicam salt - Google Patents
Method for detecting content of monohydrate of ropivacaine/meloxicam salt Download PDFInfo
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- CN116840378A CN116840378A CN202310847200.XA CN202310847200A CN116840378A CN 116840378 A CN116840378 A CN 116840378A CN 202310847200 A CN202310847200 A CN 202310847200A CN 116840378 A CN116840378 A CN 116840378A
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- ropivacaine
- meloxicam
- solution
- monohydrate
- mobile phase
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- ZKMNUMMKYBVTFN-HNNXBMFYSA-N (S)-ropivacaine Chemical class CCCN1CCCC[C@H]1C(=O)NC1=C(C)C=CC=C1C ZKMNUMMKYBVTFN-HNNXBMFYSA-N 0.000 title claims abstract description 87
- 150000004682 monohydrates Chemical class 0.000 title claims abstract description 45
- 238000000034 method Methods 0.000 title claims abstract description 43
- 229960001929 meloxicam Drugs 0.000 claims abstract description 45
- 229960001549 ropivacaine Drugs 0.000 claims abstract description 45
- ZRVUJXDFFKFLMG-UHFFFAOYSA-N Meloxicam Chemical compound OC=1C2=CC=CC=C2S(=O)(=O)N(C)C=1C(=O)NC1=NC=C(C)S1 ZRVUJXDFFKFLMG-UHFFFAOYSA-N 0.000 claims abstract description 44
- OKKJLVBELUTLKV-UHFFFAOYSA-N methanol Substances OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims abstract description 40
- 238000001514 detection method Methods 0.000 claims abstract description 28
- 239000013558 reference substance Substances 0.000 claims abstract description 14
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims abstract description 11
- 238000010828 elution Methods 0.000 claims abstract description 11
- 239000000945 filler Substances 0.000 claims abstract description 5
- YTJSFYQNRXLOIC-UHFFFAOYSA-N octadecylsilane Chemical compound CCCCCCCCCCCCCCCCCC[SiH3] YTJSFYQNRXLOIC-UHFFFAOYSA-N 0.000 claims abstract description 5
- 239000000243 solution Substances 0.000 claims description 62
- 238000004128 high performance liquid chromatography Methods 0.000 claims description 38
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 21
- 238000012360 testing method Methods 0.000 claims description 14
- 239000000523 sample Substances 0.000 claims description 12
- 239000003085 diluting agent Substances 0.000 claims description 10
- -1 ropivacaine/meloxicam salt monohydrate Chemical class 0.000 claims description 8
- 239000000126 substance Substances 0.000 claims description 7
- 239000012085 test solution Substances 0.000 claims description 7
- 239000007788 liquid Substances 0.000 claims description 6
- 238000002347 injection Methods 0.000 claims description 5
- 239000007924 injection Substances 0.000 claims description 5
- 239000012488 sample solution Substances 0.000 claims description 5
- 239000000377 silicon dioxide Substances 0.000 claims description 4
- 229910019142 PO4 Inorganic materials 0.000 claims description 3
- 238000010812 external standard method Methods 0.000 claims description 3
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 claims description 3
- 239000010452 phosphate Substances 0.000 claims description 3
- 239000011259 mixed solution Substances 0.000 claims description 2
- 229910000403 monosodium phosphate Inorganic materials 0.000 claims description 2
- 235000019799 monosodium phosphate Nutrition 0.000 claims description 2
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 claims description 2
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 abstract description 5
- 230000009286 beneficial effect Effects 0.000 abstract description 3
- 150000001875 compounds Chemical class 0.000 abstract description 2
- 239000000741 silica gel Substances 0.000 abstract 1
- 229910002027 silica gel Inorganic materials 0.000 abstract 1
- 239000012071 phase Substances 0.000 description 25
- 238000011084 recovery Methods 0.000 description 8
- 230000000052 comparative effect Effects 0.000 description 7
- 239000003814 drug Substances 0.000 description 7
- 229940079593 drug Drugs 0.000 description 5
- 239000007787 solid Substances 0.000 description 5
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 4
- 239000000843 powder Substances 0.000 description 4
- 238000002360 preparation method Methods 0.000 description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical group CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 238000004090 dissolution Methods 0.000 description 3
- 238000004811 liquid chromatography Methods 0.000 description 3
- 239000003589 local anesthetic agent Substances 0.000 description 3
- 238000003908 quality control method Methods 0.000 description 3
- 239000002904 solvent Substances 0.000 description 3
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 2
- 206010002091 Anaesthesia Diseases 0.000 description 2
- 206010061218 Inflammation Diseases 0.000 description 2
- 208000002193 Pain Diseases 0.000 description 2
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 2
- 150000001408 amides Chemical class 0.000 description 2
- 230000037005 anaesthesia Effects 0.000 description 2
- 229960000074 biopharmaceutical Drugs 0.000 description 2
- 239000013078 crystal Substances 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 230000004054 inflammatory process Effects 0.000 description 2
- 230000014759 maintenance of location Effects 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 230000036407 pain Effects 0.000 description 2
- 238000001556 precipitation Methods 0.000 description 2
- 238000000926 separation method Methods 0.000 description 2
- WGTYBPLFGIVFAS-UHFFFAOYSA-M tetramethylammonium hydroxide Chemical compound [OH-].C[N+](C)(C)C WGTYBPLFGIVFAS-UHFFFAOYSA-M 0.000 description 2
- 238000005303 weighing Methods 0.000 description 2
- WSGYTJNNHPZFKR-UHFFFAOYSA-N 3-hydroxypropanenitrile Chemical group OCCC#N WSGYTJNNHPZFKR-UHFFFAOYSA-N 0.000 description 1
- 238000012935 Averaging Methods 0.000 description 1
- 208000000094 Chronic Pain Diseases 0.000 description 1
- NNJVILVZKWQKPM-UHFFFAOYSA-N Lidocaine Chemical compound CCN(CC)CC(=O)NC1=C(C)C=CC=C1C NNJVILVZKWQKPM-UHFFFAOYSA-N 0.000 description 1
- 206010037660 Pyrexia Diseases 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 230000001154 acute effect Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000012296 anti-solvent Substances 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 239000011258 core-shell material Substances 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000000227 grinding Methods 0.000 description 1
- 229910052739 hydrogen Inorganic materials 0.000 description 1
- 239000001257 hydrogen Substances 0.000 description 1
- 238000005286 illumination Methods 0.000 description 1
- 229960004194 lidocaine Drugs 0.000 description 1
- 235000004213 low-fat Nutrition 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000002844 melting Methods 0.000 description 1
- 230000008018 melting Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 210000005036 nerve Anatomy 0.000 description 1
- 239000000041 non-steroidal anti-inflammatory agent Substances 0.000 description 1
- 229940021182 non-steroidal anti-inflammatory drug Drugs 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 229940094443 oxytocics prostaglandins Drugs 0.000 description 1
- 238000012856 packing Methods 0.000 description 1
- 229960001896 pramocaine Drugs 0.000 description 1
- DQKXQSGTHWVTAD-UHFFFAOYSA-N pramocaine Chemical compound C1=CC(OCCCC)=CC=C1OCCCN1CCOCC1 DQKXQSGTHWVTAD-UHFFFAOYSA-N 0.000 description 1
- MVFGUOIZUNYYSO-UHFFFAOYSA-N prilocaine Chemical compound CCCNC(C)C(=O)NC1=CC=CC=C1C MVFGUOIZUNYYSO-UHFFFAOYSA-N 0.000 description 1
- 229960001807 prilocaine Drugs 0.000 description 1
- 150000003180 prostaglandins Chemical class 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- QEVHRUUCFGRFIF-MDEJGZGSSA-N reserpine Chemical compound O([C@H]1[C@@H]([C@H]([C@H]2C[C@@H]3C4=C(C5=CC=C(OC)C=C5N4)CCN3C[C@H]2C1)C(=O)OC)OC)C(=O)C1=CC(OC)=C(OC)C(OC)=C1 QEVHRUUCFGRFIF-MDEJGZGSSA-N 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 239000007790 solid phase Substances 0.000 description 1
- 238000001228 spectrum Methods 0.000 description 1
- 238000013112 stability test Methods 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 238000003860 storage Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- WROMPOXWARCANT-UHFFFAOYSA-N tfa trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F.OC(=O)C(F)(F)F WROMPOXWARCANT-UHFFFAOYSA-N 0.000 description 1
- 238000009210 therapy by ultrasound Methods 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 238000001291 vacuum drying Methods 0.000 description 1
Classifications
-
- 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
- G01N30/26—Conditioning of the fluid carrier; Flow patterns
- G01N30/28—Control of physical parameters of the fluid carrier
- G01N30/34—Control of physical parameters of the fluid carrier of fluid composition, e.g. gradient
-
- 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
- G01N30/60—Construction of the column
-
- 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
- G01N30/86—Signal analysis
Landscapes
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Nitrogen Condensed Heterocyclic Rings (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention belongs to the technical field of compound detection, and provides a method for detecting the content of monohydrate of ropivacaine/meloxicam salt. The chromatographic column takes octadecylsilane chemically bonded silica gel as a filler, and the column temperature is 35-40 ℃; gradient elution is carried out by taking disodium hydrogen phosphate solution-methanol as a mobile phase; the flow rate is 1.0-1.2mL/min; the detection wavelength is 210-220nm; the monohydrate content of the ropivacaine/meloxicam salt is directly calculated by taking ropivacaine and meloxicam as reference substances, and the method is simple and has high accuracy; the method has the advantages of good stability, high precision and high repeatability; the quality detection index of the monohydrate of the ropivacaine/meloxicam salt is more comprehensive, which is beneficial to better controlling the quality of products.
Description
Technical Field
The invention belongs to the technical field of compound detection, and relates to a method for detecting the content of monohydrate of ropivacaine/meloxicam salt.
Background
Ropivacaine (RVC) is a novel amide local anesthetic with pure S (-) isomer, has the pKa of 8.16, has the characteristic of low fat solubility, and is suitable for the medication of various anesthesia modes such as epidural, lumbar anesthesia, regional nerve block and the like and the treatment of other acute and chronic pains; meloxicam is a non-steroidal anti-inflammatory drug used to treat pain and reduce inflammation by reducing the production of prostaglandins which lead to pain, fever and inflammation.
The two medicines are used simultaneously, and have the defects of poor dissolution efficiency of meloxicam, certain toxic and side effects of ropivacaine, difference of the two medicines in the aspect of administration concentration and the like. The Chinese patent application CN116041342A discloses a new crystal form of a hydrate of ropivacaine/meloxicam salt, which has stable form and definite melting point, good chemical stability, high temperature resistance and illumination resistance, and has the performances required for preparing solid preparations, higher administration concentration, better dissolution, better compressibility and disintegration, convenient storage, simpler production operation and easier quality control.
Currently, the currently common detection methods for ropivacaine and meloxicam are mainly high performance liquid chromatography. The Chinese patent No. 110596272B discloses a detection method of 16 local anesthetic components in an adult external product, which mainly adopts high performance liquid chromatography to detect amide local anesthetic components including lidocaine, prilocaine, ropivacaine, ding Bika factor and pramoxine, and the detection conditions of the high performance liquid chromatography are as follows: a C18 chromatographic column using a sub-3 μm core-shell packing; mobile phase A is dipotassium hydrogen phosphate solution, mobile phase B is methanol acetonitrile, 1:1v/v; the detection wavelength is 210nm; the elution adopts the elution procedure as follows: 0-8min,95% -14% A;8-10min,14% A; the flow rate is 1.0-2.0mL/min; the column temperature is 25-40 ℃; the method improves the detection recovery rate and accuracy, has simple and quick operation process, can obviously improve the sensitivity of the detection method, and has strong practicability.
The Chinese patent application CN107300599A discloses a method for screening acidic drugs in liquid drugs by solid phase extraction-liquid chromatography, wherein a chromatographic column of a high performance liquid chromatography detection test sample is a Hypersil ODS C18 column, a mobile phase A is deionized water, a mobile phase B is acetonitrile and 5% tetramethylammonium hydroxide and water (400-600:10:1000), pH is regulated to 3.5 by phosphoric acid, the volume ratio of the mobile phase A to the mobile phase B is 20:80, and the flow rate of the mobile phase is 1-2mL/min; the method can detect various components including meloxicam simultaneously, and has high separation degree, early peak time and high recovery rate.
The content of the monohydrate of the ropivacaine/meloxicam salt in the medicament is a critical parameter in the quality control process, and no related method can detect the content of the monohydrate of the ropivacaine/meloxicam salt at present; in addition, there are significant differences in the properties of ropivacaine, meloxicam and the monohydrate of ropivacaine/meloxicam salt due to polarity, solubility, etc., and the prior art is not suitable for the detection of the content of ropivacaine/meloxicam salt monohydrate for simultaneous detection of ropivacaine, meloxicam.
Disclosure of Invention
Aiming at the problems existing in the prior art, the technical scheme of the invention is completed by optimizing chromatographic conditions based on a High Performance Liquid Chromatography (HPLC), and the method for detecting the monohydrate content of ropivacaine/meloxicam salt is provided, and has the advantages of good stability, high precision and high repeatability; fills the blank of content detection technology in the quality control of the monohydrate substance of ropivacaine/meloxicam salt.
The technical scheme of the invention is as follows:
a method for detecting the monohydrate content of ropivacaine/meloxicam salt, said method employing high performance liquid chromatography HPLC; the conditions of the HPLC are as follows:
mobile phase: mobile phase A is sodium dihydrogen phosphate solution, mobile phase B is methanol;
gradient elution, elution procedure was as follows:
| time min | Mobile phase a/% | Mobile phase B/% |
| 0-6 | 60 | 40 |
| 6-15.1 | 20 | 80 |
| 15.1-20 | 60 | 40 |
。
For some embodiments of the invention, the mobile phase A is a phosphate solution of 8-12mmol/L and the pH is 2.0-3.0; preferably 10mmol/L of phosphate solution, pH 2.0.
For some embodiments of the invention, the HPLC conditions further comprise:
the chromatographic column uses octadecylsilane chemically bonded silica as filler, 4.6X105 mm,5 μm;
the flow rate of the mobile phase is 1.0-1.2mL/min;
the column temperature is 35-40 ℃;
the detection wavelength is 210-220nm.
Preferably, in some embodiments of the invention, the HPLC conditions are:
the flow rate of the mobile phase is 1.0mL/min;
the column temperature is 35 ℃;
the detection wavelength was 220nm.
Further, the method for the monohydrate content of ropivacaine/meloxicam salt specifically comprises the following steps:
(1) Preparing a reference substance solution and a test substance solution;
(2) Respectively taking a reference substance solution and a test substance solution, and injecting into a high performance liquid chromatograph to detect the content of the monohydrate of the ropivacaine/meloxicam salt;
(3) Calculating the content of the monohydrate of the ropivacaine/meloxicam salt according to an external standard method according to the peak area recorded by HPLC;
the high performance liquid chromatography is performed according to the requirements described in the fourth edition general rule 0512 of the 2015 edition of Chinese pharmacopoeia.
For some embodiments of the invention, the control solution comprises: meloxicam control solution, ropivacaine control solution; the concentration of the meloxicam control solution and the ropivacaine control solution are both 0.2mg/mL.
For some embodiments of the invention, the test solution is a ropivacaine/meloxicam salt monohydrate solution; the concentration was 0.2mg/mL.
As the monohydrate of the ropivacaine/meloxicam salt is combined by means of intramolecular hydrogen bonds, the solution state is in a free state, the content is measured by using the meloxicam control solution and the ropivacaine control solution, and the monohydrate of the ropivacaine/meloxicam salt is further calculated, so that the result is more accurate.
For some embodiments of the present invention, the preparation of the control solution and the test solution uses the following diluents: methanol and 0.4mol/L sodium hydroxide solution.
For some embodiments of the invention, the control solution and the test solution are prepared by adopting a mixed solution of methanol and 0.4mol/L sodium hydroxide solution according to a volume ratio of 50:3 as a diluent.
Because of the solubility difference of the monohydrate of the ropivacaine/meloxicam salt in different solvents, the diluent of the method for preparing the related substances of the meloxicam in Chinese pharmacopoeia is 100mL+0.4mol/L sodium hydroxide 6mL of 40% methanol solution, and the solubility of the diluent to the ropivacaine is poor.
For some embodiments of the invention, the injection amount of the control solution and the test solution is 10-20 μl; preferably 20. Mu.L.
In some embodiments of the invention, the actual content of monohydrate W of the ropivacaine/meloxicam salt r0 The method is calculated according to the following formula:
W r0 ＝W t0 ×W r1 +W t0 ×W r1
wherein W is t0 Theoretical content of monohydrate of ropivacaine/meloxicam salt;
W r1 the actual content of ropivacaine in the monohydrate of ropivacaine/meloxicam salt;
W r2 the actual content of meloxicam in the monohydrate of ropivacaine/meloxicam salt;
further, the method comprises the steps of,
wherein 274.401 is the molecular weight of ropivacaine;
351.401 is the molecular weight of meloxicam;
643.802 is the molecular weight of the monohydrate of ropivacaine/meloxicam salt;
still further, the method further comprises the steps of,
wherein A is t1 Ropivacaine peak area in control;
A t2 as the area of meloxicam Kang Feng in the reference substance;
A r1 peak area of ropivacaine in the test sample;
A r2 kang Feng area of meloxicam in the test sample;
Wt 1 the theoretical content of ropivacaine in the reference substance;
W t2 is the theoretical content of meloxicam in the reference substance;
still further still, the method comprises the steps of,
compared with the prior art, the invention has the following beneficial effects:
1. the chromatographic column using octadecylsilane chemically bonded silica as a filler and a specific mobile phase and elution program are subjected to gradient elution, so that the analysis of a ropivacaine alkaline system and a meloxicam Kang Suanxing system can be simultaneously satisfied; fills the technical blank of a method for detecting the content of the monohydrate of ropivacaine/meloxicam salt;
2. the separation degree of the peak to be detected is further improved by optimizing the pH value, the flow rate and the Wen Dengse spectrum conditions of the mobile phase A, so that the detection precision is improved;
3. as the monohydrate of the ropivacaine/meloxicam salt exists in a solution in a free state of ropivacaine and meloxicam, and the meloxicam Kang Youli state has lower solubility in methanol, the solubility of the monohydrate of the ropivacaine/meloxicam salt is greatly improved by adding a proper amount of sodium hydroxide solution into the methanol as a diluent, and the stability, the precision and the repeatability of a detection result are further improved;
4. the content result of the ropivacaine/meloxicam salt monohydrate obtained by calculating the theoretical content and peak area of the reference ropivacaine and meloxicam is accurate and reliable, and a separate preparation of the ropivacaine/meloxicam salt monohydrate reference substance is not needed;
5. the invention ensures that the quality detection index of the monohydrate of ropivacaine/meloxicam salt in the medicine is more comprehensive, and is beneficial to better controlling the quality of products.
Drawings
FIG. 1 is a graph of HPLC results for monohydrate of ropivacaine/meloxicam salt of lot number QYST02-020-A1 in example 2;
FIG. 2 is a graph of HPLC results for monohydrate of ropivacaine/meloxicam salt of lot number QYST02-033-A1 in example 2.
Detailed Description
The following examples are given to further illustrate the present invention, but are not to be construed as limiting the scope thereof. All techniques implemented based on the above description of the invention are included in the scope of the invention; unless otherwise indicated, materials or reagents provided in the examples are all commonly commercially available products.
(1) Instrument:
instrument model: agilent1260series high performance liquid chromatograph;
chromatographic column: c18 (XBIdge) R 4.6×250mm，5μm)；
(2) Chemical reagent:
control: ropivacaine, chinese pharmaceutical biologicals institute 100866-202003;
meloxicam, chinese pharmaceutical biologics institute 190025-202107;
test article: ropivacaine/meloxicam salt monohydrate, homemade, lot number: QYST02-020-A1, QYST02-033-A1; the preparation method is shown in the Chinese patent application example 1 of application number 202211724956.7, and is specifically as follows:
12.8g of meloxicam and 10.0g of ropivacaine are added into 2.7L of acetone (organic solvent), heated and refluxed at 70 ℃ and stirred to be completely dissolved; concentrating the clear solution under reduced pressure (45 ℃ and minus 0.1 MPa) until solid is precipitated, stopping concentrating, adding 2L of water (antisolvent), stirring at 25 ℃ (solid precipitation temperature) for 17 hours (solid precipitation time), filtering, and vacuum drying the solid sample at 50 ℃ (drying temperature) for 5 hours to obtain a monohydrate I crystal form of ropivacaine meloxicam salt;
reagent: methanol, chromatographic grade, merck company;
disodium hydrogen phosphate, analytically pure, shanghai test.
Example 1
(1) Preparing a solvent:
mobile phase a: 1.42mg of disodium hydrogen phosphate is measured, added into 100mL of water, and phosphoric acid is added to adjust the pH of the solution to 2.0, so as to obtain a disodium hydrogen phosphate solution with the concentration of 10 mmol/L;
mobile phase B: methanol;
a diluent: measuring 100mL of methanol, adding 6mL of 0.4mol/L sodium hydroxide solution, and shaking uniformly for later use;
(2) Control solution:
meloxicam control solution: weighing about 2mg of meloxicam reference substance into 10mL of diluent, and obtaining 0.2mg/mL of meloxicam reference substance solution for later use after dissolving;
ropivacaine control solution: about 2mg of ropivacaine reference substance is weighed into 10mL of diluent, and after dissolution, 0.2mg/mL of ropivacaine reference substance solution is obtained for standby.
(3) Preparing a test solution:
the method comprises the steps of respectively taking the monohydrate of ropivacaine/meloxicam salt serving as a sample of batch numbers QYST02-020-A1 and QYST02-033-A1, grinding the monohydrate into fine powder, taking about 20mg of powder, precisely weighing the powder, placing the powder into a 5mL volumetric flask for ultrasonic treatment for 10 minutes, adding a diluent to 10mL, shaking the mixture uniformly, and filtering the mixture to obtain 2mg/mL of solution for later use.
Example 2
According to the requirements recorded in the fourth code 0512 of the 2015 edition of Chinese pharmacopoeia, respectively precisely sucking 20uL of the control solution prepared in the example 1 and the sample solutions to be tested of the batch numbers QYST02-020-A1 and QYST02-033-A1, and respectively injecting the 20uL into a liquid chromatograph for detection;
the conditions of the high performance liquid chromatography are as follows:
the chromatographic column uses octadecylsilane chemically bonded silica as filler, 4.6X105 mm,5 μm; the column temperature is 35 ℃; the mobile phase A is 10mmol/L disodium hydrogen phosphate solution, and the mobile phase B is methanol; the flow rate is 1.0mL/min; the detection wavelength is 220nm; gradient elution was performed using the elution procedure of table 1;
TABLE 1 Mobile phase elution procedure
| Time/min | Mobile phase a/% | Mobile phase B/% |
| 0-6 | 60 | 40 |
| 6-15.1 | 20 | 80 |
| 15.1-20 | 60 | 40 |
The HPLC result of the high performance liquid chromatography of batch number QYST02-020-A1 is shown in figure 1, and the retention time of ropivacaine and meloxicam is 4.515min and 13.407min respectively; the HPLC result of the high performance liquid chromatography of batch number QYST02-033-A1 is shown in figure 2, and the retention time of ropivacaine and meloxicam is 4.556min and 13.409min respectively; calculating the content of meloxicam and ropivacaine in the test sample according to the external standard method according to the peak area recorded by HPLC, and calculating the content of meloxicam and ropivacaine to obtain the monohydrate of ropivacaine/meloxicam salt, wherein the result is shown in Table 2:
table 2 results of monohydrate content of ropivacaine/meloxicam salt
Example 3
The difference compared to example 2 is only that the detection wavelength by high performance liquid chromatography is 210nm.
Example 4
The only difference compared to example 2 is that the high performance liquid chromatography flow rate is 1.2mL/min.
Comparative example 1
In comparison with example 2, the only difference is that mobile phase A of high performance liquid chromatography is a 0.1% trifluoroacetic acid TFA solution at pH 2.0.
Comparative example 2
The only difference compared to example 2 is that the high performance liquid chromatography flow rate is 0.8mL/min.
Comparative example 3
The only difference compared to example 2 is that the column temperature of the high performance liquid chromatography is 30 ℃.
Comparative example 4
The only difference compared to example 2 is that the column of high performance liquid chromatography is ZORBAX Eclipse Plus C.
Comparative example 5
The only difference compared to example 2 is that the column for high performance liquid chromatography is InfinityLab Poroshell EC-C18.
Comparative example 6
In comparison with example 2, the only difference is that the column of the high performance liquid chromatography is ZORBAX Bonus-RP.
The results of the above examples and comparative examples are shown in Table 3: as can be seen from Table 3, the two reference substances of meloxicam and ropivacaine detected under the condition of liquid chromatography have better peak shapes, the solvent peak and the meloxicam and ropivacaine can be effectively separated, the baseline is stable, and the method is suitable for simultaneously measuring the content of ropivacaine and meloxicam in the monohydrate of the ropivacaine/meloxicam salt.
TABLE 3 detection results for different liquid chromatography conditions
Stability, precision, repeatability, recovery test:
(1) Stability test:
and precisely sucking 20uL of the sample solution, repeating sample injection for 6 times, respectively injecting into a liquid chromatograph at 0, 1, 2, 4 and 8 hours, detecting according to the high performance liquid chromatography condition described in the example 2, recording peak areas, and calculating the peak areas of ropivacaine and meloxicam, wherein RSD is 1.20 percent and 1.05 percent respectively, which indicates that the sample solution is stable within 8 hours.
(2) Precision test:
20uL of each of the ropivacaine control solution and the meloxicam control solution are respectively sucked precisely, the sample injection is respectively repeated 6 times, the peak areas are recorded according to the high performance liquid chromatography condition detection described in the example 2, the peak areas of ropivacaine and meloxicam are calculated, and the RSD is respectively 0.62% and 0.31%.
(3) Repeatability test:
precisely sucking 20uL of the sample solution, repeating sample injection for 6 times, measuring the content of each sample according to the high performance liquid chromatography condition described in the example 2, recording peak area, and averaging 102.55% of the labeled amount of ropivacaine in each sample, wherein RSD is 0.21%; the average meloxicam content of meloxicam Kang Meiluo was 97.97% and the RSD was 0.19%.
(4) Recovery rate test:
precisely measuring 3 parts of ropivacaine control solution and meloxicam control solution prepared in the example 1, wherein the volumes are 1mL, 1.5mL and 2mL respectively, placing the solution in a 10mL volumetric flask, measuring according to the high performance liquid chromatography condition described in the example 2, and calculating the recovery rate; the average recovery rate of ropivacaine is 102.52% and RSD is 0.31%; the average recovery rate of meloxicam is 98.21%, and the RSD is 0.63%.
It follows that the detection limit for the monohydrate of ropivacaine/meloxicam salt is low under the conditions of high performance liquid chromatography according to the present invention; stability, precision, repeatability and recovery rate are high; and the method is simple to operate and high in practicability, and provides technical support for the content detection of the monohydrate of the ropivacaine/meloxicam salt.
Finally, it should be noted that the above description is only for illustrating the technical solution of the present invention, and not for limiting the scope of the present invention, and that the simple modification and equivalent substitution of the technical solution of the present invention can be made by those skilled in the art without departing from the spirit and scope of the technical solution of the present invention.
Claims (10)
1. A method for detecting the monohydrate content of ropivacaine/meloxicam salt, characterized in that the method is performed by high performance liquid chromatography, HPLC, under the following conditions:
mobile phase: mobile phase A is sodium dihydrogen phosphate solution, mobile phase B is methanol;
gradient elution, elution procedure was as follows:
。
2. The method according to claim 1, wherein the mobile phase a is a phosphate solution of 8-12mmol/L and the pH is 2.0-3.0.
3. The method of claim 1, wherein the HPLC conditions further comprise:
the chromatographic column uses octadecylsilane chemically bonded silica as filler, 4.6X105 mm,5 μm;
the flow rate of the mobile phase is 1.0-1.2mL/min;
the column temperature is 35-40 ℃;
the detection wavelength is 210-220nm.
4. A method according to claim 3, wherein the HPLC conditions are: the flow rate of the mobile phase is 1.0mL/min; the column temperature is 35 ℃; the detection wavelength was 220nm.
5. The method according to any one of claims 1-4, characterized in that it comprises in particular the following steps:
(1) Preparing a reference substance solution and a test substance solution;
(2) Respectively taking a reference substance solution and a test substance solution, and injecting into a high performance liquid chromatograph to detect the content of the monohydrate of the ropivacaine/meloxicam salt;
(3) The content of ropivacaine/meloxicam salt monohydrate was calculated according to the external standard method from the peak area recorded by HPLC.
6. The method of claim 5, wherein the control solution comprises: meloxicam control solution, ropivacaine control solution; the concentration of the meloxicam control solution and the ropivacaine control solution are both 0.2mg/mL; the sample solution is a monohydrate solution of ropivacaine/meloxicam salt; the concentration was 0.2mg/mL.
7. The method of claim 5, wherein the control solution and the test solution are prepared using a diluent of: a mixed solution of methanol and 0.4mol/L sodium hydroxide solution.
8. The method according to claim 7, wherein the volume ratio of methanol and 0.4mol/L sodium hydroxide solution is 50:3.
9. The method according to claim 5, wherein the injection amount of the control solution and the test solution in the step (2) is 10 to 20. Mu.L.
10. The process according to claim 5, wherein in step (3) the content W of the monohydrate of ropivacaine/meloxicam salt is r0 The method is calculated according to the following formula:
W r0 ＝97.20％×W r1 +97.20％×W r2
wherein 97.20% is the theoretical content of the monohydrate of ropivacaine/meloxicam salt;
W r1 the actual content of ropivacaine in the monohydrate of ropivacaine/meloxicam salt;
W r2 the actual content of meloxicam in the monohydrate of ropivacaine/meloxicam salt;
the W is r1 、W r2 Is obtained by calculating the peak areas of ropivacaine and meloxicam in the monohydrate sample of the ropivacaine/meloxicam salt and the peak areas of ropivacaine and meloxicam in the control.
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| CN1486690A (en) * | 2002-10-02 | 2004-04-07 | ����ҩƷ��ҵ�ɷ�����˾ | Medicinal salt with local narcosis and antiphlogosis activity and its prepn process |
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