CN117825573A - Method for detecting content of hydroxylamine hydrochloride in bulk drug and application - Google Patents
Method for detecting content of hydroxylamine hydrochloride in bulk drug and application Download PDFInfo
- Publication number
- CN117825573A CN117825573A CN202410100936.5A CN202410100936A CN117825573A CN 117825573 A CN117825573 A CN 117825573A CN 202410100936 A CN202410100936 A CN 202410100936A CN 117825573 A CN117825573 A CN 117825573A
- Authority
- CN
- China
- Prior art keywords
- hydroxylamine hydrochloride
- solution
- gas chromatography
- sample
- temperature
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- WTDHULULXKLSOZ-UHFFFAOYSA-N Hydroxylamine hydrochloride Chemical compound Cl.ON WTDHULULXKLSOZ-UHFFFAOYSA-N 0.000 title claims abstract description 147
- 238000000034 method Methods 0.000 title claims abstract description 45
- 239000003814 drug Substances 0.000 title claims abstract description 22
- 229940079593 drug Drugs 0.000 title claims abstract description 18
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims abstract description 75
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical group CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims abstract description 54
- 238000004817 gas chromatography Methods 0.000 claims abstract description 26
- 238000001514 detection method Methods 0.000 claims abstract description 23
- 239000003153 chemical reaction reagent Substances 0.000 claims abstract description 18
- 238000001212 derivatisation Methods 0.000 claims abstract description 16
- 238000002360 preparation method Methods 0.000 claims abstract description 13
- 239000012488 sample solution Substances 0.000 claims abstract description 10
- 239000013558 reference substance Substances 0.000 claims abstract description 6
- 239000000243 solution Substances 0.000 claims description 38
- CGIGDMFJXJATDK-UHFFFAOYSA-N indomethacin Chemical compound CC1=C(CC(O)=O)C2=CC(OC)=CC=C2N1C(=O)C1=CC=C(Cl)C=C1 CGIGDMFJXJATDK-UHFFFAOYSA-N 0.000 claims description 35
- 239000000523 sample Substances 0.000 claims description 25
- 229960000905 indomethacin Drugs 0.000 claims description 20
- 238000002347 injection Methods 0.000 claims description 9
- 239000007924 injection Substances 0.000 claims description 9
- 239000012159 carrier gas Substances 0.000 claims description 7
- 239000012085 test solution Substances 0.000 claims description 7
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 6
- 238000002156 mixing Methods 0.000 claims description 6
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 claims description 4
- 238000007664 blowing Methods 0.000 claims description 4
- 239000003795 chemical substances by application Substances 0.000 claims description 4
- 239000001257 hydrogen Substances 0.000 claims description 4
- 229910052739 hydrogen Inorganic materials 0.000 claims description 4
- 239000000126 substance Substances 0.000 claims description 4
- 239000005485 Azilsartan Substances 0.000 claims description 3
- 239000008186 active pharmaceutical agent Substances 0.000 claims description 3
- KGSXMPPBFPAXLY-UHFFFAOYSA-N azilsartan Chemical group CCOC1=NC2=CC=CC(C(O)=O)=C2N1CC(C=C1)=CC=C1C1=CC=CC=C1C1=NOC(=O)N1 KGSXMPPBFPAXLY-UHFFFAOYSA-N 0.000 claims description 3
- 229960002731 azilsartan Drugs 0.000 claims description 3
- 229940088679 drug related substance Drugs 0.000 claims description 3
- 229910052757 nitrogen Inorganic materials 0.000 claims description 3
- 230000005526 G1 to G0 transition Effects 0.000 claims description 2
- QQQIECGTIMUVDS-UHFFFAOYSA-N N-[[4-[2-(dimethylamino)ethoxy]phenyl]methyl]-3,4-dimethoxybenzamide Chemical compound C1=C(OC)C(OC)=CC=C1C(=O)NCC1=CC=C(OCCN(C)C)C=C1 QQQIECGTIMUVDS-UHFFFAOYSA-N 0.000 claims description 2
- 239000002202 Polyethylene glycol Substances 0.000 claims description 2
- 239000001307 helium Substances 0.000 claims description 2
- 229910052734 helium Inorganic materials 0.000 claims description 2
- SWQJXJOGLNCZEY-UHFFFAOYSA-N helium atom Chemical compound [He] SWQJXJOGLNCZEY-UHFFFAOYSA-N 0.000 claims description 2
- 229960005302 itopride Drugs 0.000 claims description 2
- TZRHLKRLEZJVIJ-UHFFFAOYSA-N parecoxib Chemical compound C1=CC(S(=O)(=O)NC(=O)CC)=CC=C1C1=C(C)ON=C1C1=CC=CC=C1 TZRHLKRLEZJVIJ-UHFFFAOYSA-N 0.000 claims description 2
- 229960004662 parecoxib Drugs 0.000 claims description 2
- 229920001223 polyethylene glycol Polymers 0.000 claims description 2
- 238000004519 manufacturing process Methods 0.000 abstract description 5
- 238000000605 extraction Methods 0.000 abstract description 4
- 238000004458 analytical method Methods 0.000 abstract description 3
- 238000003908 quality control method Methods 0.000 abstract description 3
- 238000000926 separation method Methods 0.000 abstract description 3
- 238000006243 chemical reaction Methods 0.000 abstract description 2
- 125000003544 oxime group Chemical group 0.000 abstract description 2
- 239000012490 blank solution Substances 0.000 description 10
- 239000011259 mixed solution Substances 0.000 description 10
- 238000007865 diluting Methods 0.000 description 8
- 238000011084 recovery Methods 0.000 description 8
- 239000011550 stock solution Substances 0.000 description 8
- 238000012360 testing method Methods 0.000 description 8
- 238000010828 elution Methods 0.000 description 6
- 238000011835 investigation Methods 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- HUMNYLRZRPPJDN-UHFFFAOYSA-N benzaldehyde Chemical compound O=CC1=CC=CC=C1 HUMNYLRZRPPJDN-UHFFFAOYSA-N 0.000 description 4
- 238000010586 diagram Methods 0.000 description 4
- 238000005259 measurement Methods 0.000 description 4
- 239000012086 standard solution Substances 0.000 description 4
- 238000005303 weighing Methods 0.000 description 4
- 239000007789 gas Substances 0.000 description 3
- 239000012088 reference solution Substances 0.000 description 3
- 230000035945 sensitivity Effects 0.000 description 3
- JHIVVAPYMSGYDF-UHFFFAOYSA-N cyclohexanone Chemical compound O=C1CCCCC1 JHIVVAPYMSGYDF-UHFFFAOYSA-N 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 239000012467 final product Substances 0.000 description 2
- 238000004255 ion exchange chromatography Methods 0.000 description 2
- 238000012417 linear regression Methods 0.000 description 2
- 230000014759 maintenance of location Effects 0.000 description 2
- QNGNSVIICDLXHT-UHFFFAOYSA-N para-ethylbenzaldehyde Natural products CCC1=CC=C(C=O)C=C1 QNGNSVIICDLXHT-UHFFFAOYSA-N 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 238000002798 spectrophotometry method Methods 0.000 description 2
- 238000003786 synthesis reaction Methods 0.000 description 2
- 238000010521 absorption reaction Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 239000003638 chemical reducing agent Substances 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 238000006482 condensation reaction Methods 0.000 description 1
- 238000012937 correction Methods 0.000 description 1
- 238000009795 derivation Methods 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000010812 external standard method Methods 0.000 description 1
- 230000007674 genetic toxicity Effects 0.000 description 1
- 231100000025 genetic toxicology Toxicity 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 238000004895 liquid chromatography mass spectrometry Methods 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 238000004949 mass spectrometry Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000007086 side reaction Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 230000009897 systematic effect Effects 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
Landscapes
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention belongs to the technical field of medicine analysis, and relates to a method for detecting the content of hydroxylamine hydrochloride in a bulk drug and application thereof. The method for detecting the content of hydroxylamine hydrochloride in the bulk drug adopts a derivatization method and a gas chromatography to detect the content of the hydroxylamine hydrochloride, and comprises the following steps: preparing a reference substance solution, preparing a sample solution and detecting by gas chromatography; the derivatizing reagent is acetone and methanol. The reagent provided by the invention can be rapidly converted into an oxime structure which is convenient for gas phase detection by utilizing hydroxylamine hydrochloride and acetone, is easy to obtain, simple and convenient to operate, quick in reaction, free of waiting, free of complex operations such as separation and extraction, high in accuracy, good in precision, strong in specificity, wide in linear range, good in linearity between 2.0 and 40 mu g/mL, and suitable for quality control in the production process of bulk drugs, and can be immediately tested after preparation.
Description
Technical Field
The invention belongs to the technical field of medicine analysis, and relates to a method for detecting the content of hydroxylamine hydrochloride in a bulk drug and application thereof.
Background
Hydroxylamine hydrochloride is a reducing agent commonly used in the chemical synthesis industry, and in the production process of indomethacin, the hydroxylamine hydrochloride reacts with the intermediate 1 to obtain the intermediate 2, and the intermediate 2 is a reagent which is necessary to produce the indomethacin, so that the hydroxylamine hydrochloride has genetic toxicity and genetic mutation effects, the residual is detected, and the control limit is very important to ensure the medication safety of the indomethacin. The synthesis route of indomethacin intermediate 2 is as follows:
the current literature reports that analytical methods for hydroxylamine hydrochloride residue include LC-MS, ion chromatography, spectrophotometry, GC derivatization and HPLC derivatization. These methods have advantages and disadvantages, in which the sensitivity and specificity of mass spectrometry and ion chromatography are high, but the cost of the apparatus is high. Spectrophotometry has poor specificity and is easily interfered by the absorption of different drug matrixes.
Chinese patent No. 111855881A discloses a method for detecting hydroxylamine hydrochloride in azilsartan, which uses benzaldehyde as a derivatization reagent, wherein the derivatized solution is required to be stored at 8 ℃, the control requirement on test conditions is high, and the accuracy of a measurement result is at risk; the indometacin intermediate 2 has primary amine groups on the structure, and can generate condensation reaction with benzaldehyde to cause interference of side reaction.
Chinese patent No. 103913537A discloses a method for rapidly determining hydroxylamine hydrochloride content based on gas-phase derivatization chromatography, wherein cyclohexanone is used for derivatization under alkaline conditions, operations such as extraction, centrifugal drying and the like are required, the steps are relatively complicated, the extraction easily causes sample loss in the pretreatment process of the sample, the accuracy is influenced, and the reproducibility of the method is difficult to ensure.
Therefore, it is necessary to explore a method for detecting the hydroxylamine hydrochloride content, which has the advantages of good repeatability, high precision, strong specificity, high sensitivity and good accuracy.
Disclosure of Invention
Aiming at the problems existing in the prior art, the invention aims to provide a method for detecting the content of hydroxylamine hydrochloride in a bulk drug and application thereof.
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:
the invention provides a method for detecting the content of hydroxylamine hydrochloride in a bulk drug, which adopts a derivatization method and a gas chromatography method to detect the content of the hydroxylamine hydrochloride and comprises the following steps:
preparing a reference substance solution, preparing a test sample solution and detecting by gas chromatography.
The derivation principle is as follows:
further, the derivatizing agents are acetone and methanol.
Further, the volume ratio of the acetone to the methanol is 1: (1-5).
Further, the volume ratio of the acetone to the methanol is 1:1.
Further, the preparation of the test solution comprises the following steps: and taking a sample to be detected, dissolving the sample by using a derivatization reagent, and uniformly mixing to obtain a sample solution.
Further, the preparation of the reference solution comprises the following steps: dissolving hydroxylamine hydrochloride standard substance with derivatization reagent, and mixing to obtain reference substance solution.
Further, in the gas chromatography, the gas chromatography detection is performed by adopting a temperature programming mode to perform elution, and the temperature programming program is as follows: the initial temperature is 60-70 ℃, the temperature is kept for 1-5min, then 15-30 ℃/min is raised to 160-180 ℃, and then 25-40 ℃/min is raised to 220-280 ℃ and the temperature is kept for 2-8min.
Further, the gas chromatography detection adopts a temperature programming mode to perform elution, and the temperature programming program is as follows: the initial temperature is 60 ℃, kept for 1min, then 20 ℃/min is raised to 160 ℃, and then 30 ℃/min is raised to 250 ℃ and kept for 5min.
Further, in the gas chromatography detection, the sample injection mode is split sample injection, and the split ratio is 4:1-8:1.
Still further, the split ratio was 5:1.
Further, in the gas chromatography detection, the carrier gas is nitrogen or helium.
Further, in the gas chromatography detection, the carrier gas flow rate is 1.0-2.0 mL.min -1 The sample injection amount is 1-5 mu L.
Further, in the gas chromatography detection, the carrier gas flow rate was 2.0mL min -1 The sample loading was 1. Mu.L.
Further, in the gas chromatography detection, the chromatographic column stationary phase is polyethylene glycol.
Further, the column was an Agilent DB-wax (30 m. Times.0.25 mm,0.25 μm)
Further, in the gas chromatography detection, the temperature of the sample inlet is 230-250 ℃, and the temperature of the detector is 225-245 ℃.
Further, the temperature of the sample inlet was 250℃and the temperature of the detector was 245 ℃.
Further, in the gas chromatography detection, the type of the detector is a FID detector.
Further, in the gas chromatography detection, the hydrogen flow is 36-50mL/min, and further, the hydrogen flow is 40mL/min.
Further, in the gas chromatography detection, the tail blowing flow rate is 10-50mL/min, and further, the tail blowing flow rate is 30mL/min.
Further, in the gas chromatography detection, the air flow rate is 350-450mL/min.
Further, in the gas chromatography detection, the air flow rate was 400mL/min.
The invention also provides application of the method in detecting the content of hydroxylamine hydrochloride in the bulk drug.
Further, the bulk drug is azilsartan, itopride hydrochloride, parecoxib or indomethacin intermediate 2.
In some specific embodiments, a method for detecting the content of hydroxylamine hydrochloride in a drug substance comprises the steps of
(1) Preparation of derivatizing reagents:
mixing acetone and methanol at a volume ratio of 1:1, placing into a volumetric flask, and shaking.
(2) Preparing a blank solution:
taking the prepared derivatization reagent, namely a blank solution.
(3) Preparation of a control solution:
stock solution 1: weighing 20 mg+/-2.0 mg hydroxylamine hydrochloride standard substance, precisely weighing, placing in a 50mL volumetric flask, dissolving with a mixed solution of acetone and methanol (1:1, v/v), diluting to a fixed volume to scale, and shaking uniformly to obtain the final product. Two parts are prepared by the same method. (hydroxylamine hydrochloride: 0.4 mg/mL).
Stock solution 2: and precisely transferring 1mL of the stock solution into a volumetric flask with 1 to 10mL, diluting and fixing the volume to the scale by using a mixed solution (1:1, v/v) of acetone and methanol, and shaking uniformly to obtain the product. Two parts are prepared by the same method. (hydroxylamine hydrochloride: 40.0. Mu.g/mL)
Control solution: accurately transferring 1mL of the stock solution into a volumetric flask with 2 to 10mL, diluting and fixing the volume to the scale by using a mixed solution of acetone and methanol (1:1, v/v), and shaking uniformly. (hydroxylamine hydrochloride: 4.0. Mu.g/mL, 100 ppm) two portions, labeled STD1 and STD2, respectively, were prepared in the same manner.
(4) Preparation of test solution:
a400 mg.+ -.40 mg sample of indomethacin intermediate 2 was precisely weighed, placed in a 10mL volumetric flask, dissolved with a mixed solution of acetone and methanol (1:1, v/v), diluted to volume to scale, and shaken well. Obtaining the product. Two parts labeled TS1 and TS2, respectively, were prepared in the same manner. (Indometacin intermediate 2:40 mg/mL)
(5) Detection by gas chromatography:
the column was Agilent DB-wax (30 m. Times.0.25 mm,0.25 μm); eluting by adopting a temperature programming mode, wherein the temperature programming program is as follows: the initial temperature is 60 ℃, the temperature is kept for 1min, then 20 ℃/min is increased to 160 ℃, and then 30 ℃/min is increased to 250 ℃ and the temperature is kept for 5min; the sample injection mode is split sample injection, and the split ratio is 5:1; the carrier gas is nitrogen, and the flow rate of the carrier gas is 2.0 mL.min -1 The method comprises the steps of carrying out a first treatment on the surface of the The sample injection amount is 1 mu L, the temperature of the sample injection port is 250 ℃, the temperature of the detector is 245 ℃, the type of the detector is an FID detector, the hydrogen flow is 40mL/min, the tail blowing flow is 30mL/min, and the air flow is 400mL/min.
Compared with the prior art, the invention has the following beneficial effects:
the invention adopts derivatization method and gas chromatography to detect the content of hydroxylamine hydrochloride in the bulk drug, utilizes acetone and methanol as derivatization reagents, can rapidly convert the hydroxylamine hydrochloride and the acetone into oxime structures which are convenient for gas phase detection, has rapid reaction, does not need waiting, does not need complicated steps of separation, extraction and the like, can be tested immediately after preparation, has simple operation, reliable result, good precision and strong specificity, has wide linear range, has good linearity between 2.0 mu g/mL and 40 mu g/mL, is suitable for quality control in the production process of the bulk drug, particularly in the production process of indomethacin intermediate 2, and provides theoretical and practical basis for comprehensive and effective control of the quality of the bulk drug.
Drawings
FIG. 1 is a chromatogram of a blank solution;
FIG. 2 is an enlarged partial chromatogram of a blank solution;
FIG. 3 is a chromatogram of the temperature-programmed elution condition (1), in which the chromatographic peaks represent the components: 1. hydroxylamine hydrochloride;
FIG. 4 is a chromatogram of control solution STD (4.0. Mu.g/mL hydroxylamine hydrochloride), with the chromatographic peaks representing the components: 1. hydroxylamine hydrochloride;
FIG. 5 is a partial enlarged view of a chromatogram of the control solution STD (4.0. Mu.g/mL hydroxylamine hydrochloride), in which the chromatographic peaks represent the components: 1. hydroxylamine hydrochloride;
FIG. 6 is a chromatogram of a quantitative limiting solution (2.0. Mu.g/mL hydroxylamine hydrochloride), with the chromatographic peaks representing the components: 1. hydroxylamine hydrochloride;
FIG. 7 is a partial enlarged view of a chromatogram of a quantitative limiting solution (2.0. Mu.g/mL hydroxylamine hydrochloride), in which the chromatographic peaks represent the components: 1. hydroxylamine hydrochloride;
FIG. 8 is a chromatogram of a test solution (40 mg/mL indomethacin intermediate 2);
FIG. 9 is a partial enlarged view of a chromatogram of a test solution (40 mg/mL indomethacin intermediate 2);
FIG. 10 is a linear regression diagram of hydroxylamine hydrochloride;
fig. 11 is a residual diagram of hydroxylamine hydrochloride.
Detailed Description
It is to be noted that the raw materials used in the present invention are all common commercial products, and the sources thereof are not particularly limited. The following instruments and sources of reagents are exemplary illustrations.
Instrument for measuring and controlling the intensity of light
Gas chromatograph (agilent company); MS105DU type electronic balance (mertrel).
Reagent(s)
Methanol and acetone are chromatographic purity, the methanol is purchased from high-purity solvent limited company of Anhui Tiandi in China, and the acetone is purchased from chemical reagent limited company of national medicine group; hydroxylamine hydrochloride standard was purchased from aladine, model J2115489; indometacin intermediate 2 is self-made by the company, and batch number AP2449-2-9319-030-01.
Examples
1. Experimental method investigation
1.1 elution conditions
The elution conditions of temperature programming are optimized and selected under the condition that other chromatographic conditions are kept unchanged.
(1) The elution procedure at the programmed temperature was as follows: the initial temperature is 100 ℃ for 3min, then 20 ℃/min is raised to 180 ℃, and then 30 ℃/min is raised to 250 ℃ for 5min, (the chromatogram is shown in figure 3).
The results are found according to the chromatogram, in FIG. 3, hydroxylamine hydrochloride has poorer peak type, early peak time and does not reach baseline separation.
2. Experimental method
2.1 materials
2.1.1 instruments
Gas chromatograph (agilent company); MS105DU type electronic balance (mertrel).
2.1.2 reagents
Methanol and acetone are chromatographic purity, the methanol is purchased from high-purity solvent limited company of Anhui Tiandi in China, and the acetone is purchased from chemical reagent limited company of national medicine group; hydroxylamine hydrochloride standard was purchased from aladine, lot J2115489; indometacin intermediate 2 is self-made by the company, and batch number AP2449-2-9319-030-01.
2.2 chromatographic conditions
TABLE 1
2.3 preparation of solutions
2.3.1 derivatizing agents
Mixing acetone and methanol at a volume ratio of 1:1, placing into a volumetric flask, and shaking.
2.3.2 blank solution
Taking the derivatization reagent prepared by 2.3.1, namely the blank solution.
2.3.3 preparation of control solution
(1) Stock solution 1
Weighing 20 mg+/-2.0 mg hydroxylamine hydrochloride standard substance, precisely weighing, placing in a 50mL volumetric flask, dissolving with a mixed solution of acetone and methanol (1:1, v/v), diluting to a fixed volume to scale, and shaking uniformly to obtain the final product. Two parts are prepared by the same method. (hydroxylamine hydrochloride: 0.4 mg/mL).
(2) Stock solution 2
And precisely transferring 1mL of the stock solution into a volumetric flask with 1 to 10mL, diluting and fixing the volume to the scale by using a mixed solution (1:1, v/v) of acetone and methanol, and shaking uniformly to obtain the product. Two parts are prepared by the same method. (hydroxylamine hydrochloride: 40.0. Mu.g/mL)
(3) Reference substance solution
Accurately transferring 1mL of the stock solution into a volumetric flask with 2 to 10mL, diluting and fixing the volume to the scale by using a mixed solution of acetone and methanol (1:1, v/v), and shaking uniformly. (hydroxylamine hydrochloride: 4.0. Mu.g/mL, 100 ppm) two portions, labeled STD1 and STD2, respectively, were prepared in the same manner.
2.3.4 preparation of sample solutions
A400 mg.+ -.40 mg sample of indomethacin intermediate 2 was precisely weighed, placed in a 10mL volumetric flask, dissolved with a mixed solution of acetone and methanol (1:1, v/v), diluted to volume to scale, and shaken well. Obtaining the product. Two parts labeled TS1 and TS2, respectively, were prepared in the same manner. (Indometacin intermediate 2:40 mg/mL)
2.3.5 preparation of quantitative Limited and detection Limited solutions
(1) Quantitative limiting solution
Precisely transferring 5mL of STD1 solution into a 10mL volumetric flask, diluting and fixing the volume to the scale by using a mixed solution of acetone and methanol (1:1, v/v), and shaking uniformly. Labeled LOQ. (hydroxylamine hydrochloride: 2.0. Mu.g/mL, 50 ppm)
(1) Detection limiting solution
Precisely transferring 5mL of LOQ solution into a 10mL volumetric flask, diluting and fixing the volume to the scale by using a mixed solution of acetone and methanol (1:1, v/v), and shaking uniformly. Labeled LOD. (hydroxylamine hydrochloride: 1.0. Mu.g/mL, 25 ppm)
2.4 detection procedure
The sample was introduced according to the following sequence:
TABLE 2
| Solution name | Number of sample injection needles |
| Blank solution | ≥2* |
| LOQ solution | 1 |
| Standard solution (STD 1) | 4 |
| Standard solution (STD 2) | 2 |
| Sample solution 1 (TS 1) | 1 |
| Sample solution 2 (TS 2) | 1 |
| Blank solution | 1 |
| Standard solution (STD 1) | 2 |
Remarks: * The last needle is used as a system blank, and the other needles are used as balance systems;
ensuring that the system meets the following parameters:
(1) The blank has no interference (the area of the interference peak in the blank is less than 50% of the LOQ area at the peak position of hydroxylamine hydrochloride);
(2) The signal to noise ratio of LOQ is greater than 1;
(3) The response factor RSD of all STDs 1 is less than or equal to 10.0 percent;
(4) Recovery between STD1 and STD2 was 90.0-110.0%.
2.5 calculation
The content of hydroxylamine hydrochloride is calculated by an external standard method, and the formula is as follows:
wherein:
R i response factor of hydroxylamine hydrochloride in the sample solution;
rs represents the response factor mean of hydroxylamine hydrochloride in all STD1 solutions;
the content of Ps, hydroxylamine hydrochloride standard,%.
Remarks: response factor = peak area ≡concentration
TABLE 3 Table 3
| Name of the name | Retention time (min) | Relative retention time | Correction factor |
| Hydroxylamine hydrochloride | 4.2 | 1.0 | NA |
2.6 reporting results requirement
(1) Only hydroxylamine hydrochloride was integrated and the remaining peaks were not integrated.
(2) The average of the two samples is reported.
Reporting < 50ppm when the calculated result is less than 50ppm, and reporting the actual measurement value when the calculated result is more than 50 ppm.
3. Methodology investigation
3.1 investigation of specificity
Blank solution (acetone: methanol=1:1), quantitative limiting solution (2.0 μg/mL hydroxylamine hydrochloride), control solution (4.0 μg/mL hydroxylamine hydrochloride) and test solution (40 mg/mL indomethacin intermediate 2) were precisely aspirated, measured according to the chromatographic conditions under the "2.2" item, and the chromatograms were recorded. The results of the graphs shown in figures 1-2 and figures 4-9 show that the blank solution has no interference and good specificity at the peak position of hydroxylamine hydrochloride.
3.2 System applicability
In the same sequence, a hydroxylamine hydrochloride standard solution of "4 needles (STD 1) +2 needles (STD 2) +2 needles (STD 1)" was continuously introduced, and the measurement was performed under the chromatographic condition under the item "2.2". The criteria that demonstrate the systematic applicability of this method is that the RSD of all 6-needle STD1 response factors is less than 10% and the recovery of STD1 and STD2 is between 90-110%. The result shows that the RSD of the response factor of the STD1 of all control solutions is 1.74%, the recovery rate of the STD1 and the STD2 is 98.95%, and the system applicability is good.
Table 4 system suitability investigation
3.3 linear relationship investigation
Preparing 6 parts (2.0 mu g/mL,4.0 mu g/mL,6.0 mu g/mL,8.0 mu g/mL,20 mu g/mL and 40 mu g/mL) of hydroxylamine hydrochloride reference solution, injecting according to the chromatographic condition under the item "2.2", sequentially feeding two needles respectively, recording peak areas, taking the peak areas as an ordinate (Y), taking the concentration of the reference solution as an abscissa (X), drawing a regression curve by a least square method, recording a regression equation, and recording a correlation coefficient r 2 Sum of squares of residuals, require r 2 Greater than 0.997. The regression equation is shown in Table 6, the linear regression diagram is shown in FIG. 10, and the residual diagram of hydroxylamine hydrochloride is shown in FIG. 11.
TABLE 5 examination of the linear relationship of hydroxylamine hydrochloride
TABLE 6
| Name of the name | Linear range | Linear equation | Correlation coefficient r 2 |
| Hydroxylamine hydrochloride | 2.0-40μg/mL(LOQ-1000％) | y＝2.015x-0.285 | 1.000 |
3.5 repeatability test
6 parts of the indomethacin intermediate 2 solution with the concentration of 40mg/mL are prepared in parallel, the test solution is prepared in parallel under the condition of 2.3.4, the test is carried out according to the chromatographic condition under the condition of 2.2, the content of hydroxylamine hydrochloride in the test is tested, and the RSD between 6 parts of results is evaluated, wherein the requirement is less than 10%. RSD is 1.71%, which shows that the method has good repeatability and high precision.
TABLE 7
3.6 sample recovery test
15 parts of indomethacin intermediate 2 solution with the concentration of 40mg/mL is prepared, 4 levels of hydroxylamine hydrochloride (blank 3,2.0 mug/mL 3,4.0 mug/mL 3,8.0 mug/mL 3, 40 mug/mL 3) are added to the solution, a test sample solution is prepared according to the item "2.3.4", the ratio between an actual measurement value and an added value is calculated according to the chromatographic condition under the item "2.2", the recovery rate is calculated, and the recovery rate is required to be 75-125%. As a result, the LOQ concentration level, the 100% concentration level, the 200% concentration level and the 1000% concentration level are respectively 87.8% -88.5%, 86.6% -92.2%, 87.8% -91.3% and 89.0% -91.0% in the sample recovery rate intervals, so that the method is good in accuracy.
Table 8 hydroxylamine hydrochloride sample recovery test results (n=3)
3.7 quantitative limit and detection limit
6 parts of hydroxylamine hydrochloride solution having a concentration of 1.0. Mu.g/mL was prepared as a detection limit solution (LOD), 6 needles were continuously introduced, and the signal to noise ratio of the hydroxylamine hydrochloride peak was evaluated. A signal to noise ratio greater than 3 is required.
6 portions of hydroxylamine hydrochloride solution having a concentration of 2.0. Mu.g/mL was prepared as a limiting solution (LOQ), 6 needles were continuously introduced, and the signal to noise ratio of the hydroxylamine hydrochloride peak was evaluated. A signal to noise ratio greater than 10 is required.
The results show that the signal to noise ratio S/N of the detection limiting solution is between 10 and 15 (table 9), and the signal to noise ratio S/N of the quantitative limiting solution is between 47 and 54 (table 10), thereby meeting the requirements.
TABLE 9
Table 10
3.8 method verification results summarization
TABLE 11
4. Conclusion(s)
The method for detecting the content of the hydroxylamine hydrochloride in the bulk drug has the advantages of strong specificity, good repeatability, high precision, high accuracy, high sensitivity (up to 2.0 mug/mL) and wide linear range (2.0-40 mug/mL), can be applied to the production process and quality control of the bulk drug, and particularly can detect the residual hydroxylamine hydrochloride in the indomethacin intermediate 2.
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. The method for detecting the content of hydroxylamine hydrochloride in the bulk drug is characterized by adopting a derivatization method and a gas chromatography method to detect the content of the hydroxylamine hydrochloride, and comprises the following steps:
preparing a reference substance solution, preparing a sample solution and detecting by gas chromatography;
the derivatizing reagent is acetone and methanol.
2. The method according to claim 1, wherein the derivatizing agent, acetone and methanol are present in a volume ratio of 1:
(1-5)。
3. the method according to claim 2, wherein the derivatizing agent, acetone and methanol are present in a volume ratio of 1:1.
4. the method according to claim 1, wherein the preparation of the test solution comprises the steps of: and taking a sample to be detected, dissolving the sample by using a derivatization reagent, and uniformly mixing to obtain a sample solution.
5. The method of claim 1, wherein the preparation of the control solution comprises the steps of: dissolving hydroxylamine hydrochloride standard substance with derivatization reagent, and mixing to obtain reference substance solution.
6. The method according to claim 1, wherein in the gas chromatography, the gas chromatography detection is performed by eluting with a temperature programmed method, and the temperature programmed program is: the initial temperature is 60-70 ℃, the temperature is kept for 1-5min, then 15-30 ℃/min is raised to 160-180 ℃, and then 25-40 ℃/min is raised to 220-280 ℃ and the temperature is kept for 2-8min.
7. The method according to claim 1, wherein the gas chromatography is performed by split-feeding, and the split ratio is 4:1-8:1.
8. The method according to claim 1, wherein in the gas chromatography, the carrier gas is nitrogen or helium, and the carrier gas flow rate is 1.0-2.0 mL-min -1 ；
The sample injection amount is 1-5 mu L;
the chromatographic column stationary phase is polyethylene glycol;
the temperature of the sample inlet is 230-250 ℃, and the temperature of the detector is 225-245 ℃;
the type of detector is a FID detector;
the flow rate of the hydrogen is 36-50mL/min; the tail blowing flow is 10-50mL/min.
9. Use of the method of any one of claims 1-8 for detecting the content of hydroxylamine hydrochloride in a drug substance.
10. The use according to claim 9, wherein the drug substance is azilsartan, itopride hydrochloride, parecoxib or indomethacin intermediate 2.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410100936.5A CN117825573A (en) | 2024-01-24 | 2024-01-24 | Method for detecting content of hydroxylamine hydrochloride in bulk drug and application |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410100936.5A CN117825573A (en) | 2024-01-24 | 2024-01-24 | Method for detecting content of hydroxylamine hydrochloride in bulk drug and application |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN117825573A true CN117825573A (en) | 2024-04-05 |
Family
ID=90513550
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202410100936.5A Pending CN117825573A (en) | 2024-01-24 | 2024-01-24 | Method for detecting content of hydroxylamine hydrochloride in bulk drug and application |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117825573A (en) |
-
2024
- 2024-01-24 CN CN202410100936.5A patent/CN117825573A/en active Pending
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN111289676B (en) | Method for detecting residual tert-butylamine in terbutaline sulfate bulk drug | |
| CN109406690B (en) | Method for detecting related substances in chloral hydrate | |
| CN115166080B (en) | Method for detecting impurity A and impurity B in ifosfamide bulk drug | |
| CN114994212A (en) | High performance liquid chromatography detection method for hydroxylamine residue in medicine | |
| CN112710758A (en) | Method for detecting residual solvent in tapentadol hydrochloride raw material medicine | |
| CN117825573A (en) | Method for detecting content of hydroxylamine hydrochloride in bulk drug and application | |
| CN114264752B (en) | Detection method of methyl bromoacetate | |
| CN110376302B (en) | Method for detecting m-fluorobenzaldehyde and m-fluorobenzene cinnamaldehyde | |
| CN112710762A (en) | Method for measuring residual quantity of dimethyl sulfate | |
| CN111999400A (en) | Method for separating and determining impurities of bulk drugs of baricitinib by using HPLC (high performance liquid chromatography) | |
| CN111380992A (en) | Method for detecting residual quantity of organic solvent in formoterol bulk drug | |
| CN115825292B (en) | Method for detecting S-glycidol in glycidyl R-butyrate | |
| CN115436525B (en) | LLTS-M3 and detection method and application of related substances thereof | |
| CN117074578B (en) | LC-MS/MS quantitative detection method of 2- (methylamino) -ethanol | |
| CN112305100B (en) | Method for detecting content of genotoxic impurity benzyl bromide in medicine | |
| CN113777204B (en) | Detection method of p-hydroxyacetophenone related substances | |
| CN111175388B (en) | Method for determining DCC content in indapamide bulk drug | |
| CN117538457A (en) | Method for detecting 1, 4-butyrolactone in succinic anhydride | |
| WO2023065521A1 (en) | Gas chromatography-mass spectrometry combined method for determining genotoxic impurity 1,3-dichloro-2-propanol | |
| CN115754077A (en) | Method for determining related substances in pyrazinamide | |
| CN116879478A (en) | Headspace gas chromatography detection method for multiple residual solvents in amiodarone hydrochloride | |
| CN115754076A (en) | Gas chromatography for measuring related substances in 4-methylene piperidine hydrochloride | |
| CN116223667A (en) | Method for detecting genotoxic impurities in tadalafil | |
| CN117310015A (en) | Method for measuring etomidate content in electronic tobacco tar | |
| CN115932103A (en) | Method for detecting benzaldehyde in clindamycin phosphate injection |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |