CN106706785B - A method of using substance related in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece - Google Patents
A method of using substance related in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece Download PDFInfo
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- CN106706785B CN106706785B CN201611205823.3A CN201611205823A CN106706785B CN 106706785 B CN106706785 B CN 106706785B CN 201611205823 A CN201611205823 A CN 201611205823A CN 106706785 B CN106706785 B CN 106706785B
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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
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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
- G01N2030/022—Column chromatography characterised by the kind of separation mechanism
- G01N2030/027—Liquid chromatography
Abstract
The invention belongs to Pharmaceutical Analysis fields, and in particular to the analysis method of liquid chromatogram separation determination irbesartan and hydrochlorthiazide piece impurity.(1) chromatographic column, column temperature, Detection wavelength and mobile phase are first passed through to determine chromatographic condition；(2) preparation of test sample and reference substance solution；(3) measuring method then measures reference substance solution and test solution using accurate respectively, injects liquid chromatograph, records chromatogram, calculates by Self-control method and correction factor own control hair method；Test solution main peak is consistent with the retention time of reference substance solution main peak, and realizes the present invention.
Description
Technical field
The invention belongs to analytical chemistry fields, and in particular to a kind of to use high performance liquid chromatography detection Irbesartan esodrix
In relation to the method for substance in piperazine piece.
Background technique
Irbesartan and hydrochlorthiazide piece is the development of Sanofi company, trade name An Bonuo (COAPROVEL).In
It lists in Britain within 1998, and was successfully listed in 2004 in China.Since the Hypotensive Mechanism of two kinds of drugs is different, compound is formed
Synergistic effect is generated after preparation to be depressured and enhance antihypertensive effect, and its deficiency that can complement each other, it is long-term to reduce or offset single medicine
The adverse reaction that medication is likely to occur, to increase the compliance of patient's medication.According to document and USP standard, esodrix
Oneself in piperazine bulk pharmaceutical chemicals and tablet knows that impurity is chloro- 1,3 benzene disulfonic acid amide of 4- amino -6-, it is possible to produce catabolite have
Chlorothiazide.Oneself knows that impurity is since the physicochemical property difference of two kinds of main ingredients is larger for Irbesartan impurity A in Irbesartan Tablets
Two kinds of main ingredients can be made to efficiently separate with impurity, and can be saved analysis time, need to establish a kind of detection method and control it.
Currently, only describing the method for irbesartan and hydrochlorthiazide piece defects inspecting in United States Pharmacopeia forum, but in the method
Impurity essence sulfanilamide (SN) retention time excessively shift to an earlier date, and solvent peak exist interference, Irbesartan appearance time rearward, peak type compared with
Difference, the review time is too long, and Hydrochioro is not completely separated with its impurity chlorothiazide.Therefore, establish one it is accurate targetedly
Gradient elution program detection irbesartan and hydrochlorthiazide piece and its method of impurity have very important significance.
Summary of the invention
The purpose of the present invention is to provide a kind of using related in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece
The analysis method of substance guarantees the compound preparation to realize the separation and measurement of irbesartan and hydrochlorthiazide and its impurity
Purity realizes the quality control of its finished product.
The object of the present invention is achieved like this: a kind of using in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece
Method in relation to substance, the detection method include the following steps:
(1) chromatographic condition
Chromatographic column: cyano key and silicagel column
Mobile phase: 0.01mol/L phosphate buffer solution is as mobile phase A, the mixed solution conduct stream of methanol and acetonitrile
Dynamic phase B；
Detection wavelength: 220nm
Flow velocity: 1.0ml/min
Sampling volume: 10 μ L
Column temperature: 25 DEG C
Solvent: methanol, acetonitrile, acid water mixed solution
Gradient elution:
| Time (min) | Mobile phase A (%) | Mobile phase B (%) |
| 0 | 75 | 25 |
| 10 | 75 | 25 |
| 11 | 60 | 40 |
| 20 | 60 | 40 |
| 21 | 75 | 25 |
| 30 | 75 | 25 |
(2) test sample: accurately weighing test sample fine powder about Irbesartan 150mg, set in 100ml measuring bottle, solubilizer 30ml,
Ultrasound 10 minutes, lets cool, is diluted to scale with solvent, shakes up, considered；Precision measures subsequent filtrate 7.5ml, sets in 50ml measuring bottle,
It is diluted to scale with solvent, shakes up to obtain the final product；
(3) reference substance: reference substance solution (a): taking Irbesartan reference substance 15mg, accurately weighed, sets in 25ml measuring bottle, adds
Solvent 5ml ultrasound makes to dissolve for 2 minutes, is diluted to scale with solvent, shake up to get；
Reference substance solution (b): taking Hydrochioro reference substance 20mg, accurately weighed, sets in 200ml measuring bottle, solubilizer 10ml
Ultrasound make to dissolve within 2 minutes, be diluted to scale with solvent, shake up to get；
Reference substance solution (c): Irbesartan impurity A reference substance stock solution: Irbesartan impurity A reference substance 2.5mg is taken, is set
In 25ml measuring bottle, solubilizer ultrasound make within 2 minutes to dissolve and be diluted to scale, shake up to get；
Reference substance solution (d): smart sulfanilamide (SN) reference substance stock solution: smart sulfanilamide (SN) reference substance 2.5mg is taken, sets in 50ml measuring bottle, adds
Make to dissolve and be diluted to scale within solvent supersonic 2 minutes, shake up to get；
System suitability reference substance solution: precision measures above-mentioned reference substance solution stock solution (a), (b), (c), (d) respectively
4.2ml, 2.5ml, 0.5ml, 3.0ml are set in same 100ml measuring bottle, and solubilizer is diluted to scale, shake up to get；
(4) measuring method: taking 20 μ l of contrast solution to inject liquid chromatograph, adjusts detection sensitivity, makes Irbesartan peak
Height is about the 5% of full scale；It is accurate again to measure test solution and each 20 μ l of contrast solution, it is injected separately into liquid chromatograph, is remembered
Chromatogram is recorded to 2 times of Irbesartan peak retention time；The content of each impurity is calculated according to the following formula；Other single impurity peaks faces
Product is not greater than the 0.2% of the main peak area of contrast solution, and the sum of each impurity peak area is not greater than contrast solution main peak area
1.0%.
(5) calculation formula
The specification of the cyano key and silicagel column be 250 × 4.6mm, 5 μm；The cyano key and silicagel column are selected from ZORBAX
CN；The preparation method of the 0.01mol/L phosphate buffer is are as follows: weighs potassium dihydrogen phosphate 1.36g, is dissolved in water and constant volume
To 900mL, make to dissolve, add triethylamine 2ml, with phosphoric acid,diluted tune pH value to 3.2, is diluted with water to 1000ml；The Mobile phase B
For the mixed solution of methanol and acetonitrile, volume ratio is methanol: acetonitrile=10:15；In step (1), in the solvent, first
Alcohol: acetonitrile: the volume ratio of acid water is 30:45:25；The acid water is the aqueous solution that phosphorus acid for adjusting pH value is 2.0.
Detailed description of the invention
Fig. 1 Irbesartan UV scanning spectrogram
Fig. 2 Hydrochioro UV scanning spectrogram
The specificity spectrogram of Fig. 3 irbesartan and hydrochlorthiazide and its impurity.
Fig. 4 changes pH of cushioning fluid chromatogram
Fig. 5 difference liquidity ratio chromatogram
The chromatogram of Fig. 6 different manufacturers chromatographic column
Specific embodiment
It will be helpful to understand the present invention by following examples, but the contents of the present invention cannot be limited to.
Experimental example one:
Irbesartan and hydrochlorthiazide piece measuring method according to the invention, the self-control strategic point that measurement lot number is 20,131,104 one years
The acceleration samples of Bei Shatan hydrochlorothiazide tablets check the content of each impurity in relation to substance item.
It takes the sample lots 10 finely ground, weighs 150.23mg, set in 100ml measuring bottle, solubilizer 30ml, 10 points of ultrasound
Clock is let cool, and is diluted to scale with solvent, is shaken up, considered；Precision measures subsequent filtrate 7.5ml, sets in 50ml measuring bottle, dilute with solvent
It releases to scale, shakes up to obtain the final product.
Reference substance: reference substance solution (a): accurately weighing Irbesartan reference substance 15.14mg, accurately weighed, sets 25ml amount
In bottle, solubilizer 5ml ultrasound makes to dissolve for 2 minutes, is diluted to scale with solvent, shake up to get；
Reference substance solution (b): taking Hydrochioro reference substance 20.21mg, accurately weighed, sets in 200ml measuring bottle, solubilizer
10ml ultrasound make to dissolve within 2 minutes, be diluted to scale with solvent, shake up to get；
Reference substance solution (c): Irbesartan impurity A reference substance stock solution: taking Irbesartan impurity A reference substance 2.56mg,
Set in 25ml measuring bottle, solubilizer ultrasound make within 2 minutes to dissolve and be diluted to scale, shake up to get；
Reference substance solution (d): smart sulfanilamide (SN) reference substance stock solution: smart sulfanilamide (SN) reference substance 2.52mg is taken, sets in 50ml measuring bottle, adds
Make to dissolve and be diluted to scale within solvent supersonic 2 minutes, shake up to get；
System suitability reference substance solution: precision measures above-mentioned reference substance solution stock solution (a), (b), (c), (d) respectively
4.2ml, 2.5ml, 0.5ml, 3.0ml are set in same 100ml measuring bottle, and solubilizer is diluted to scale, shake up to get；
Chromatographic column selects Agilent cyano key and silicagel column, mobile phase: weighs potassium dihydrogen phosphate 1.36g, is dissolved in water simultaneously
It is settled to 900mL, makes to dissolve, adds triethylamine 2ml, with phosphoric acid,diluted tune pH value to 3.2, is diluted with water to 1000ml as mobile phase
A.The mixed solution of methanol 400ml, acetonitrile 600ml are measured as Mobile phase B；Detection wavelength: 220nm；Flow velocity: 1.0ml/min；
Sampling volume: 10 μ L；Column temperature: 25 DEG C；Solvent: the mixed solvent of methanol 300ml, acetonitrile 450ml, acid water 250ml are measured.
According to following gradient elution:
| Time (min) | Mobile phase A (%) | Mobile phase B (%) |
| 0 | 75 | 25 |
| 10 | 75 | 25 |
| 11 | 60 | 40 |
| 20 | 60 | 40 |
| 21 | 75 | 25 |
| 30 | 75 | 25 |
Measuring method: taking 20 μ l of contrast solution to inject liquid chromatograph, adjusts detection sensitivity, makes Irbesartan peak height about
It is the 5% of full scale；It is accurate again to measure test solution and each 20 μ l of contrast solution, it is injected separately into liquid chromatograph, records color
Spectrogram is to 2 times of Irbesartan peak retention time.The content of each impurity is calculated according to the following formula.Other single impurity peak areas are not
0.2% of the main peak area greater than contrast solution is obtained, the sum of each impurity peak area is not greater than contrast solution main peak area
1.0%.
Calculation formula
It is obtained according to calculation formula: smart sulfanilamide (SN): 0.50%, Irbesartan impurity A: 0.08%, single miscellaneous: 0.05%, it is total miscellaneous
0.65%.Experimental example two
The selection of 1.1 Detection wavelengths
Precision weighs Irbesartan, Irbesartan A reference substance in right amount with the dissolution of above-mentioned solvent and constant volume, is prepared into about 20 μ
The mixed reference substance solution of g/ml；Precision weighs Hydrochioro reference substance in right amount with solvent dissolution and constant volume, is prepared into 20 μ g/ml
The mixed reference substance solution of solution swept in 200-400nm wave-length coverage using solvent as blank according to ultraviolet spectrophotometry
It retouches, records ultra-violet absorption spectrum.
Shown by 1 Irbesartan of attached drawing and 2 Hydrochioro of attached drawing it is found that in 200-300nm wave-length coverage Hydrochioro
There is absorption maximum at 220nm (see appended drawing reference 1 in Fig. 2), 270nm and 312nm；Irbesartan and its impurity Irbesartan A
There is absorption maximum (see appended drawing reference 1 in Fig. 1) at 206nm, each acromion at 225nm and 242nm.Select the 220nm wave
A length of Detection wavelength for measuring the Compound Tablet Related substances separation.
The selection of 1.2 mobile phases
1.2.1 the selection of the water phase of mobile phase
Investigate influence of the various concentration potassium dihydrogen phosphate 0.01mol/L-0.2mol/L to main peak, as a result, it has been found that influence compared with
It is small, it is contemplated that high salt concentration be easy be precipitated, be easy be precipitated from mobile phase, it is possible to damage chromatographic column and instrument thus select compared with
The potassium dihydrogen phosphate of low concentration 0.01mol/L.
1.2.2 the selection of phosphate buffer pH
Investigate pH respectively 3.2,4.0,5.0 influence of the buffer to chromatographic column.It is shown in Table 1
Influence of the aqueous pH values to chromatographic column in 1 mobile phase of table
The results show that pH is the phosphate-buffered salt of 3.2-5.0, the tailing factor and theoretical tray of irbesartan and hydrochlorthiazide
Number meets the requirements, and when pH is 3.2, effect is best.Raising of the Hydrochioro summit with pH in chromatogram, appearance time
Shorten, but under gradient condition, pH value influences the appearance time of Irbesartan and little, only there is a small peak before Irbesartan,
Consider the separating degree and mobile phase ratio, appearance time of the two, final determination uses the buffer salt solution of pH3.2 as water phase.
1.2.3 in mobile phase organic phase selection
Through pharmacopeia and literature query it is found that physical and chemical properties of drugs difference is larger, grope to determine when organic phase is through overtesting
The volume ratio of methanol and acetonitrile obtains preferable peak type and separating property when being 40:60.It see the table below 2
Influence of organic Phase Proportion to chromatographic column in 2 mobile phase of table
1.2.4 the selection of mobile phase ratio
It can be seen that Irbesartan and Hydrochioro polarity spectrum are bigger from the physicochemical property of two kinds of drugs, work as organic phase
When ratio is higher, Hydrochioro appearance is too early, but if watr-proportion is turned up, the retention time of Irbesartan can postpone compared with
It is more, the retention time of Hydrochioro is influenced smaller.From save testing cost angle, organic phase (methanol: acetonitrile=
It is 10:15) relatively reasonable for 25:75 with water phase (0.01mol/L phosphate buffer solution) volume ratio, and in this, as mobile phase ratio
Example selection.
Embodiment 2
The foundation of methodology
2.1 specificity
Specificity solution is prepared to be prepared by the preparation method of reference substance (2) in this patent, and precision measures above-mentioned linear solvent
Each 20 μ l injects liquid chromatograph, records chromatogram, sees Fig. 3.Label is Irbesartan in attached drawing 3, is marked in attached drawing 3
1 is Irbesartan A, and label is smart sulfanilamide (SN) in attached drawing, and label is Hydrochioro in attached drawing.
The chromatographic isolation table of 3 specificity of table
Test result shows that solvent appearance is early, and related substance is not interfered to detect；Impurity mixing contrast solution and system are applicable in
Property testing liquid in see that attached drawing 3 can obtain, smart sulfanilamide (SN) (see appended drawing reference 3 in Fig. 3) and Hydrochioro (see appended drawing reference 4 in Fig. 3),
And Irbesartan impurity A (see appended drawing reference 1 in Fig. 3) and Irbesartan (see appended drawing reference 2 in attached drawing 3), between separating degree
1.5 are all larger than, separation is good；And there is absorption maximum in 220nm or so.As it can be seen that this method is used for irbesartan and hydrochlorthiazide
The Related substances separation of piece, specificity are good.
2.2 the range of linearity
Linear stock solution: precision measure smart sulfanilamide (SN) stock solution, Irbesartan impurity A stock solution, Irbesartan stock solution and
Each 5ml of Hydrochioro stock solution is set in same 50ml measuring bottle, and solubilizer is diluted to scale, shake up to get.
Precision measures linear stock solution 0.2,0.3,0.5ml, splits in 100ml measuring bottle, solubilizer is diluted to scale, shakes
It is even, 1., 2., 3. as linear solvent.Precision measures linear stock solution 0.1,0.2,0.4,0.5,0.7,1.0ml, splits 10ml
In measuring bottle, solubilizer is diluted to scale, shakes up, 4., 5., 6., 7., 8., 9. as linear solvent.
Precision measures above-mentioned each 20 μ l of linear solvent, injects liquid chromatograph, records chromatogram, is vertical sit with peak area
Mark, linear solvent concentration are abscissa, carry out linear regression with least square method.
Test result:
Smart sulfanilamide (SN) A=110204.74C+813.20r=0.9997 essence sulfanilamide (SN) is in the 0.007 μ g/ml concentration of μ g/ml~0.365
In range, concentration and peak area are in good linear relationship.
Irbesartan impurity A A=53428.39C-1570.86r=0.9999 Irbesartan impurity A is in 0.070 μ g/ml
In~1.396 μ g/ml concentration ranges, concentration and peak area are in good linear relationship.
Irbesartan A=56185.08C-1565.52r=0.9997 Irbesartan is in 0.135 μ of μ g/ml~4.503 g/ml
In concentration range, concentration and peak area are in good linear relationship.
Hydrochioro A=88153.33C+217.56r=0.9998 Hydrochioro is in 0.008 μ of μ g/ml~0.389 g/ml
In concentration range, concentration and peak area are in good linear relationship.
2.3 correction factors calculate
According to above-mentioned linear test as a result, with impurity essence sulfanilamide (SN), Irbesartan impurity A and principal component Hydrochioro, E Bei
The slope of husky smooth linear equation, calculate separately smart sulfanilamide (SN) relative to principal component Hydrochioro, Irbesartan impurity A relative to it is main at
The correction factor f of point Irbesartan, it is as a result as follows: calculation formula: the slope of f=impurity linear equation/principal component linear equation
Slope
4 irbesartan and hydrochlorthiazide piece Related substances separation correction factor calculated result of table
Test result shows that impurity essence sulfanilamide (SN) correction factor is 1.3, the correction factor of Irbesartan impurity A be 1.0 (
In 0.9~1.1 range), in Related substances separation, smart sulfanilamide (SN) uses the Self-control method of the correction up factor, and Irbesartan is miscellaneous
Matter A is using the Self-control method that correction factor is not added.
2.4 recovery test
Test solution: stock solution is made by the preparation method of this patent test sample (2).
Rate of recovery solution:
(1) test sample stock solution 1.5ml is taken, is set in 10ml measuring bottle, precision addition impurity stock solution 1.0ml is simultaneously dilute with solvent
It releases to scale, shakes up, as 100% rate of recovery solution, prepare 3 parts with method.
(2) test sample stock solution 1.5ml is taken, is set in 10ml measuring bottle, precision addition impurity stock solution 0.8ml is simultaneously dilute with solvent
It releases to scale, shakes up, as 80% rate of recovery solution, prepare 3 parts with method.
(3) test sample stock solution 1.5ml is taken, is set in 10ml measuring bottle, precision addition impurity stock solution 0.5ml is simultaneously dilute with solvent
It releases to scale, shakes up, as 50% rate of recovery solution, prepare 3 parts with method.
Precision measures above-mentioned impurity reference substance solution, test solution, each 20 μ l of rate of recovery solution, injects liquid chromatogram
Instrument records chromatogram, calculates the impurity rate of recovery, as a result see the table below:
5 irbesartan and hydrochlorthiazide piece Related substances separation recovery test result of table
Test result show the rate of recovery of smart sulfanilamide (SN) in 98.9~103.7% ranges, the recycling of Irbesartan impurity A
Rate is in 99.2~103.6% ranges；This product Related substances separation method accuracy is good.
2.5 quantitative limits, detection limit
Take smart sulfanilamide (SN), Irbesartan impurity A, Hydrochioro, Irbesartan reference substance each appropriate, accurately weighed, solubilizer
Ultrasound makes to dissolve for 5 minutes, then is gradually quantitatively diluted with solvent, when its signal-to-noise ratio S/N is about 10:1, respective concentration and injection instrument
The amount of device is quantitative limit, and by quantitative limit strength solution continuous sample introduction 6 times, calculates the relative standard deviation of gained peak area；
When its signal-to-noise ratio S/N is about 3:1, the amount of respective concentration and injection instrument is to detect limit.
6 irbesartan and hydrochlorthiazide piece Related substances separation quantitative limit test result of table
From above-mentioned test result: this method impurity essence sulfanilamide (SN), Irbesartan impurity A quantitative limit be respectively
0.12ng,1.34ng；Quantitative limit strength solution continuous sample introduction 6 times, the RSD of retention time are respectively 1.0%, 0.4%, respectively less than
2.0%；The RSD of peak area is respectively 4.7%, 4.7%, respectively less than 5.0%.
7 irbesartan and hydrochlorthiazide piece Related substances separation of table detection limit test result
From the above results: the detection of main ingredient Hydrochioro, Irbesartan and impurity essence sulfanilamide (SN), Irbesartan impurity A
Limit is respectively 0.04ng, 0.06ng, 0.31ng and 0.42ng
2.6 durability
2.6.1, pH of cushioning fluid
It is smart respectively according to preparation system employment and suitability test (E & ST) solution and test solution in this patent Related substance method
20 μ l of close measurement, are tested, map is shown in attached drawing 4 according to above-mentioned condition.1 in attached drawing 4 be 2.9 pH value buffer, in attached drawing 4
2 be 3.0 pH value buffer, 3 in attached drawing 4 be 3.2 pH value buffer.
To the inspection result of Drug-related Substances under 8 difference pH of table
Conclusion: by the test result of table 8 show the 1-3 pH of cushioning fluid marked from attached drawing 4 occur minor change when pair
Related substances separation result is without influence, good tolerance.
2.6.2 change mobile phase ratio: spectrogram is shown in attached drawing 5, and primary condition 1, changing liquidity ratio is respectively that 2-5 is shown in
Table 9
The mobile phase of label 1 is phosphate buffer (pH 3.0 ± 0.1)-methanol-acetonitrile (73:10:17) in attached drawing 5
The mobile phase of label 2 is phosphate buffer (pH 3.0 ± 0.1)-methanol-acetonitrile (74:9:17) in attached drawing 5
The mobile phase of label 3 is phosphate buffer (pH 3.0 ± 0.1)-methanol-acetonitrile (72:10:18) in attached drawing 5
The mobile phase of label 4 is phosphate buffer (pH 3.0 ± 0.1)-methanol-acetonitrile (72:11:17) in attached drawing 5
The mobile phase of label 5 is phosphate buffer (pH 3.0 ± 0.1)-methanol-acetonitrile (75:15:10) in attached drawing 5
Inspection result: 9 are shown in Table
The flowing Relative drug of 9 different proportion of table and its influence of impurity
The variation tendency of the mobile phase for the 1-5 for showing to mark from attached drawing 5 by the test result of table 9, it can be deduced that, change
Become mobile phase ratio to Related substances separation result without influence, good tolerance
2.6.3 different manufacturers chromatographic column:
The chromatographic column for choosing different manufacturers is as follows:
Label 1 is chromatographic column 1 in attached drawing 6: Agilent cyano key and silicagel column 4.6 × 250mm, 5 μm
P.N.880952-705S.N.USl0019652
Label 2 is chromatographic column 2 in attached drawing 6: Agilent cyano key and silicagel column 4.6 × 250mm, 5 μm
P.N.880952-705S.N.USl0019777
Label 3 is chromatographic column 3 in attached drawing 6: Waters cyano key and silicagel column 4.6 × 250mm, 5 μm of Part
Number:Ult5cn425Serial Number:251101594
The A that chromatographic column 1-3 is marked in attached drawing 6 is Irbesartan impurity A
10 different manufacturers chromatogram inspection result of table
From the experimental result of table 10: Irbesartan impurity A is overlapped with Irbesartan in chromatographic column 3, and separating degree is not inconsistent
Close regulation；2 system suitabilities of chromatographic column 1 and chromatographic column in attached drawing 6 are good, and Related substances separation result is consistent；Therefore
4.6 × 250mm of recommendation cyano key and silicagel column in Related substances separation, 5 μm.
Claims (3)
1. a kind of using the method in relation to substance in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece, which is characterized in that
The detection method includes the following steps:
(1) chromatographic condition
Chromatographic column: cyano key and silicagel column
Mobile phase: 0.01mol/L phosphate buffer solution is as mobile phase A；The mixed solution of methanol and acetonitrile is as mobile phase
B, volume ratio are methanol: acetonitrile=10:15；The preparation method of the 0.01mol/L phosphate buffer are as follows: weigh biphosphate
Potassium 1.36g is dissolved in water and is settled to 900mL, make to dissolve, add triethylamine 2ml, with phosphoric acid,diluted tune pH value to 3.2, is diluted with water
To 1000ml；
Detection wavelength: 220nm
Flow velocity: 1.0ml/min
Sampling volume: 10 μ L
Column temperature: 25 DEG C
Solvent: methanol, acetonitrile, acid water mixed solution
Gradient elution:
(2) test sample: accurately weighing test sample fine powder about Irbesartan 150mg, set in 100ml measuring bottle, solubilizer 30ml, ultrasound
It 10 minutes, lets cool, is diluted to scale with solvent, shakes up, considered；Precision measures subsequent filtrate 7.5ml, sets in 50ml measuring bottle, use is molten
Dilution agent shakes up to scale to obtain the final product；
(3) reference substance: reference substance solution (a): taking Irbesartan reference substance 15mg, accurately weighed, sets in 25ml measuring bottle, solubilizer
5ml ultrasound make to dissolve within 2 minutes, be diluted to scale with solvent, shake up to get；
Reference substance solution (b): taking Hydrochioro reference substance 20mg, accurately weighed, sets in 200ml measuring bottle, solubilizer 10ml ultrasound 2
Minute make to dissolve, be diluted to scale with solvent, shake up to get；
Reference substance solution (c): Irbesartan impurity A reference substance stock solution: Irbesartan impurity A reference substance 2.5mg is taken, 25ml is set
In measuring bottle, solubilizer ultrasound make within 2 minutes to dissolve and be diluted to scale, shake up to get；
Reference substance solution (d): smart sulfanilamide (SN) reference substance stock solution: smart sulfanilamide (SN) reference substance 2.5mg is taken, is set in 50ml measuring bottle, solubilizer
Ultrasound make within 2 minutes to dissolve and be diluted to scale, shake up to get；
Reference substance solution (e): chlorothiazide reference substance stock solution: chlorothiazide reference substance 2.5mg is taken, is set in 50ml measuring bottle, solubilizer
Ultrasound make within 2 minutes to dissolve and be diluted to scale, shake up to get；
System suitability reference substance solution: precision measures above-mentioned reference substance solution stock solution (a), (b), (c), (d), (e) respectively
4.2ml, 2.5ml, 0.5ml, 3.0ml, 1.0ml are set in same 100ml measuring bottle, and solubilizer is diluted to scale, shake up to get；
(4) measuring method: taking 20 μ l of contrast solution to inject liquid chromatograph, adjusts detection sensitivity, makes Irbesartan peak height about
It is the 5% of full scale；It is accurate again to measure test solution and each 20 μ l of contrast solution, it is injected separately into liquid chromatograph, records color
Spectrogram is to 2 times of Irbesartan peak retention time；The content of each impurity is calculated according to the following formula；Other single impurity peak areas are not
0.2% of the main peak area greater than contrast solution is obtained, the sum of each impurity peak area is not greater than contrast solution main peak area
1.0%；
(5) calculation formula
2. according to claim 1 a kind of using substance related in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece
Method, which is characterized in that the specification of the cyano key and silicagel column be 250 × 4.6mm, 5 μm；The cyano key and silicagel column
Selected from ZORBAX CN.
3. according to claim 1 a kind of using substance related in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece
Method, which is characterized in that in step (1), in the solvent, methanol: acetonitrile: the volume ratio of acid water be 30:45:25；
The acid water is the aqueous solution that the pH value that phosphoric acid is adjusted is 2.0.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201611205823.3A CN106706785B (en) | 2016-12-23 | 2016-12-23 | A method of using substance related in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece |
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| CN201611205823.3A CN106706785B (en) | 2016-12-23 | 2016-12-23 | A method of using substance related in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece |
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| CN106706785B true CN106706785B (en) | 2019-02-26 |
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| CN111175385B (en) * | 2018-11-12 | 2023-02-07 | 珠海润都制药股份有限公司 | Irbesartan isomer testing method in irbesartan |
| CN110988180A (en) * | 2019-12-19 | 2020-04-10 | 山东达因海洋生物制药股份有限公司 | Method for analyzing related substances of esomeprazole magnesium based on hybrid mass spectrometry |
| CN113030352B (en) * | 2019-12-25 | 2024-03-22 | 上海奥博生物医药股份有限公司 | Determination and analysis method for NMBA content in irbesartan |
| CN111220730A (en) * | 2020-01-19 | 2020-06-02 | 合肥科颖医药科技有限公司 | Analysis method of related substances in irbesartan and hydrochlorothiazide compound preparation |
| CN113686977B (en) * | 2020-05-18 | 2023-08-08 | 武汉中博绿亚生物科技有限公司 | Method for measuring related substances in compound fenbendazole preparation |
| CN112229923B (en) * | 2020-09-30 | 2022-11-11 | 东北制药集团股份有限公司 | Method for detecting 15-ketone and related substances thereof by adopting high performance liquid chromatography |
| CN114778711A (en) * | 2022-03-14 | 2022-07-22 | 江苏金申医药科技有限公司 | Method for analyzing related substances of sulfadoxine |
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| CN1203527A (en) * | 1995-11-03 | 1998-12-30 | 萨诺费公司 | Stable freeze-dried pharmaceutical formulation |
| CN1732953A (en) * | 2005-09-02 | 2006-02-15 | 姚俊华 | Dispersible tablet for treating hypertension |
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