CN106706785A - Method for detecting related substances in irbesartan hydrochlorothiazide tablets by adopting high performance liquid chromatography - Google Patents
Method for detecting related substances in irbesartan hydrochlorothiazide tablets by adopting high performance liquid chromatography Download PDFInfo
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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- 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
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Abstract
The invention belongs to the field of pharmaceutical analysis and particularly relates to a method for detecting related substances in irbesartan hydrochlorothiazide tablets by adopting high performance liquid chromatography. The method comprises the steps that 1, chromatographic conditions are firstly determined through a chromatographic column, column temperature, detection wave length and a mobile phase; 2, a test sample solution and a reference substance solution are prepared; 3, a determination method is adopted to perform precision determination of the test sample solution and the reference substance solution respectively, the solutions are injected into a liquid chromatograph, a chromatogram is recorded, and calculation is performed according to a self-contrasting method and a correction factor self-contrasting method; a main peak of the test sample solution and a main peak of the reference substance solution are consistent in retention time, and accordingly the method is achieved.
Description
Technical field
The invention belongs to analytical chemistry field, and in particular to one kind uses high performance liquid chromatography detection Irbesartan esodrix
About the method for material in piperazine piece.
Background technology
Irbesartan and hydrochlorthiazide piece is developed for Sanofi companies, trade name An Bonuo (COAPROVEL).In
Listed in Britain within 1998, and successfully listed in China in 2004.Because two kinds of Hypotensive Mechanisms of medicine are different, compound is constituted
Synergy is produced to be depressured and strengthen antihypertensive effect after preparation, and its deficiency that can complement each other, reduce or offset single medicine long-term
The adverse reaction that medication is likely to occur, so as to increase the compliance of patient's medication.According to document and USP standard, esodrix
Oneself in piperazine bulk drug and tablet knows that impurity is the benzene disulfonic acid amides of 4- amino -6- chloro- 1,3, it is possible to produce catabolite have
Chlorothiazide.Oneself knows that impurity, for Irbesartan impurity A, because the physicochemical property difference of two kinds of main ingredients is larger, is in Irbesartan Tablets
Two kinds of main ingredients can be made to be efficiently separated with impurity, can be saved analysis time again, needed badly and set up a kind of detection method and control it.
At present, the method for irbesartan and hydrochlorthiazide piece defects inspecting is only described in American 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 long, and Hydrochioro is not completely separated with its impurity chlorothiazide.Therefore, set up one it is accurate targetedly
Gradient elution program detection irbesartan and hydrochlorthiazide piece and its method for impurity have very important significance.
The content of the invention
It is an object of the invention to provide one kind using relevant in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece
The analysis method of material, so as to realize separation and the measure of irbesartan and hydrochlorthiazide and its impurity, it is ensured that the compound preparation
Purity, realizes the quality control of its finished product.
The object of the present invention is achieved like this：One kind is using in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece
About the method for material, the detection method comprises the following steps：
(1) chromatographic condition
Chromatographic column：Cyano group key and silicagel column
Mobile phase：Used as mobile phase A, the mixed solution of methyl alcohol and acetonitrile is used as stream for 0.01mol/L PBSs
Dynamic phase B；
Detection wavelength：220nm
Flow velocity：1.0ml/min
Sampling volume：10μL
Column temperature：25℃
Solvent：Methyl alcohol, acetonitrile, the mixed solution of acid water
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：Test sample fine powder about Irbesartan 150mg accurately is weighed, in putting 100ml measuring bottles, solubilizer 30ml,
Ultrasound 10 minutes, is let cool, and scale is diluted to solvent, is shaken up, and is considered；Precision measures subsequent filtrate 7.5ml, in putting 50ml measuring bottles,
Scale is diluted to solvent, is shaken up and is obtained final product；
(3) reference substance：Reference substance solution (a)：Irbesartan reference substance 15mg is taken, it is accurately weighed, in putting 25ml measuring bottles, plus
Solvent 5ml ultrasounds make dissolving in 2 minutes, and scale is diluted to solvent, shake up, and obtain final product；
Reference substance solution (b)：Hydrochioro reference substance 20mg is taken, it is accurately weighed, in putting 200ml measuring bottles, solubilizer 10ml
Ultrasound makes dissolving in 2 minutes, and scale is diluted to solvent, shakes up, and obtains final product；
Reference substance solution (c)：Irbesartan impurity A reference substance stock solution：Irbesartan impurity A reference substance 2.5mg is taken, is put
In 25ml measuring bottles, solubilizer ultrasound makes to dissolve and be diluted to scale for 2 minutes, shakes up, and obtains final product；
Reference substance solution (d)：Smart sulfanilamide (SN) reference substance stock solution：Smart sulfanilamide (SN) reference substance 2.5mg is taken, in putting 50ml measuring bottles, plus
Solvent supersonic makes to dissolve and be diluted to scale for 2 minutes, shakes up, and obtains final product；
System suitability reference substance solution：Precision measures above-mentioned reference substance solution storing solution (a), (b), (c), (d) respectively
4.2ml, 2.5ml, 0.5ml, 3.0ml are put in same 100ml measuring bottles, and solubilizer is diluted to scale, shakes up, and obtains final product；
(4) assay method:The μ l of contrast solution 20 injection liquid chromatographs are taken, detection sensitivity is adjusted, makes Irbesartan peak
Height is about the 5% of full scale；Precision measures need testing solution and each 20 μ l of contrast solution again, is injected separately into liquid chromatograph, remembers
Record chromatogram is to 2 times of Irbesartan peak retention time；The content of each impurity is calculated by following equation；Other single impurity peaks faces
Product cannot be greater than contrast solution main peak area 0.2%, each impurity peak area and cannot be greater than contrast solution main peak area
1.0%.
(5) computing formula
The specification of the cyano group key and silicagel column is 250 × 4.6mm, 5 μm；The cyano group key and silicagel column are selected from ZORBAX
CN；The compound method of the 0.01mol/L phosphate buffers for：Potassium dihydrogen phosphate 1.36g is weighed, is dissolved in water and constant volume
To 900mL, make dissolving, plus triethylamine 2ml, adjust pH value to 3.2 with phosphoric acid,diluted, be diluted with water to 1000ml；Described Mobile phase B
It is methyl alcohol and the mixed solution of acetonitrile, its volume ratio is methyl alcohol：Acetonitrile=10:15；In step (1), in the solvent, first
Alcohol：Acetonitrile：The volume ratio of acid water is 30:45:25；Described acid water is the aqueous solution that phosphorus acid for adjusting pH value is 2.0.
Brief description of the drawings
Fig. 1 Irbesartan UV scanning spectrograms
Fig. 2 Hydrochioro UV scanning spectrograms
The specificity spectrogram of Fig. 3 irbesartan and hydrochlorthiazides and its impurity.
Fig. 4 changes pH of cushioning fluid chromatogram
Fig. 5 difference liquidity ratio chromatograms
The chromatogram of Fig. 6 different manufacturers chromatographic columns
Specific embodiment
Will be helpful to understand the present invention by following examples, but present disclosure can not be limited to.
Experimental example one：
According to irbesartan and hydrochlorthiazide piece assay method of the invention, the self-control strategic point that lot number is 20,131,104 one years is determined
The acceleration samples of Bei Shatan hydrochlorothiazide tablets, check the content of each impurity in about material.
Take the sample lots 10 finely ground, weigh 150.23mg, put in 100ml measuring bottles, solubilizer 30ml is ultrasonic 10 points
Clock, is let cool, and scale is diluted to solvent, is shaken up, and is considered；Precision measures subsequent filtrate 7.5ml, dilute with solvent in putting 50ml measuring bottles
Release to scale, shake up and obtain final product.
Reference substance：Reference substance solution (a)：It is accurate to weigh Irbesartan reference substance 15.14mg, it is accurately weighed, put 25ml amounts
In bottle, solubilizer 5ml ultrasounds make dissolving in 2 minutes, and scale is diluted to solvent, shake up, and obtain final product；
Reference substance solution (b)：Hydrochioro reference substance 20.21mg is taken, it is accurately weighed, in putting 200ml measuring bottles, solubilizer
10ml ultrasounds make dissolving in 2 minutes, and scale is diluted to solvent, shake up, and obtain final product；
Reference substance solution (c)：Irbesartan impurity A reference substance stock solution：Irbesartan impurity A reference substance 2.56mg is taken,
Put in 25ml measuring bottles, solubilizer ultrasound makes to dissolve and be diluted to scale for 2 minutes, shakes up, and obtains final product；
Reference substance solution (d)：Smart sulfanilamide (SN) reference substance stock solution：Smart sulfanilamide (SN) reference substance 2.52mg is taken, in putting 50ml measuring bottles, plus
Solvent supersonic makes to dissolve and be diluted to scale for 2 minutes, shakes up, and obtains final product；
System suitability reference substance solution：Precision measures above-mentioned reference substance solution storing solution (a), (b), (c), (d) respectively
4.2ml, 2.5ml, 0.5ml, 3.0ml, put in same 100ml measuring bottles, and solubilizer is diluted to scale, shakes up, and obtains final product；
Chromatographic column selects Agilent cyano group key and silicagel column, mobile phase：Potassium dihydrogen phosphate 1.36g is weighed, is dissolved in water simultaneously
900mL is settled to, makes dissolving, plus triethylamine 2ml, adjust pH value to 3.2 with phosphoric acid,diluted, be diluted with water to 1000ml as mobile phase
A.Methyl alcohol 400ml, the mixed solution of acetonitrile 600ml are measured as Mobile phase B；Detection wavelength：220nm；Flow velocity：1.0ml/min；
Sampling volume：10μL；Column temperature：25℃；Solvent：Measure methyl alcohol 300ml, acetonitrile 450ml, the mixed solvent of acid water 250ml.
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 |
Assay method:The μ l of contrast solution 20 injection liquid chromatographs are taken, detection sensitivity is adjusted, makes Irbesartan peak height about
It is the 5% of full scale；Precision measures need testing solution and each 20 μ l of contrast solution again, 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 by following equation.Other single impurity peak areas are not
Must be more than contrast solution main peak area 0.2%, each impurity peak area and cannot be greater than contrast solution main peak area
1.0%.
Computing formula
Drawn according to computing formula：Smart sulfanilamide (SN)：0.50%, Irbesartan impurity A：0.08%, it is 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 substances and is dissolved and constant volume with above-mentioned solvent in right amount, is prepared into about 20 μ
The mixed reference substance solution of g/ml；Precision weighs the appropriate solvent of Hydrochioro reference substance and dissolves and constant volume, is prepared into 20 μ g/ml
Solution mixed reference substance solution, with solvent as blank, swept in 200-400nm wave-length coverages according to ultraviolet spectrophotometry
Retouch, record ultra-violet absorption spectrum.
Shown from the Irbesartan of accompanying drawing 1 and the Hydrochioro of accompanying drawing 2, the Hydrochioro in 200-300nm wave-length coverages
There is absorption maximum at 220nm (reference 1 in see Fig. 2), 270nm and 312nm；Irbesartan and its impurity Irbesartan A
(reference 1 in see Fig. 1) has absorption maximum at 206nm, each acromion at 225nm and 242nm.From the 220nm ripples
A length of Detection wavelength for determining 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 influences of the various concentrations potassium dihydrogen phosphate 0.01mol/L-0.2mol/L to main peak, as a result find influence compared with
It is small, it is contemplated that high salt concentration is easily separated out, easily from mobile phase separate out, it is possible to damage chromatographic column and instrument thus from 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 solution to chromatographic column.It is shown in Table 1
Influence of the aqueous pH values to chromatographic column in the mobile phase of table 1
Result shows 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.Hydrochioro summit in chromatogram is with the rising of pH, appearance time
Shorten, but under gradient condition, appearance time influence of the pH value on Irbesartan is simultaneously little, simply there is a small peak before Irbesartan,
Consider both separating degrees and mobile phase ratio, appearance time, the final buffer salt solution for determining to use pH3.2 is used as water phase.
1.2.3 in mobile phase organic phase selection
Understand that physical and chemical properties of drugs difference is larger through pharmacopeia and literature query, grope to determine when organic phase is through overtesting
Methyl alcohol is 40 with the volume ratio of acetonitrile:Preferable peak type and separating property are obtained when 60.See the table below 2
Organic Phase Proportion is checked colors and composes the influence of post in the mobile phase of table 2
1.2.4 the selection of mobile phase ratio
From the physicochemical property of two kinds of medicines can be seen that Irbesartan with Hydrochioro polarity spectrum than larger, work as organic phase
When ratio is higher, Hydrochioro appearance too early, but if watr-proportion is heightened, the retention time of Irbesartan can postpone compared with
Many, the retention time influence on Hydrochioro is smaller.From the angle for saving testing cost, organic phase (methyl alcohol：Acetonitrile=
10:15) it is 25 with water phase (0.01mol/L PBSs) volume ratio:75 is relatively reasonable, and in this, as mobile phase ratio
Example selection.
Embodiment 2
The foundation of methodology
2.1 specificities
Specificity solution prepares the compound method preparation for pressing reference substance (2) in this patent, and precision measures above-mentioned linear solvent
Each 20 μ l, inject liquid chromatograph, record chromatogram, see Fig. 3.What mark was in accompanying drawing 3 is Irbesartan, is marked in accompanying drawing 3
1 is Irbesartan A, and what mark was in accompanying drawing is smart sulfanilamide (SN), and what mark was in accompanying drawing is Hydrochioro.
The chromatographic isolation table of the specificity of table 3
Result of the test shows that solvent appearance morning does not disturb relevant material to detect；Impurity mixing contrast solution is applicable with system
Property testing liquid in see that accompanying drawing 3 can be obtained, smart sulfanilamide (SN) (see reference 3 in Fig. 3) and Hydrochioro (see reference 4 in Fig. 3),
And Irbesartan impurity A (see reference 1 in Fig. 3) and Irbesartan (see reference 2 in accompanying drawing 3), separating degree between it
1.5 are all higher than, are separated good；And have absorption maximum in 220nm or so.It can be seen that, this method is used for irbesartan and hydrochlorthiazide
The Related substances separation of piece, specificity is good.
2.2 ranges 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, puts in same 50ml measuring bottles, and solubilizer is diluted to scale, shakes up, and obtains final product.
Precision measures linear stock solution 0.2,0.3,0.5ml, splits in 100ml measuring bottles, and solubilizer is diluted to scale, shakes
It is even, as linear solvent 1., 2., 3..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, as linear solvent 4., 5., 6., 7., 8., 9..
Precision measures each 20 μ l of above-mentioned linear solvent, injects liquid chromatograph, records chromatogram, is vertical seat with peak area
Mark, linear solvent concentration is abscissa, and linear regression is carried out with least square method.
Result of the test：
Smart sulfanilamide (SN) A=110204.74C+813.20r=0.9997 essences sulfanilamide (SN) is in 0.007 μ g/ml~0.365 μ g/ml concentration
In the range of, concentration is in good linear relationship with peak area.
Irbesartan impurity A A=53428.39C-1570.86r=0.9999 Irbesartans impurity A is in 0.070 μ g/ml
In~1.396 μ g/ml concentration ranges, concentration is in good linear relationship with peak area.
Irbesartan A=56185.08C-1565.52r=0.9997 Irbesartans are in 0.135 μ g/ml~4.503 μ g/ml
In concentration range, concentration is in good linear relationship with peak area.
Hydrochioro A=88153.33C+217.56r=0.9998 Hydrochioros are in 0.008 μ g/ml~0.389 μ g/ml
In concentration range, concentration is in good linear relationship with peak area.
2.3 correction factors are calculated
According to above-mentioned linear test result, with impurity essence sulfanilamide (SN), Irbesartan impurity A and principal component Hydrochioro, E Bei
The slope of husky smooth linear equation, calculate respectively smart sulfanilamide (SN) relative to principal component Hydrochioro, Irbesartan impurity A relative to it is main into
The correction factor f of point Irbesartan, it is as a result as follows：Computing formula：The slope of f=impurity linear equations/principal component linear equation
Slope
The irbesartan and hydrochlorthiazide piece Related substances separation correction factor result of calculation of table 4
Result of the test shows, impurity essence sulfanilamide (SN) correction factor is 1.3, the correction factor of Irbesartan impurity A for 1.0 (
In the range of 0.9~1.1), 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 for being not added with correction factor.
2.4 recovery tests
Need testing solution：Storing solution is made by the compound method of this patent test sample (2).
Rate of recovery solution：
(1) test sample stock solution 1.5ml is taken, is put in 10ml measuring bottles, precision addition impurity stock solution 1.0ml is simultaneously dilute with solvent
Release to scale, shake up, as 100% rate of recovery solution, 3 parts are prepared with method.
(2) test sample stock solution 1.5ml is taken, is put in 10ml measuring bottles, precision addition impurity stock solution 0.8ml is simultaneously dilute with solvent
Release to scale, shake up, as 80% rate of recovery solution, 3 parts are prepared with method.
(3) test sample stock solution 1.5ml is taken, is put in 10ml measuring bottles, precision addition impurity stock solution 0.5ml is simultaneously dilute with solvent
Release to scale, shake up, as 50% rate of recovery solution, 3 parts are prepared with method.
Precision measures each 20 μ l of above-mentioned impurity reference substance solution, need testing solution, rate of recovery solution, injects liquid chromatogram
Instrument, records chromatogram, calculates the impurity rate of recovery, as a result see the table below：
The irbesartan and hydrochlorthiazide piece Related substances separation recovery test result of table 5
Result of the test shows, the rate of recovery of smart sulfanilamide (SN) in the range of 98.9~103.7%, the recovery of Irbesartan impurity A
Rate is in the range of 99.2~103.6%；This product Related substances separation method degree of accuracy is good.
2.5 quantitative limits, test limit
Take smart sulfanilamide (SN), Irbesartan impurity A, Hydrochioro, Irbesartan reference substance each appropriate, accurately weighed, solubilizer
Ultrasound makes dissolving for 5 minutes, then is progressively quantitatively diluted with solvent, when its signal to noise ratio S/N is about 10:When 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:When 1, the amount of respective concentration and injection instrument is test limit.
The irbesartan and hydrochlorthiazide piece Related substances separation quantitative limit result of the test of table 6
From above-mentioned result of the test：This method impurity essence sulfanilamide (SN), the quantitative limit of Irbesartan impurity A are respectively
0.12ng、1.34ng；Quantitative limit strength solution continuous sample introduction 6 times, the RSD of retention time is 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%.
The irbesartan and hydrochlorthiazide piece Related substances separation test limit result of the test of table 7
From the above results：Main ingredient Hydrochioro, Irbesartan and impurity essence sulfanilamide (SN), the detection of Irbesartan impurity A
Limit is respectively 0.04ng, 0.06ng, 0.31ng and 0.42ng
2.6 durabilities
2.6.1, pH of cushioning fluid
It is smart respectively according to preparation system employment and suitability test (E ＆ ST) solution and need testing solution in this patent Related substance method
It is close to measure 20 μ l, to be tested according to above-mentioned condition, collection of illustrative plates is shown in accompanying drawing 4.In accompanying drawing 41 is 2.9 pH value buffer solution, in accompanying drawing 4
2 for 3.0 pH value buffer solution, in accompanying drawing 43 for 3.2 pH value buffer solution.
To the inspection result of Drug-related Substances under the difference of table 8 pH
Conclusion：Shown by the result of the test of table 8, from accompanying drawing 4 mark 1-3 pH of cushioning fluid occur minor variations when pair
Related substances separation result is without influence, good tolerance.
2.6.2 mobile phase ratio is changed：Spectrogram is shown in accompanying drawing 5, and primary condition is 1, and change liquidity ratio is respectively 2-5 and sees
Table 9
The mobile phase of mark 1 is phosphate buffer (pH 3.0 ± 0.1)-methanol-acetonitrile (73 in accompanying drawing 5：10：17)
The mobile phase of mark 2 is phosphate buffer (pH 3.0 ± 0.1)-methanol-acetonitrile (74 in accompanying drawing 5：9：17)
The mobile phase of mark 3 is phosphate buffer (pH 3.0 ± 0.1)-methanol-acetonitrile (72 in accompanying drawing 5：10：18)
The mobile phase of mark 4 is phosphate buffer (pH 3.0 ± 0.1)-methanol-acetonitrile (72 in accompanying drawing 5：11：17)
The mobile phase of mark 5 is phosphate buffer (pH 3.0 ± 0.1)-methanol-acetonitrile (75 in accompanying drawing 5：15：10)
Inspection result：It is shown in Table 9
The flowing Relative drug of the different proportion of table 9 and its influence of impurity
Shown by the result of the test of table 9, the variation tendency of the mobile phase of the 1-5 marked from accompanying drawing 5, 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：
Mark 1 is chromatographic column 1 in accompanying drawing 6:Agilent cyano group key and silicagel column 4.6 × 250mm, 5 μm
P.N.880952-705S.N.USl0019652
Mark 2 is chromatographic column 2 in accompanying drawing 6:Agilent cyano group key and silicagel column 4.6 × 250mm, 5 μm
P.N.880952-705S.N.USl0019777
Mark 3 is chromatographic column 3 in accompanying drawing 6:This cyano group key and silicagel column of water 4.6 × 250mm, 5 μm of Part
Number:Ult5cn425Serial Number:251101594
The A that chromatographic column 1-3 is marked in accompanying drawing 6 is Irbesartan impurity A
The different manufacturers chromatogram inspection result of table 10
From the experimental result of table 10：Irbesartan impurity A overlaps 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 accompanying drawing 6 are good, and Related substances separation result is consistent；Therefore
4.6 × 250mm of recommendation cyano group key and silicagel column, 5 μm in Related substances separation.
Claims (5)
1. about the method for material in a kind of use high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece, it is characterised in that
The detection method comprises the following steps：
(1) chromatographic condition
Chromatographic column：Cyano group key and silicagel column
Mobile phase：Used as mobile phase A, the mixed solution of methyl alcohol and acetonitrile is used as mobile phase for 0.01mol/L PBSs
B；
Detection wavelength：220nm
Flow velocity：1.0ml/min
Sampling volume：10μL
Column temperature：25℃
Solvent：Methyl alcohol, acetonitrile, the mixed solution of acid water
Gradient elution：
(2) test sample：Test sample fine powder about Irbesartan 150mg accurately is weighed, is put in 100ml measuring bottles, solubilizer 30ml is ultrasonic
10 minutes, let cool, scale is diluted to solvent, shake up, considered；Precision measures subsequent filtrate 7.5ml, in putting 50ml measuring bottles, with molten
Dilution agent shakes up and obtains final product to scale；
(3) reference substance：Reference substance solution (a)：Irbesartan reference substance 15mg is taken, it is accurately weighed, in putting 25ml measuring bottles, solubilizer
5ml ultrasounds make dissolving in 2 minutes, and scale is diluted to solvent, shake up, and obtain final product；
Reference substance solution (b)：Hydrochioro reference substance 20mg is taken, it is accurately weighed, in putting 200ml measuring bottles, solubilizer 10ml ultrasounds 2
Minute makes dissolving, and scale is diluted to solvent, shakes up, and obtains final product；
Reference substance solution (c)：Irbesartan impurity A reference substance stock solution：Irbesartan impurity A reference substance 2.5mg is taken, 25ml is put
In measuring bottle, solubilizer ultrasound makes to dissolve and be diluted to scale for 2 minutes, shakes up, and obtains final product；
Reference substance solution (d)：Smart sulfanilamide (SN) reference substance stock solution：Smart sulfanilamide (SN) reference substance 2.5mg is taken, in putting 50ml measuring bottles, solubilizer
Ultrasound makes to dissolve and be diluted to scale for 2 minutes, shakes up, and obtains final product；
Reference substance solution (e)：Chlorothiazide reference substance storing solution：Chlorothiazide reference substance 2.5mg is taken, in putting 50ml measuring bottles, solubilizer
Ultrasound makes to dissolve and be diluted to scale for 2 minutes, shakes up, and obtains final product；
System suitability reference substance solution：Precision measures above-mentioned reference substance solution storing solution (a), (b), (c), (d), (e) respectively
4.2ml, 2.5ml, 0.5ml, 3.0ml, 1.0ml, put in same 100ml measuring bottles, and solubilizer is diluted to scale, shakes up, and obtains final product；
(4) assay method:The μ l of contrast solution 20 injection liquid chromatographs are taken, detection sensitivity is adjusted, makes Irbesartan peak height about
It is the 5% of full scale；Precision measures need testing solution and each 20 μ l of contrast solution again, 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 by following equation；Other single impurity peak areas are not
Must be more than contrast solution main peak area 0.2%, each impurity peak area and cannot be greater than contrast solution main peak area
1.0%；
(5) computing formula
2. according to claim 1 a kind of using relevant material in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece
Method, it is characterised in that the specification of the cyano group key and silicagel column be 250 × 4.6mm, 5 μm；The cyano group key and silicagel column
Selected from ZORBAX CN.
3. according to claim 1 a kind of using relevant material in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece
Method, it is characterised in that the compound method of the 0.01mol/L phosphate buffers for：Weigh potassium dihydrogen phosphate
1.36g, is dissolved in water and is settled to 900mL, makes dissolving, plus triethylamine 2ml, adjusts pH value to 3.2 with phosphoric acid,diluted, is diluted with water to
1000ml。
4. according to claim 1 a kind of using relevant material in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece
Method, it is characterised in that described Mobile phase B is the mixed solution of methyl alcohol and acetonitrile, and volume ratio is methyl alcohol：Acetonitrile=10:
15。
5. according to claim 1 a kind of using relevant material in high performance liquid chromatography detection irbesartan and hydrochlorthiazide piece
Method, it is characterised in that in step (1), in the solvent, methyl alcohol：Acetonitrile：The volume ratio of acid water is 30:45:25；
Described acid water is the aqueous solution that the pH value that phosphoric acid is adjusted is 2.0.
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| 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 |
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| CN111175385A (en) * | 2018-11-12 | 2020-05-19 | 珠海润都制药股份有限公司 | Irbesartan isomer testing method 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 |
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