CN104749286B - A kind of analyzing detecting method of valganciclovir hydrochloride impurity - Google Patents
A kind of analyzing detecting method of valganciclovir hydrochloride impurity Download PDFInfo
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Abstract
The invention belongs to technical field of analytical chemistry, specifically disclose a kind of HPLC analyzing detecting method of valganciclovir hydrochloride impurity, more particularly, it relates to a kind of analysis detection guanine, GCV, methoxy guanine, 1 N methylvaline ester of GCV, monovinylphenyl GCV, monochloro are for GCV or the method containing the medicine or its preparation of impurity in above-mentioned 6.The mobile phase in performance liquid chromatographic column of the solution to be measured injection with phenyl silane bonded silica gel as filler, being constituted with chromatographically pure organic solvent with the inorganic salt solution containing amine is rinsed and is separated, then detected with ultraviolet light analysis.This method creatively adds certain density ammonium acetate or ammonium formate first in mobile phase, the difficult problem that main peak and impurity peaks and other impurities peak between separate difficulty is thoroughly solved, easy to operate, separating degree is big, sensitivity is high, accuracy is good, by literature search, there is no similar document report.
Description
Technical field
The present invention relates to technical field of analytical chemistry, and in particular to a kind of HPLC analysis inspection of valganciclovir hydrochloride impurity
Survey method, more particularly, it relates to a kind of analysis detection guanine, GCV, methoxy guanine, GCV 1-
N- methylvaline ester, monovinylphenyl GCV, monochloro are for GCV or the medicine containing impurity in above-mentioned 6 or its preparation
Method.
Background technology
Valganciclovir hydrochloride bulk drug was placed in synthesis process and in valganciclovir Hydrochloride solid preparation
Six kinds of major impurities, respectively guanine, GCV, methoxy guanine, GCV 1-N- may be produced in journey
Methylvaline ester, monovinylphenyl GCV, monochloro are for GCV.
Above-mentioned 6 kinds of impurity, the number of its content directly affects the quality of valganciclovir hydrochloride or its preparation, easily draw
Human body bad reaction and medicine irritation is played, also determines the shelf-life length of medicine.Valganciclovir hydrochloride in American Pharmacopeia
The relevant substance detecting method of tablet adopts special phenyl post, and particle diameter is 3.0～3.5 μm, in actual applications chromatogram column pressure
Height, easily blocks, and needs to add pre-column process, causes chromatographic system pressure bigger, and cost increases, and is not suitable in actual production
Popularization and application, i.e., do not have broad applicability；Clearly do not report in existing valganciclovir hydrochloride and its preparation document this 6
The qualitatively and quantitatively method of impurity is planted, also this 6 kinds of detection can be just precisely separating using a kind of common chromatographic column nothing document report miscellaneous
The method of matter.
Content of the invention
For the deficiencies in the prior art, the technical problem to be solved is how to adopt general phenyl
Chromatographic column analyzes above-mentioned 6 kinds of impurity in detection valganciclovir hydrochloride bulk drug and its preparation exactly.The present invention for solve with
Upper technical problem, creatively adds the certain density formates containing ammonium or acetate first in mobile phase, thoroughly solves
A difficult problem for difficulty is separated between main peak and impurity peaks and other impurities peak, and easy to operate, sensitivity is high, accuracy is good.Carry
Supplied a kind of relevant substance detecting method of valganciclovir hydrochloride, for analyze detection valganciclovir hydrochloride bulk drug or its
Impurity in preparation, this method are particularly suitable for using general silane group silicagel column, especially phenyl silane bonded silica gel color
6 kinds of impurity in spectrum post analysis detection valganciclovir hydrochloride bulk drug or its preparation, by literature search, there is no similar document report
Road.
The present invention provide technical scheme be：
A kind of valganciclovir hydrochloride comprises the steps about the analyzing detecting method of material：
A. valganciclovir hydrochloride medicine or its preparation is taken, is mixed with water, watery hydrochloric acid, the aqueous solution of organic solvent or flowing
Solution, is configured to the solution of 400～600 μ g/ml of hydrochloric valganciclovir, preferably 500 μ g/ml；
The aqueous solution of the organic solvent is selected from alcohols or the aqueous solution of nitrile, preferably methyl alcohol or acetonitrile solution.
It is preferred that with watery hydrochloric acid or flowing phased soln, the watery hydrochloric acid concentration be 0.0005～0.002mol/L, most preferably
0.001mol/L；
B. take step a resulting solution appropriate, be injected in high performance liquid chromatograph, be rinsed with mobile phase, the height
Performance liquid chromatographic column in effect liquid phase chromatogram instrument with silane group silica gel as filler with phenyl silane bonded silica gel is preferably
Filler；
C. using the ultraviolet light detection that wavelength is 240～260nm, best detection wavelength is 254nm.
The valganciclovir hydrochloride medicine or its preparation are selected from valganciclovir hydrochloride bulk drug, valganciclovir hydrochloride
Piece, valganciclovir hydrochloride capsule, valganciclovir hydrochloride granule etc..
Containing one or more in following six kinds of impurity in the valganciclovir hydrochloride medicine or its preparation, excellent
Select five kinds or six kinds：Guanine, GCV, methoxy guanine, GCV 1-N- methylvaline ester, single acetyl
Epoxide GCV, monochloro are for GCV.
The mobile phase is made up of amine-inorganic mixed salt solution and chromatographically pure organic solvent, and the amine-inorganic salts are mixed
The volume ratio for solution being closed with chromatographically pure organic solvent is 99：1～90：10, volume ratio is preferably 98：2～92：8, more preferably
97：3～93：7, optimum is 95：5.
The preferred methyl alcohol of the chromatographically pure organic solvent or acetonitrile, optimum is methyl alcohol.
The pH value of the amine-inorganic mixed salt solution is 5.0～6.0, preferably 5.2～5.8.Amine-the inorganic salts
Contain amine 0.1%～0.5% (v/v), 0.01～0.05mol/L containing inorganic salts, the amine-inorganic salts mixing in mixed solution
Also contain the organic acid for adjusting pH value in solution.
The amine-inorganic mixed salt solution can be understood as cushioning liquid.
The preferred diethylamine of the amine, triethylamine, optimum is triethylamine.
The preferred ammonium formate of the inorganic salts, ammonium acetate, optimum is ammonium acetate.
The preferred formic acid of the organic acid, trifluoroacetic acid, optimum is trifluoroacetic acid.
Compared with prior art, the advantage of analyzing detecting method of the present invention and beneficial effect are：
The valganciclovir hydrochloride of the present invention, about the analyzing detecting method of material, by literature search, is to flow first
The certain density formates containing ammonium or acetate is creatively added in phase, thoroughly solves main peak and impurity peaks and adjacent
A difficult difficult problem is separated between impurity peaks, easy to operate, sensitivity is high, accuracy is good.For analyzing detection hydrochloric acid figured silk fabrics more former times Lip river
When Wei or its preparation impurity, between main peak and impurity, other impurities, all there is relatively satisfactory separating degree, each impurity all has low inspection
Limit is surveyed, LDL is about 0.02 μ g/ml, quantitative limit is about 0.05 μ g/ml.In each impurity concentration in 0.05～20 μ g/ml
In the range of have good linear relationship, coefficient correlation is up to 0.9999.
Description of the drawings
Fig. 1 is the HPLC analysis chart of valganciclovir hydrochloride bulk drug in embodiment 1；
Fig. 2 is the HPLC analysis chart of valganciclovir hydrochloride tablet in embodiment 1；
Fig. 3 is the HPLC analysis chart that valganciclovir hydrochloride bulk drug adds six impurity in embodiment 2；
Fig. 4 is the HPLC analysis chart that valganciclovir hydrochloride tablet adds five impurity in embodiment 3；
Fig. 5 is the HPLC analysis chart that valganciclovir hydrochloride tablet adds five impurity in embodiment 4；
Fig. 6 is the HPLC analysis chart that valganciclovir hydrochloride Capsule content adds five impurity in embodiment 5；
Fig. 7 is the HPLC analysis chart that valganciclovir hydrochloride Capsule content adds five impurity in embodiment 6；
Fig. 8 is the HPLC analysis chart that valganciclovir hydrochloride granule adds five impurity in embodiment 7；
Fig. 9 is the HPLC analysis chart that valganciclovir hydrochloride granule adds five impurity in embodiment 8.
Specific embodiment
For ease of skilled artisan understands that present invention, below applicant will retouch further in conjunction with specific embodiments
Technical scheme is stated, but herein below limits the present invention never in any form.
Below in all embodiments, main agents used and raw material are described below：
Methoxymethylguanine (methoxy guanine) standard items；
Ganciclovir Mono-N-Methyl Valinate (GCV 1-N- methylvaline ester) standard items；
Isomonochloro Ganciclovir (monochloro is for GCV) standard items；
Monoacetoxy Ganciclovir (monovinylphenyl GCV) standard items, CAS Number：88110-
89-8；
Valganciclovir hydrochloride bulk drug, tablet, capsule, granule be applicant according to conventional prior from
System.
Other are conventional reagent and raw material.
Below in all embodiments, the percentage of unreceipted unit refers both to mass percent.
Embodiment 1
Chromatographic condition：With phenyl silane bonded silica gel as filler, mobile phase is buffered for the triethylamine ammonium formate of pH=5.6
Liquid (contains three kinds of component triethylamines, ammonium formate and trifluoroacetic acids, its content is respectively than water：0.2% (v/v) three second
Amine, 0.05mol/L ammonium formate and appropriate trifluoroacetic acid, wherein appropriate trifluoroacetic acid in order to adjust the pH of the buffer solution to
5.6)-methyl alcohol, the buffer solution are 95 with methyl alcohol volume ratio：5, flow velocity 1.0ml/min, Detection wavelength 254nm, chromatographic column：
COSMOSIL 150 × 4.6mm of 5PE-MS, 5 μm, 35 DEG C of column temperature, sample size：50μl.
Prepared by need testing solution：
Valganciclovir hydrochloride bulk drug about 50mg is taken, accurately weighed, put in 100ml volumetric flask, plus 0.001mol/L salt
Acid dissolving is simultaneously diluted to scale, used as valganciclovir hydrochloride need testing solution.
Valganciclovir hydrochloride piece is taken, finely ground, precision is weighed and (is approximately equivalent to hydrochloric valganciclovir main ingredient in right amount
50mg) put in 100ml volumetric flask, plus 0.001mol/L dissolving with hydrochloric acid scale is diluted to, supply as valganciclovir hydrochloride piece
Test sample solution.
Prepared by poly-doped impurity reference substance solution：
Take guanine, GCV and other four kinds of impurity each appropriate, accurately weighed, put in 100ml volumetric flask, use
0.001mol/L hydrochloric acid makees diluent, is configured to 5 μ g/ml of guanine, 10 μ g/ml of GCV and methoxy bird fast
Purine, GCV 1-N- methylvaline ester, monovinylphenyl GCV, monochloro are miscellaneous for the mixing of the equal 1 μ g/ml of GCV
Matter reference substance solution.
Three of the above solution respectively takes 50 μ l injection liquid chromatograph, records chromatogram, calculates bird by area with external standard method fast
Purine and the content of GCV, are calculated the content of other impurities by area normalization method.
It is guanine 0.007% to measure each impurity content in valganciclovir hydrochloride bulk drug, GCV 0.416%,
GCV 1-N- methylvaline ester 0.120%, monovinylphenyl GCV 0.117%, other two impurity are not detected.
Shown in the HPLC analysis chart as accompanying drawing 1 of valganciclovir hydrochloride bulk drug, wherein retention time 3.036min is guanine chromatogram
Peak, 3.826min are GCV chromatographic peak, and 7.550min and 8.211min is two chromatogram main peaks of valganciclovir hydrochloride,
8.950min and 9.978min is GCV 1-N- methylvaline two chromatographic peaks of ester, and 10.777min is monovinylphenyl
GCV chromatographic peak.
In valganciclovir hydrochloride piece, each impurity content is guanine 0.019%, GCV 0.278%, GCV
1-N- methylvaline ester 0.175%, monovinylphenyl GCV 0.068%, other two impurity are not detected.Hydrochloric acid figured silk fabrics is more
Shown in the HPLC analysis chart as accompanying drawing 2 of VCV piece, wherein retention time 3.038min is guanine chromatographic peak, and 3.834min is
GCV chromatographic peak, 7.570min and 8.232min be two chromatogram main peaks of valganciclovir hydrochloride, 8.992min and
9.997min is GCV 1-N- methylvaline ester chromatographic peak, and 10.789min is monovinylphenyl GCV chromatographic peak.
Embodiment 2
Chromatographic condition：With phenyl silane bonded silica gel as filler, mobile phase is buffered for the triethylamine acetate ammonium of pH=5.3
Liquid (contains three kinds of component triethylamines, ammonium acetate and formic acid, its content is respectively than water：0.25% (v/v) triethylamine,
0.03mol/L ammonium acetate and appropriate formic acid, wherein appropriate formic acid is in order to adjust the pH value of the buffer solution to 5.3)-first
Alcohol, the buffer solution are 95 with methyl alcohol volume ratio：5, flow velocity 1.0ml/min, Detection wavelength 254nm, chromatographic column：COSMOSIL
150 × 4.6mm of 5PE-MS, 5 μm, 35 DEG C of column temperature, sample size：50μl.
Prepared by need testing solution：
Valganciclovir hydrochloride bulk drug about 50mg is taken, accurately weighed, put in 100ml volumetric flask, add six impurity each
In right amount, 5 μ g/ml of guanine, 10 μ g/ml of GCV and methoxy guanine, GCV 1-N- methyl are configured to
L-valine ester, monovinylphenyl GCV, monochloro are for the equal 1 μ g/ml of GCV, plus 0.001mol/L dissolving with hydrochloric acid dilute
To scale, as valganciclovir hydrochloride mixed solution.
50 μ l valganciclovir hydrochloride mixed solutions injection liquid chromatograph is taken, records chromatogram.
The theoretical cam curve of second valganciclovir hydrochloride main peak is 6973, and separating degree is：Guanine and GCV
For 4.84, GCV is 12.79 with methoxy guanine, methoxy guanine and first hydrochloric acid figured silk fabrics more former times Lip river
Wei main peak is 2.09, and second valganciclovir hydrochloride main peak with first GCV 1-N- methylvaline ester peak is
1.72, second GCV 1-N- methylvaline ester peak is 1.63 with monovinylphenyl GCV peak, monochloro more former times in generation
Luo Wei is 1.89 with adjacent unknown impuritie peak.Each impurity LDL is about 0.02 μ g/ml, and quantitative limit is about 0.05 μ g/
ml.There is good linear relationship in each impurity concentration in the range of 0.05～20 μ g/ml, coefficient correlation is 0.9999.
Valganciclovir hydrochloride bulk drug adds shown in the HPLC analysis chart as accompanying drawing 3 of six impurity, wherein retention time
3.028min is guanine chromatographic peak, and 3.835min is GCV chromatographic peak, and 6.821min is methoxy guanine color
Spectral peak, 7.489min and 8.143min are two chromatogram main peaks of valganciclovir hydrochloride, and 8.937min and 9.847min is more former times
Lip river Wei 1-N- methylvaline ester chromatographic peak, 10.645min are monovinylphenyl GCV chromatographic peak, and 14.346min is single
Chloro GCV chromatographic peak.
Embodiment 3
Chromatographic condition：With phenyl silane bonded silica gel as filler, mobile phase is buffered for the triethylamine acetate ammonium of pH=5.4
Liquid (contains three kinds of component triethylamines, ammonium acetate and trifluoroacetic acids, its content is respectively than water：0.35% (v/v) three second
Amine, 0.03mol/L ammonium acetate and appropriate trifluoroacetic acid, wherein appropriate trifluoroacetic acid in order to adjust the pH of the buffer solution to
5.4)-methyl alcohol, the buffer solution are 96 with methyl alcohol volume ratio：4, flow velocity 1.0ml/min, Detection wavelength 254nm, chromatographic column：
COSMOSIL 150 × 4.6mm of 5PE-MS, 5 μm, 30 DEG C of column temperature, sample size：50μl.
Prepared by need testing solution：
Valganciclovir hydrochloride piece is taken, finely ground, precision is weighed and (is approximately equivalent to hydrochloric valganciclovir main ingredient in right amount
50mg) put in 100ml volumetric flask, add guanine, GCV, methoxy guanine, GCV 1-N- methyl figured silk fabrics
Propylhomoserin ester, five impurity of monovinylphenyl GCV are each appropriate, be configured to 5 μ g/ml of guanine, 10 μ g/ml of GCV with
And methoxy guanine, GCV 1-N- methylvaline ester, the equal 1 μ g/ml of monovinylphenyl GCV, add water molten
Solve and scale is diluted to, as valganciclovir hydrochloride piece mixed solution.
50 μ l valganciclovir hydrochloride piece mixed solutions injection liquid chromatograph is taken, records chromatogram.
The theoretical cam curve of second valganciclovir hydrochloride main peak is 7233, and separating degree is：Guanine and GCV
For 5.42, GCV is 13.83 with methoxy guanine, methoxy guanine and first hydrochloric acid figured silk fabrics more former times Lip river
Wei main peak is 2.01, and second valganciclovir hydrochloride main peak with first GCV 1-N- methylvaline ester peak is
1.69, second GCV 1-N- methylvaline ester peak is 2.40 with monovinylphenyl GCV peak.Each impurity is most
Low test limit is about 0.024 μ g/ml, and quantitative limit is about 0.07 μ g/ml.Each impurity concentration 0.07～20 μ g/ml scope
Inside there is good linear relationship, coefficient correlation is 0.9999.
Valganciclovir hydrochloride piece adds shown in the HPLC analysis chart as accompanying drawing 4 of five impurity, wherein retention time
3.188min is guanine chromatographic peak, and 4.090min is GCV chromatographic peak, and 7.393min is methoxy guanine color
Spectral peak, 8.058min and 8.798min are two chromatogram main peaks of valganciclovir hydrochloride, and 9.616min and 10.642min is more former times
Lip river Wei 1-N- methylvaline two chromatographic peaks of ester, 11.800min are monovinylphenyl GCV chromatographic peak.
Embodiment 4
Chromatographic condition：With phenyl silane bonded silica gel as filler, mobile phase is buffered for the triethylamine acetate ammonium of pH=5.5
Liquid (contains three kinds of component triethylamines, ammonium acetate and trifluoroacetic acids, its content is respectively than water：0.15% (v/v) three second
Amine, 0.01mol/L ammonium acetate and appropriate trifluoroacetic acid, wherein appropriate trifluoroacetic acid in order to adjust the pH of the buffer solution to
5.5)-methyl alcohol, the buffer solution are 95 with methyl alcohol volume ratio：5, flow velocity 1.0ml/min, Detection wavelength 254nm, chromatographic column：
COSMOSIL 150 × 4.6mm of 5PE-MS, 5 μm, 35 DEG C of column temperature, sample size：50μl.
Prepared by need testing solution：
Valganciclovir hydrochloride piece is taken, finely ground, precision is weighed and (is approximately equivalent to hydrochloric valganciclovir main ingredient in right amount
60mg) put in 100ml volumetric flask, add guanine, GCV, methoxy guanine, GCV 1-N- methyl figured silk fabrics
Propylhomoserin ester, five impurity of monovinylphenyl GCV are each appropriate, be configured to 5 μ g/ml of guanine, 10 μ g/ml of GCV with
And methoxy guanine, GCV 1-N- methylvaline ester, the equal 1 μ g/ml of monovinylphenyl GCV, add water-
Methyl alcohol (volume ratio 95：5) dissolve and scale is diluted to, as valganciclovir hydrochloride piece mixed solution.
50 μ l valganciclovir hydrochloride piece mixed solutions injection liquid chromatograph is taken, records chromatogram.
The theoretical cam curve of second valganciclovir hydrochloride main peak is 6318, and separating degree is：Guanine and GCV
For 4.68, GCV is 12.39 with methoxy guanine, methoxy guanine and first hydrochloric acid figured silk fabrics more former times Lip river
Wei main peak is 2.02, and second valganciclovir hydrochloride main peak with first GCV 1-N- methylvaline ester peak is
1.65, second GCV 1-N- methylvaline ester peak is 1.66 with monovinylphenyl GCV peak.Each impurity is most
Low test limit is about 0.02 μ g/ml, and quantitative limit is about 0.06 μ g/ml.In each impurity concentration in the range of 0.06～20 μ g/ml
There is good linear relationship, coefficient correlation is 0.9997～0.9999.
Valganciclovir hydrochloride piece adds shown in the HPLC analysis chart as accompanying drawing 5 of five impurity, wherein retention time
3.002min is guanine chromatographic peak, and 3.804min is GCV chromatographic peak, and 6.739min is methoxy guanine color
Spectral peak, 7.401min and 8.043min are two chromatogram main peaks of valganciclovir hydrochloride, and 8.782min and 9.688min is more former times
Lip river Wei 1-N- methylvaline two chromatographic peaks of ester, 10.480min are monovinylphenyl GCV chromatographic peak.
Embodiment 5
Chromatographic condition：With phenyl silane bonded silica gel as filler, mobile phase is the diethylamine ammonium acetate buffer of pH=5.7
Liquid (contains three kinds of component diethylamine, ammonium acetate and trifluoroacetic acids, its content is respectively than water：0.45% (v/v) diethyl
Amine, 0.05mol/L ammonium acetate and appropriate trifluoroacetic acid, wherein appropriate trifluoroacetic acid in order to adjust the pH of the buffer solution to
5.7)-methyl alcohol, the buffer solution are 94 with methyl alcohol volume ratio：6, flow velocity 1.0ml/min, Detection wavelength 254nm, chromatographic column：
COSMOSIL 150 × 4.6mm of 5PE-MS, 5 μm, 35 DEG C of column temperature, sample size：50μl.
Prepared by need testing solution：
Valganciclovir hydrochloride capsule 's content is taken, finely ground, precision is weighed and (is approximately equivalent to hydrochloric valganciclovir in right amount
Main ingredient 50mg) put in 100ml volumetric flask, add guanine, GCV, methoxy guanine, GCV 1-N- first
Base L-valine ester, five impurity of monovinylphenyl GCV are each appropriate, are configured to 5 μ g/ml of guanine, 10 μ g/ of GCV
Ml and methoxy guanine, GCV 1-N- methylvaline ester, the equal 1 μ g/ml of monovinylphenyl GCV, plus
Flowing phased soln is simultaneously diluted to scale, used as valganciclovir hydrochloride capsule mixed solution.
50 μ l valganciclovir hydrochloride capsule mixed solutions injection liquid chromatograph is taken, records chromatogram.
The theoretical cam curve at second valganciclovir hydrochloride peak is 6404, and separating degree is：Guanine with GCV is
3.95, GCV and methoxy guanine are 11.60, methoxy guanine and first valganciclovir hydrochloride
Main peak is 1.58, and second valganciclovir hydrochloride main peak and first GCV 1-N- methylvaline ester peak are 1.68,
Second GCV 1-N- methylvaline ester peak is 1.33 with monovinylphenyl GCV peak.Each impurity lowest detection
Limit is about 0.02 μ g/ml, and quantitative limit is about 0.07 μ g/ml.Have in the range of 0.07～20 μ g/ml well in each impurity concentration
Linear relationship, coefficient correlation be 0.9995～0.9999.
Valganciclovir hydrochloride capsule 's content adds shown in the HPLC analysis chart as accompanying drawing 6 of five impurity, when wherein retaining
Between 2.918min be guanine chromatographic peak, 3.573min be GCV chromatographic peak, 6.207min be methoxy guanine
Chromatographic peak, 6.687min and 7.229min are two chromatogram main peaks of valganciclovir hydrochloride, and 7.893min and 8.655min is for more
VCV 1-N- methylvaline two chromatographic peaks of ester, 9.240min are monovinylphenyl GCV chromatographic peak.
Embodiment 6
Chromatographic condition：With phenyl silane bonded silica gel as filler, mobile phase is buffered for the triethylamine acetate ammonium of pH=5.3
Liquid (contains three kinds of component triethylamines, ammonium acetate and trifluoroacetic acids, its content is respectively than water：0.45% (v/v) three second
Amine, 0.02mol/L ammonium acetate and appropriate trifluoroacetic acid, wherein appropriate trifluoroacetic acid in order to adjust the pH of the buffer solution to
5.3)-methyl alcohol, the buffer solution are 95 with the volume ratio of methyl alcohol：5, flow velocity 1.0ml/min, Detection wavelength 254nm, chromatographic column：
Thermo BetaBasic 250 × 4.6mm of Phenyl, 5 μm, 35 DEG C of column temperature, sample size：50μl.
Prepared by need testing solution：
Valganciclovir hydrochloride capsule 's content is taken, finely ground, precision is weighed and (is approximately equivalent to hydrochloric valganciclovir in right amount
Main ingredient 50mg) put in 100ml volumetric flask, add guanine, GCV, methoxy guanine, GCV 1-N- first
Base L-valine ester, five impurity of monovinylphenyl GCV are each appropriate, are configured to 5 μ g/ml of guanine, 10 μ g/ of GCV
Ml and methoxy guanine, GCV 1-N- methylvaline ester, the equal 1 μ g/ml of monovinylphenyl GCV, plus
Flowing phased soln is simultaneously diluted to scale, used as valganciclovir hydrochloride capsule mixed solution.
50 μ l valganciclovir hydrochloride capsule mixed solutions injection liquid chromatograph is taken, records chromatogram.
The theoretical cam curve at second valganciclovir hydrochloride peak is 7639, and separating degree is：Guanine with GCV is
5.45, GCV and methoxy guanine are 13.74, methoxy guanine and first valganciclovir hydrochloride
Main peak is 2.69, and second valganciclovir hydrochloride main peak and first GCV 1-N- methylvaline ester peak are 1.76,
Second GCV 1-N- methylvaline ester peak is 1.53 with monovinylphenyl GCV peak.Each impurity lowest detection
Limit is about 0.03 μ g/ml, and quantitative limit is about 0.08 μ g/ml.Have in the range of 0.08～20 μ g/ml well in each impurity concentration
Linear relationship, coefficient correlation be 0.9996～0.9999.
Valganciclovir hydrochloride capsule adds shown in the HPLC analysis chart as accompanying drawing 7 of five impurity, wherein retention time
3.065min is guanine chromatographic peak, and 3.950min is GCV chromatographic peak, and 7.068min is methoxy guanine color
Spectral peak, 7.922min and 8.642min are two chromatogram main peaks of valganciclovir hydrochloride, and 9.500min and 10.498min is more former times
Lip river Wei 1-N- methylvaline two chromatographic peaks of ester, 11.242min are monovinylphenyl GCV chromatographic peak.
Embodiment 7
Chromatographic condition：With phenyl silane bonded silica gel as filler, mobile phase is the diethylamine ammonium acetate buffer of pH=5.7
Liquid (contains three kinds of component diethylamine, ammonium acetate and trifluoroacetic acids, its content is respectively than water：0.45% (v/v) diethyl
Amine, 0.05mol/L ammonium acetate and appropriate trifluoroacetic acid, wherein appropriate trifluoroacetic acid in order to adjust the pH of the buffer solution to
5.7)-acetonitrile, the buffer solution are 94 with acetonitrile volume ratio：6, flow velocity 1.0ml/min, Detection wavelength 254nm, chromatographic column：
COSMOSIL 150 × 4.6mm of 5PE-MS, 5 μm, 35 DEG C of column temperature, sample size：50μl.
The preparation of need testing solution：
Valganciclovir hydrochloride particle is taken, finely ground, precision is weighed and (is approximately equivalent to hydrochloric valganciclovir main ingredient in right amount
50mg) put in 100ml volumetric flask, add guanine, GCV, methoxy guanine, GCV 1-N- methyl figured silk fabrics
Propylhomoserin ester, five impurity of monovinylphenyl GCV are each appropriate, be configured to 5 μ g/ml of guanine, 10 μ g/ml of GCV with
And methoxy guanine, GCV 1-N- methylvaline ester, the equal 1 μ g/ml of monovinylphenyl GCV, plus flowing
Phased soln is simultaneously diluted to scale, used as valganciclovir hydrochloride particle mixed solution.
50 μ l valganciclovir hydrochloride particle mixed solutions injection liquid chromatograph is taken, records chromatogram.
The theoretical cam curve at second valganciclovir hydrochloride peak is 8107, and separating degree is：Guanine with GCV is
4.81, GCV and methoxy guanine are 13.08, methoxy guanine and first valganciclovir hydrochloride
Main peak is 2.70, and second valganciclovir hydrochloride main peak and first GCV 1-N- methylvaline ester peak are 1.86,
Second GCV 1-N- methylvaline ester peak is 0.92 with monovinylphenyl GCV peak.Each impurity lowest detection
Limit is about 0.02 μ g/ml, and quantitative limit is about 0.06 μ g/ml.Have in the range of 0.06～20 μ g/ml well in each impurity concentration
Linear relationship, coefficient correlation be 0.9995～0.9999.
Valganciclovir hydrochloride particle adds shown in the HPLC analysis chart as accompanying drawing 8 of five impurity, wherein retention time
2.857min is guanine chromatographic peak, and 3.557min is GCV chromatographic peak, and 6.167min is methoxy guanine color
Spectral peak, 6.902min and 7.475min are two chromatogram main peaks of valganciclovir hydrochloride, and 8.185min and 8.979min is more former times
Lip river Wei 1-N- methylvaline two chromatographic peaks of ester, 9.328min are monovinylphenyl GCV chromatographic peak.
Embodiment 8
Chromatographic condition：With phenyl silane bonded silica gel as filler, mobile phase is buffered for the triethylamine acetate ammonium of pH=5.4
Liquid (contains three kinds of component triethylamines, ammonium acetate and trifluoroacetic acids, its content is respectively than water：0.45% (v/v) three second
Amine, 0.05mol/L ammonium acetate and appropriate trifluoroacetic acid, wherein appropriate trifluoroacetic acid in order to adjust the pH of the buffer solution to
5.4)-methyl alcohol, the buffer solution are 94 with methyl alcohol volume ratio：6, flow velocity 1.0ml/min, Detection wavelength 254nm, chromatographic column：
COSMOSIL 150 × 4.6mm of 5PE-MS, 5 μm, 35 DEG C of column temperature, sample size：50μl.
The preparation of need testing solution：
Valganciclovir hydrochloride particle is taken, finely ground, precision is weighed and (is approximately equivalent to hydrochloric valganciclovir main ingredient in right amount
40mg) put in 100ml volumetric flask, add guanine, GCV, methoxy guanine, GCV 1-N- methyl figured silk fabrics
Propylhomoserin ester, five impurity of monovinylphenyl GCV are each appropriate, be configured to 5 μ g/ml of guanine, 10 μ g/ml of GCV with
And methoxy guanine, GCV 1-N- methylvaline ester, the equal 1 μ g/ml of monovinylphenyl GCV, plus flowing
Phased soln is simultaneously diluted to scale, used as valganciclovir hydrochloride particle mixed solution.
50 μ l valganciclovir hydrochloride particle mixed solutions injection liquid chromatograph is taken, records chromatogram.
The theoretical cam curve at second valganciclovir hydrochloride peak is 8107, and separating degree is：Guanine with GCV is
4.41, GCV and methoxy guanine are 11.78, methoxy guanine and first valganciclovir hydrochloride
Main peak is 1.90, and second valganciclovir hydrochloride main peak and first GCV 1-N- methylvaline ester peak are 1.65,
Second GCV 1-N- methylvaline ester peak is 1.46 with monovinylphenyl GCV peak.Each impurity lowest detection
Limit is about 0.02 μ g/ml, and quantitative limit is about 0.05 μ g/ml.Have in the range of 0.05～20 μ g/ml well in each impurity concentration
Linear relationship, coefficient correlation be 0.9997～0.9999.
Valganciclovir hydrochloride particle adds shown in the HPLC analysis chart as accompanying drawing 9 of five impurity, wherein retention time
2.992min is guanine chromatographic peak, and 3.784min is GCV chromatographic peak, and 6.701min is methoxy guanine color
Spectral peak, 7.355min and 7.992min are two chromatogram main peaks of valganciclovir hydrochloride, and 8.713min and 9.632min is more former times
Lip river Wei 1-N- methylvaline two chromatographic peaks of ester, 10.428min are monovinylphenyl GCV chromatographic peak.
Claims (11)
1. a kind of valganciclovir hydrochloride comprises the steps about the analyzing detecting method of material：
A. valganciclovir hydrochloride medicine or its preparation are taken, with water, watery hydrochloric acid, the aqueous solution of organic solvent or flowing phased soln,
It is configured to the solution of 400 ~ 600 g/ml of hydrochloric valganciclovir；
The aqueous solution of the organic solvent is selected from alcohols or the aqueous solution of nitrile；
The watery hydrochloric acid concentration is 0.0005 ~ 0.002mol/L；
B. step a resulting solution is taken, is injected in high performance liquid chromatograph, is rinsed with mobile phase；
Performance liquid chromatographic column in the high performance liquid chromatograph is with phenyl silane bonded silica gel as filler；
C. the ultraviolet light using wavelength for 240 ~ 260nm is detected；
The mobile phase is made up of amine-inorganic mixed salt solution and chromatographically pure organic solvent, and the amine-inorganic salts mixing is molten
Liquid is 99 with the volume ratio of chromatographically pure organic solvent：1~90：10；
The chromatographically pure organic solvent selects methyl alcohol or acetonitrile；
The pH value of the amine-inorganic mixed salt solution is 5.2 ~ 5.8；Contain amine in the amine-inorganic mixed salt solution
0.1%~0.5%（v/v）, 0.01 ~ 0.05mol/L containing inorganic salts, in the amine-inorganic mixed salt solution also contain for adjusting
The organic acid of pH value；
The amine is diethylamine and/or triethylamine；
The inorganic salts are ammonium acetate；
The organic acid is formic acid and/or trifluoroacetic acid；
Containing five kinds or six kinds in following six kinds of impurity in the valganciclovir hydrochloride medicine or its preparation：Guanine, more
In VCV, methoxy guanine, GCV 1-N- methylvaline ester, monovinylphenyl GCV, monochloro generation, are more
VCV.
2. analyzing detecting method according to claim 1, it is characterised in that：The valganciclovir hydrochloride medicine or its system
Agent is selected from valganciclovir hydrochloride bulk drug, valganciclovir hydrochloride piece, valganciclovir hydrochloride capsule, valganciclovir hydrochloride
Granule.
3. according to the analyzing detecting method described in claim 1, it is characterised in that：The aqueous solution of the organic solvent is methyl alcohol or second
The nitrile aqueous solution.
4. according to the analyzing detecting method described in claim 1, it is characterised in that：Hydrochloric acid figured silk fabrics more former times is dissolved with watery hydrochloric acid or mobile phase
Lip river Wei medicine or its preparation.
5. according to arbitrary described analyzing detecting method in claim 1-4, it is characterised in that：The chromatographically pure organic solvent is first
Alcohol.
6. analyzing detecting method according to claim 1, it is characterised in that：The amine-inorganic mixed salt solution and color
The volume ratio for composing pure organic solvent is 98：2~92：8.
7. analyzing detecting method according to claim 6, it is characterised in that：The amine-inorganic mixed salt solution and color
The volume ratio for composing pure organic solvent is 97：3~93：7.
8. analyzing detecting method according to claim 6, it is characterised in that：The amine-inorganic mixed salt solution and color
The volume ratio for composing pure organic solvent is 95：5.
9. analyzing detecting method according to claim 1, it is characterised in that：The Detection wavelength is 254nm.
10. according to arbitrary described analyzing detecting method in claim 4, it is characterised in that：The amine is triethylamine.
11. according to arbitrary described analyzing detecting method in claim 1-4, it is characterised in that：The organic acid is trifluoroacetic acid.
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| CN108872405B (en) * | 2017-05-11 | 2021-06-01 | 武汉先路医药科技股份有限公司 | HPLC analysis detection method for relative substances of lodoxylamine tromethamine |
| CN108267519B (en) * | 2017-12-05 | 2021-02-12 | 湖北丽益医药科技有限公司 | HPLC detection method for valganciclovir hydrochloride intermediate hydrolysate isomer |
| CN110763792A (en) * | 2019-10-24 | 2020-02-07 | 湖北省宏源药业科技股份有限公司 | Method for detecting guanine-related substances |
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| CN102718765B (en) * | 2011-03-31 | 2015-01-07 | 四川科伦药物研究有限公司 | Method for preparing and purifying Valganciclovir hydrochloride |
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| CN109212093A (en) * | 2018-10-29 | 2019-01-15 | 湖北丽益医药科技有限公司 | A kind of HPLC detection method of valganciclovir hydrochloride intermediate condensate isomers |
| CN109212093B (en) * | 2018-10-29 | 2021-06-01 | 湖北丽益医药科技有限公司 | HPLC detection method of valganciclovir hydrochloride intermediate condensation compound isomer |
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