CN105929031B - The separation method of impurity in the compound of a kind of angiotensin receptor antagonist and nep inhibitor - Google Patents
The separation method of impurity in the compound of a kind of angiotensin receptor antagonist and nep inhibitor Download PDFInfo
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- CN105929031B CN105929031B CN201510958419.2A CN201510958419A CN105929031B CN 105929031 B CN105929031 B CN 105929031B CN 201510958419 A CN201510958419 A CN 201510958419A CN 105929031 B CN105929031 B CN 105929031B
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- 0 CCCCC(N(C*)Cc(cc1)ccc1-c(cccc1)c1C([N-])=N*=N)=O Chemical compound CCCCC(N(C*)Cc(cc1)ccc1-c(cccc1)c1C([N-])=N*=N)=O 0.000 description 2
- VLHKRWQICGMBAB-DEOSSOPVSA-O CCOC(CCCC[C@@H](Cc(cc1)ccc1-c1ccccc1)NC(CCC([OH2+])=C)=O)=O Chemical compound CCOC(CCCC[C@@H](Cc(cc1)ccc1-c1ccccc1)NC(CCC([OH2+])=C)=O)=O VLHKRWQICGMBAB-DEOSSOPVSA-O 0.000 description 1
- ZZLCFHIKESPLTH-UHFFFAOYSA-N Cc(cc1)ccc1-c1ccccc1 Chemical compound Cc(cc1)ccc1-c1ccccc1 ZZLCFHIKESPLTH-UHFFFAOYSA-N 0.000 description 1
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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
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/26—Conditioning of the fluid carrier; Flow patterns
- G01N30/28—Control of physical parameters of the fluid carrier
- G01N30/34—Control of physical parameters of the fluid carrier of fluid composition, e.g. gradient
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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
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Abstract
The present invention provides the separation methods of impurity in the compound of a kind of angiotensin receptor antagonist and nep inhibitor, including：Impurity separation is carried out to the pharmaceutical composition of angiotensin receptor antagonist and nep inhibitor using high performance liquid chromatography；Mobile phase in the high performance liquid chromatography includes water phase and organic phase；The water phase includes phosphate buffer, trifluoroacetic acid solution or formic acid solution；The organic phase includes organic mixture or acetonitrile, and the organic mixture includes acetonitrile and methanol；The volume ratio of the water phase and organic phase is (45%~65%)：(55%~35%).The present invention carries out impurity separation using high performance liquid chromatography, the mobile phase selected during high performance liquid chromatography includes the water phase and organic phase of special component, and the plurality of impurities in the compound of angiotensin receptor antagonist and nep inhibitor can be effectively separated using this mobile phase.
Description
Technical field
The present invention relates to the composite technology fields more particularly to one of angiotensin receptor antagonist and nep inhibitor
The separation method of impurity in the compound of kind angiotensin receptor antagonist and nep inhibitor.
Background technology
Angiotensin receptor blocker (ARBs, angiotensin-ii antagonist) can be by preventing Angiotensin II
It is combined with its receptor on vascular wall, is reduced so as to cause blood pressure.Since AT1 receptors can be inhibited, so such antagonist
It can be used for anti-hypertension, or for treating congestive heart failure and other indications.Neutral endopeptidase (EC
3.4.24.11；Enkephalinase；atriopeptidase；NEP it is) metalloproteinases containing zinc, it can crack various hydrophobic residual
Peptide substrates on the aminoterminal of base, the blood plasma level that nep inhibitor (neutral endopeptidase inhibitor, NEPi) can reduce ANP, because
And natruresis can be promoted and there is diuresis.Angiotensin receptor antagonist and nep inhibitor compound ([3- ((1S,
3R) -1- biphenyl -4- ylmethyls -3- ethoxy carbonyl -1- butylcarbamoyls) -3 '-methyl -2 ' of propionic acid-(S)-(valeryl
{ 2 "-(tetrazolium -5- bases) biphenyl -4 '-ylmethyl } amino) butyric acid] half pentahydrate of trisodium) it is a kind of answering with double action
Object is closed, there is structure shown in Formulas I：
Angiotensin receptor antagonist and nep inhibitor compound have extensive clinical generalization value.
Since the compound drug products of angiotensin receptor antagonist and nep inhibitor are from synthesis starting material to end
Product, during which all there may be impurity for each process, as that may bring starting material, reagent, intermediate and pair in production process into
The impurity such as product, these Impurities Upon Product Qualities produce a very large impact.Therefore, in order to ensure the safe and effective of drug, to drug
In the separation of each impurity be of great significance.
Currently, yet there are no about angiotensin receptor antagonist in nep inhibitor compound impurity detach it is related
Report.
Invention content
In view of this, the purpose of the present invention is to provide a kind of angiotensin receptor antagonist and nep inhibitor are compound
The separation method of impurity in object, method provided by the invention can efficiently separate angiotensin receptor antagonist and NEP suppressions
Plurality of impurities in inhibitor complex.
The present invention provides the separation sides of impurity in the compound of a kind of angiotensin receptor antagonist and nep inhibitor
Method, including：
The pharmaceutical composition of angiotensin receptor antagonist and nep inhibitor is carried out using high performance liquid chromatography miscellaneous
Matter detaches；
Mobile phase in the high performance liquid chromatography includes water phase and organic phase；
The water phase includes phosphate buffer, trifluoroacetic acid solution or formic acid solution；
The organic phase includes organic mixture or acetonitrile, and the organic mixture includes acetonitrile and methanol；
The volume ratio of the water phase and organic phase is (45%~65%):(55%~35%).
Preferably, the molar concentration of the phosphate buffer is 25mmol/L~45mmol/L.
Preferably, the volumetric concentration of the trifluoroacetic acid solution is 0.05%~0.15%.
Preferably, the volumetric concentration of the formic acid solution is 0.05%~0.15%.
Preferably, the volume ratio of the acetonitrile in the organic mixture and methanol is (50~60)：(50~40).
Preferably, the volume ratio of the water phase in the high performance liquid chromatography and organic phase is (50%~60%)：(50%
~40%).
Preferably, the flow rate of mobile phase in the high performance liquid chromatography is 0.5mL/min~1.5mL/min.
Preferably, the column temperature in the high performance liquid chromatography is 25 DEG C~45 DEG C.
Preferably, the chromatography column packing in the high performance liquid chromatography is octadecylsilane chemically bonded silica or eight alkane
Base silane bonded silica gel.
Preferably, the Detection wavelength in the high performance liquid chromatography is 240nm~260nm.
The separation method of impurity is adopted in the compound of angiotensin receptor antagonist and nep inhibitor provided by the invention
Impurity separation is carried out with high performance liquid chromatography, the mobile phase selected during high performance liquid chromatography includes the water of special component
Mutually and organic phase, answering for angiotensin receptor antagonist and nep inhibitor can be effectively separated using this mobile phase
Close the plurality of impurities in object.The experimental results showed that the angiotensins obtained using impurity separation method provided by the invention by
The mass content ＜ 0.1% of single impurity in body antagonist and nep inhibitor compound sterling.
In addition, in the compound of angiotensin receptor antagonist provided by the invention and nep inhibitor impurity separation
Method and process is simple, favorable reproducibility, and this separation method is accurate and reliable.
Description of the drawings
In order to more clearly explain the embodiment of the invention or the technical proposal in the existing technology, to embodiment or will show below
There is attached drawing needed in technology description to be briefly described, it should be apparent that, the accompanying drawings in the following description is only this
The embodiment of invention for those of ordinary skill in the art without creative efforts, can also basis
The attached drawing of offer obtains other attached drawings.
Fig. 1 is the preparation of the compound of angiotensin receptor antagonist and nep inhibitor that the embodiment of the present invention 1 provides
The flow chart of technique；
Fig. 2 is the compound of angiotensin receptor antagonist and nep inhibitor that the embodiment of the present invention 1 is prepared
Nmr spectrum；
Fig. 3 is compound of the method that provides of the embodiment of the present invention 2 to angiotensin receptor antagonist and nep inhibitor
The chromatogram that middle impurity obtains after being detached；
Fig. 4 is compound of the method that provides of the embodiment of the present invention 3 to angiotensin receptor antagonist and nep inhibitor
The chromatogram that middle impurity obtains after being detached；
Fig. 5 is compound of the method that provides of the embodiment of the present invention 4 to angiotensin receptor antagonist and nep inhibitor
The chromatogram that middle impurity obtains after being detached；
Fig. 6 is compound of the method that provides of comparative example 1 of the present invention to angiotensin receptor antagonist and nep inhibitor
The chromatogram that middle impurity obtains after being detached.
Specific implementation mode
The technical scheme in the embodiments of the invention will be clearly and completely described below, it is clear that described implementation
Example is only a part of the embodiment of the present invention, instead of all the embodiments.Based on the embodiments of the present invention, this field is common
The every other embodiment that technical staff is obtained without making creative work belongs to the model that the present invention protects
It encloses.
The present invention provides the separation sides of impurity in the compound of a kind of angiotensin receptor antagonist and nep inhibitor
Method, including：
The pharmaceutical composition of angiotensin receptor antagonist and nep inhibitor is carried out using high performance liquid chromatography miscellaneous
Matter detaches；
Mobile phase in the high performance liquid chromatography includes water phase and organic phase；
The water phase includes phosphate buffer, trifluoroacetic acid solution or formic acid solution；
The organic phase includes organic mixture or acetonitrile, and the organic mixture includes acetonitrile and methanol；
The volume ratio of the water phase and organic phase is (45%~65%):(55%~35%).
The present invention is carried out the compound of angiotensin receptor antagonist and nep inhibitor using high performance liquid chromatography
Impurity detaches.In the present invention, the compound of the angiotensin receptor antagonist and nep inhibitor is [3- ((1S, 3R)-
1- biphenyl -4- ylmethyl -3- ethoxy carbonyl -1- butylcarbamoyls) -3 '-methyl -2 ' of propionic acid-(S)-(valeryl { 2 " -
(tetrazolium -5- bases) biphenyl -4 '-ylmethyl } amino) butyric acid] half pentahydrate of trisodium, there is structure shown in Formulas I：
The present invention does not have the source of the compound of the angiotensin receptor antagonist and nep inhibitor special limit
System, can by market buy obtain, can be also prepared by method well known to those skilled in the art, such as can according to application No. is
200680001733.0 Chinese patent " pharmaceutical combination product of the inhibitor of angiotensin receptor antagonist and NEP " institute is public
The method opened is prepared.In an embodiment of the present invention, the angiotensin receptor antagonist and nep inhibitor compound
Preparation method be：
It will be reacted with the compound of structure, Valsartan and sodium hydroxide shown in formula 1, obtain angiotensin receptor
The compound of antagonist and nep inhibitor：
In an embodiment of the present invention, the temperature of the compound of structure, Valsartan shown in the formula 1 and sodium hydroxide reaction
It is 20 DEG C~30 DEG C.In an embodiment of the present invention, the compound of structure, Valsartan shown in the formula 1 and sodium hydroxide reaction
Time be 10h.In an embodiment of the present invention, mole of the compound of structure, Valsartan and sodium hydroxide shown in the formula 1
Than being 1:1:3.In an embodiment of the present invention, the preparation method of the compound of structure shown in the formula 1 is：
It will be reacted with the compound of structure and hydrochloric acid shown in formula 2, obtain the compound with structure shown in formula 1：
In an embodiment of the present invention, the compound of structure shown in the formula 2 and the temperature of hydrochloric acid reaction are 20 DEG C~30
℃.In an embodiment of the present invention, the time of the compound of structure shown in the formula 2 and hydrochloric acid reaction is 2h.The present invention's
In embodiment, the compound of structure shown in the formula 2 and the molar ratio of hydrochloric acid ratio are 1:3.In an embodiment of the present invention, described
The preparation method of the compound of structure shown in formula 2 is：
It will be reacted with the compound of structure and calcium chloride shown in formula 3, obtain the chemical combination with structure shown in formula 2
Object：
In an embodiment of the present invention, the compound of structure shown in the formula 3 and calcium chloride reaction temperature be 60 DEG C~
70℃.In an embodiment of the present invention, the time of the compound of structure shown in the formula 3 and calcium chloride reaction is 2h.In this hair
In bright embodiment, the compound of structure and the molar ratio of calcium chloride shown in the formula 3 are 1:1.In an embodiment of the present invention,
The preparation method of the compound of structure shown in the formula 3 is：
It will in water be reacted with the compound of structure shown in formula 4 and sodium hydroxide, obtain the change of structure shown in formula 4
Close object：
In an embodiment of the present invention, the compound of structure shown in the formula 4 and the temperature of sodium hydroxide reaction are 20 DEG C
~30 DEG C.In an embodiment of the present invention, the time of the compound of structure shown in the formula 4 and sodium hydroxide reaction is 1h.
In the embodiment of the present invention, the compound of structure and the molar ratio of sodium hydroxide shown in the formula 4 are 1:2.The present invention is to 4 institute of formula
Show that the source of the compound of structure does not have special limitation, can be bought and be obtained by market, it also can be ripe according to those skilled in the art
The method known is prepared.
In an embodiment of the present invention, miscellaneous in the compound of the angiotensin receptor antagonist and nep inhibitor
Matter is one or more of formula a~formula o：
In the present invention, the high performance liquid chromatography, using high pressure transfusion system, will have not using liquid as mobile phase
The mobile phases such as the single solvent of same polarity or the mixed solvent of different proportion, buffer solution are pumped into the chromatographic column equipped with stationary phase,
It after each ingredient is detached in column, is detected into detector, to realize that the separation to sample is analyzed.
In the present invention, the mobile phase in the high performance liquid chromatography includes water phase and organic phase.In the reality of the present invention
It applies in example, the water phase includes phosphate buffer, trifluoroacetic acid solution or formic acid solution.In an embodiment of the present invention, institute
The molar concentration for stating phosphate buffer is 25mmol/L~45mmol/L；In other examples, the phosphate-buffered
The molar concentration of liquid is 30mmol/L~40mmol/L；In a further embodiment, the molar concentration of the phosphate buffer
For 32mmol/L~38mmol/L；In a further embodiment, the molar concentration of the phosphate buffer be 34mmol/L~
36mmol/L；In a further embodiment, the molar concentration of the phosphate buffer is 35mmol/L.
In an embodiment of the present invention, the volumetric concentration of the trifluoroacetic acid solution is 0.05%~0.15%；At other
Embodiment in, the volumetric concentration of the trifluoroacetic acid solution is 0.08%~0.12%；In a further embodiment, described three
The volumetric concentration of fluoroacetic acid solution is 0.1%.In an embodiment of the present invention, the trifluoroacetic acid solution is the water of trifluoroacetic acid
Solution.
In an embodiment of the present invention, the volumetric concentration of the formic acid solution is 0.05%~0.15%；Other real
It applies in example, the volumetric concentration of the formic acid solution is 0.08%~0.12%；In a further embodiment, the formic acid solution
Volumetric concentration is 0.1%.In an embodiment of the present invention, the formic acid solution is the aqueous solution of formic acid.
In the present invention, the organic phase includes organic mixture or acetonitrile, and the organic mixture includes acetonitrile and first
Alcohol.In an embodiment of the present invention, the volume ratio of the acetonitrile in the organic mixture and methanol is (50~60)：(50~
40)；In other examples, the volume ratio of the acetonitrile in the organic mixture and methanol is (52~58)：(48~42)；
In a further embodiment, the volume ratio of the acetonitrile in the organic mixture and methanol is (54~56)：(46~44)；Another
In outer embodiment, the volume ratio of acetonitrile and methanol in the organic mixture is 55：45.
In an embodiment of the present invention, the initial volume ratio of the water phase in the high performance liquid chromatography and organic phase is
(45%~65%)：(55%~35%)；In other examples, the water phase and organic phase in the high performance liquid chromatography
Initial volume ratio be (50%~60%)：(50%~40%)；In a further embodiment, in the high performance liquid chromatography
Water phase and organic phase initial volume ratio be (52%~58%)：(48%~42%)；In a further embodiment, the height
The initial volume ratio of water phase and organic phase in effect liquid phase chromatogram method is (54%~56%)：(46%~44%)；Other
In embodiment, the initial volume ratio of water phase and organic phase in the high performance liquid chromatography is 50：50.
In an embodiment of the present invention, the flow rate of mobile phase in the high performance liquid chromatography be 0.5mL/min~
1.5mL/min；In other examples, the flow rate of mobile phase in the high performance liquid chromatography be 0.8mL/min~
1.2mL/min；In a further embodiment, the flow rate of mobile phase in the high performance liquid chromatography is 1mL/min.
In an embodiment of the present invention, the method that gradient elution may be used makes mobile phase pass through chromatographic column, the gradient
The method of elution is specially：
In 0~35min, volume content of the water phase in mobile phase is reduced to 5%~15% by 45%~55%, organic phase
Volume content in mobile phase has 45%~55% to increase to 85%~95%；
In 35min~45min, volume content of the water phase in mobile phase keeps 5%~15%, and organic phase is in mobile phase
Volume content keep 85%~95%.
In other examples, the method for the gradient elution is specially：
In 0~35min, volume content of the water phase in mobile phase is reduced to 8%~12% by 48%~52%, organic phase
Volume content in mobile phase has 48%~52% to increase to 88%~92%；
In 35min~45min, volume content of the water phase in mobile phase keeps 8%~12%, and organic phase is in mobile phase
Volume content keep 88%~92%.
In a further embodiment, the method for the gradient elution is specially：
In 0~35min, volume content of the water phase in mobile phase is reduced to 10% by 50%, and organic phase is in mobile phase
Volume content there is 50% to increase to 90%；
In 35min~45min, volume content of the water phase in mobile phase keeps 10%, body of the organic phase in mobile phase
Product content keeps 90%.
In an embodiment of the present invention, the column temperature in the high performance liquid chromatography is 25 DEG C~45 DEG C；Other real
It applies in example, the column temperature in the high performance liquid chromatography is 30 DEG C~40 DEG C；In a further embodiment, the high-efficient liquid phase color
Column temperature in spectrometry is 35 DEG C.
In an embodiment of the present invention, the chromatography column packing in the high performance liquid chromatography is octadecylsilane key
Close silica gel or eight alkyl silane bonded silica gels；In other examples, the chromatographic column filling in the high performance liquid chromatography
Agent is octadecylsilane chemically bonded silica.
In an embodiment of the present invention, the Detection wavelength in the high performance liquid chromatography is 240nm~260nm；At it
In his embodiment, the Detection wavelength in the high performance liquid chromatography is 245nm~255nm；In a further embodiment, institute
It is 250nm to state the Detection wavelength in high performance liquid chromatography.
In an embodiment of the present invention, impurity in the compound of the angiotensin receptor antagonist and nep inhibitor
Separation method be：
The pharmaceutical composition of angiotensin receptor antagonist and nep inhibitor is carried out using high performance liquid chromatography miscellaneous
Matter detaches；
Mobile phase in the high performance liquid chromatography includes the phosphate buffer and body that molar concentration is 35mmol/L
Product is than being 55：45 acetonitrile and the mixed liquor of methanol；
The initial volume ratio of the mixed liquor of phosphate buffer and acetonitrile and methanol is 50 in the high performance liquid chromatography：
50；
Flow rate of mobile phase in the high performance liquid chromatography is 1mL/min；
Column temperature in the high performance liquid chromatography is 35 DEG C；
Chromatography column packing is octadecylsilane chemically bonded silica in the high performance liquid chromatography；
Detection wavelength in the high performance liquid chromatography is 250nm；
Mobile phase is set to pass through chromatographic column using the method for gradient elution, the method for the gradient elution is specially：
In 0~35min, volume content of the water phase in mobile phase is reduced to 10% by 50%, and organic phase is in mobile phase
Volume content there is 50% to increase to 90%；
In 35min~45min, volume content of the water phase in mobile phase keeps 10%, body of the organic phase in mobile phase
Product content keeps 90%.
Impurity is carried out to the compound of angiotensin receptor antagonist and nep inhibitor using method provided by the invention
Separation, the angiotensin receptor antagonist obtained after detaching using efficient liquid-phase chromatography method, test and nep inhibitor are answered
Close the impurity content in object sterling；Testing result is the angiotensin receptor that impurity separation method provided by the invention obtains
The mass content ＜ 0.1% of single impurity, vasotonia provided by the invention in the compound sterling of antagonist and nep inhibitor
The method that impurity detaches in the compound of hormone receptor antagonists and nep inhibitor has preferable effect.
According to《Chinese Pharmacopoeia》The 4th 0512 efficient liquid-phase chromatography method of general rule of version in 2015, test are provided by the invention
The reproducibility for the method that impurity detaches in the compound of angiotensin receptor antagonist and nep inhibitor, by counting impurity
Retention time reproducibility and peak area reproducibility result are it is found that angiotensin receptor antagonist provided by the invention and NEP suppressions
The method that impurity detaches in the compound of preparation has preferable reproducibility, method accurate and reliable.
Embodiment 1
The compound of angiotensin receptor antagonist and nep inhibitor is prepared according to flow chart shown in FIG. 1, Fig. 1 is
The flow chart of the preparation process of the compound of angiotensin receptor antagonist and nep inhibitor that the embodiment of the present invention 1 provides：
Compound with structure shown in formula 4 is commercial product.
There is the compound of structure and the sodium hydroxide of 1.9g shown in formula 4 to carry out 1 at 20~30 DEG C in water 10g
The reaction of hour, obtains the compound with structure shown in formula 3；
10g is subjected to 2 hours anti-with the compound of structure and the calcium chloride of 2.6g shown in formula 3 at 60~70 DEG C
It answers, obtains the compound with structure shown in formula 2；
15g is subjected to 2 hours anti-with the compound of structure and the 1N hydrochloric acid of 52g shown in formula 2 at 20~30 DEG C
It answers, obtains the compound with structure shown in formula 1；
By the sodium hydroxide with the compound of structure, the Valsartan of 10.6g and 2.9g shown in formula 1 of 10g at 20-30 DEG C
The lower reaction for carrying out 10 hours, obtains the compound of angiotensin receptor antagonist and nep inhibitor.
According to《Chinese Pharmacopoeia》The 4th 0441 NMR spectrum method of general rule of version in 2015 makes the embodiment of the present invention 1
The compound of standby obtained angiotensin receptor antagonist and nep inhibitor carries out structure detection, testing result such as Fig. 2 institutes
Show, Fig. 2 is the nuclear-magnetism of the compound of angiotensin receptor antagonist and nep inhibitor that the embodiment of the present invention 1 is prepared
Resonate spectrogram, and as shown in Figure 2, angiotensin receptor antagonist that the embodiment of the present invention 1 is prepared and nep inhibitor are answered
Closing object has structure shown in Formulas I.
Embodiment 2
The angiotensin receptor antagonist and NEP that the embodiment of the present invention 1 is prepared using high performance liquid chromatography
The compound of inhibitor carries out impurity separation, and the chromatography column packing in the high performance liquid chromatography is octadecylsilane key
Silica gel is closed, mobile phase includes the phosphate buffer (water phase) that molar concentration is 35mmol/L and volume ratio is 55：45 acetonitrile
With the mixed liquor (organic phase) of methanol, the initial volume ratio of phosphate buffer and the mixed liquor of acetonitrile and methanol is 50：50, stream
The flow velocity of dynamic phase is 1mL/min, and column temperature is 35 DEG C, Detection wavelength 250nm；Mobile phase is set to pass through using the method for gradient elution
The method of chromatographic column, the gradient elution is specially：In 0~35min, volume content of the water phase in mobile phase is reduced by 50%
To 10%, volume content of the organic phase in mobile phase has 50% to increase to 90%；In 35min~45min, water phase is in mobile phase
In volume content keep 10%, volume content of the organic phase in mobile phase keeps 90%.
After the embodiment of the present invention 2 carries out impurity separation to the compound of angiotensin receptor antagonist and nep inhibitor
Obtained chromatogram as shown in figure 3, the method that Fig. 3, which is the embodiment of the present invention 2, to be provided to angiotensin receptor antagonist and NEP
The chromatogram that impurity obtains after being detached in the compound of inhibitor, from the figure 3, it may be seen that the method that the embodiment of the present invention 2 provides
LC-Z09 impurity (the structure chemical combination shown in formula h in the compound of angiotensin receptor antagonist and nep inhibitor is detached
Object), LC-01 impurity (compound of structure shown in formula o), LC-Z01 impurity (compound of structure shown in formula a), LC-Z17 impurity
(compound of structure shown in formula m), LC-Z02 (compound of structure shown in formula b), the LC-Z03 (chemical combination of structure shown in formula c
Object), LC-Z18 (compound of structure shown in formula n), LC-Z15 impurity (compound of structure shown in formula k), LC-Z04 impurity (formulas
The compound of structure shown in d), LC-Z12 impurity (compound of structure shown in formula i), (change of structure shown in formula f of LC-Z06 impurity
Close object), LC-Z14 (compound of structure shown in formula j), LC-Z08 impurity (compound of structure shown in formula g), LC-Z16 impurity
(compound of structure shown in formula l), LC-Z05 impurity (compound of structure shown in formula e).
According to the method described in above-mentioned technical proposal, the angiotensin receptor antagonist and NEP that are obtained after test separation
Impurity content and yield in the compound sterling of inhibitor；Testing result is the impurity separation side that the embodiment of the present invention 2 provides
The mass content of single impurity is respectively less than in the compound sterling for the angiotensin receptor antagonist and nep inhibitor that method obtains
0.1%.
According to the method described in above-mentioned technical proposal, angiotensin receptor antagonist and NEP provided by the invention are tested
The reproducibility for the method that impurity detaches in the compound of inhibitor, test result is as shown in Table 1 and Table 2, and table 1 is that the present invention is implemented
The method impurity retention time test result that example 2 provides, table 2 are the survey for the method impurity peak area that the embodiment of the present invention 2 provides
Test result.
The method impurity retention time test result that 1 embodiment of the present invention 2 of table provides
The test result for the method impurity peak area that 2 embodiment of the present invention 2 of table provides
Note：In Tables 1 and 2, a~o is related impurities (impurity of structure shown in formula a~formula o)；Based on LC-03 and LC-2
Peak；SS is system suitability solution
By Tables 1 and 2 it is found that angiotensin receptor antagonist that the embodiment of the present invention 2 provides and nep inhibitor are answered
Closing the method that impurity detaches in object has preferable reproducibility, method accurate and reliable.
Embodiment 3
The angiotensin receptor antagonist and NEP that the embodiment of the present invention 1 is prepared using high performance liquid chromatography
The compound of inhibitor carries out impurity separation, and the chromatography column packing in the high performance liquid chromatography is octadecylsilane key
Silica gel is closed, mobile phase includes the trifluoroacetic acid aqueous solution (water phase) that mass concentration is 0.1% and mass ratio is 50：50 acetonitrile and
The initial volume ratio of the mixed liquor (organic phase) of methanol, trifluoroacetic acid aqueous solution and the mixed liquor of acetonitrile and methanol is 55：45, stream
The flow velocity of dynamic phase is 0.8mL/min, and column temperature is 25 DEG C, Detection wavelength 250nm；Flowing is set to communicate using the method for gradient elution
Chromatographic column is crossed, the method for the gradient elution is specially：In 0~35min, volume content of the water phase in mobile phase is by 55% drop
As low as 5%, volume content of the organic phase in mobile phase has 45% to increase to 95%；In 35min~45min, water phase is flowing
Volume content in phase keeps 5%, and volume content of the organic phase in mobile phase keeps 95%.
After the embodiment of the present invention 3 carries out impurity separation to the compound of angiotensin receptor antagonist and nep inhibitor
Obtained chromatogram as shown in figure 4, the method that Fig. 4, which is the embodiment of the present invention 3, to be provided to angiotensin receptor antagonist and NEP
The chromatogram that impurity obtains after being detached in the compound of inhibitor, as shown in Figure 4, the method that the embodiment of the present invention 3 provides
LC-Z09 impurity (the structure chemical combination shown in formula h in the compound of angiotensin receptor antagonist and nep inhibitor is detached
Object), LC-01 impurity (compound of structure shown in formula o), LC-Z01 impurity (compound of structure shown in formula a), LC-Z17 impurity
(compound of structure shown in formula m), LC-Z02 (compound of structure shown in formula b), the LC-Z03 (chemical combination of structure shown in formula c
Object), LC-Z18 (compound of structure shown in formula n), LC-Z15 impurity (compound of structure shown in formula k), LC-Z04 impurity (formulas
The compound of structure shown in d), LC-Z12 impurity (compound of structure shown in formula i), (change of structure shown in formula f of LC-Z06 impurity
Close object), LC-Z14 (compound of structure shown in formula j), LC-Z08 impurity (compound of structure shown in formula g), LC-Z16 impurity
(compound of structure shown in formula l), LC-Z05 impurity (compound of structure shown in formula e).
According to the method described in above-mentioned technical proposal, the angiotensin receptor antagonist and NEP that are obtained after test separation
Impurity content and yield in the compound sterling of inhibitor；Testing result is the impurity separation side that the embodiment of the present invention 3 provides
The mass content of single impurity is respectively less than in the compound sterling for the angiotensin receptor antagonist and nep inhibitor that method obtains
0.1%.
According to the method described in above-mentioned technical proposal, angiotensin receptor antagonist and NEP provided by the invention are tested
The reproducibility for the method that impurity detaches in the compound of inhibitor, test result is as shown in Table 3 and Table 4, and table 3 is that the present invention is implemented
The method impurity retention time test result that example 3 provides, table 4 are the survey for the method impurity peak area that the embodiment of the present invention 3 provides
Test result.
The method impurity retention time test result that 3 embodiment of the present invention 3 of table provides
The test result for the method impurity peak area that 4 embodiment of the present invention 3 of table provides
Note：A-o is related impurities；LC-03 and LC-2 is main peak；SS is system suitability solution
By table 3 and table 4 it is found that angiotensin receptor antagonist that the embodiment of the present invention 3 provides and nep inhibitor are answered
Closing the method that impurity detaches in object has preferable reproducibility, method accurate and reliable.
Embodiment 4
The angiotensin receptor antagonist and NEP that the embodiment of the present invention 1 is prepared using high performance liquid chromatography
The compound of inhibitor carries out impurity separation, and the chromatography column packing in the high performance liquid chromatography is octadecylsilane key
Close silica gel, mobile phase include mass concentration be 0.1% aqueous formic acid (water phase) and acetonitrile (mobile phase), aqueous formic acid with
The initial volume ratio of acetonitrile is 45：55, the flow velocity of mobile phase is 1.2mL/min, and column temperature is 30 DEG C, Detection wavelength 250nm；It adopts
Mobile phase is set to pass through chromatographic column with the method for gradient elution, the method for the gradient elution is specially：In 0~35min, water phase exists
Volume content in mobile phase is reduced to 15% by 45%, and volume content of the organic phase in mobile phase has 55% to increase to
85%；In 35min~45min, volume content of the water phase in mobile phase keeps 15%, and volume of the organic phase in mobile phase contains
Amount keeps 85%.
After the embodiment of the present invention 4 carries out impurity separation to the compound of angiotensin receptor antagonist and nep inhibitor
Obtained chromatogram as shown in figure 5, the method that Fig. 5, which is the embodiment of the present invention 4, to be provided to angiotensin receptor antagonist and NEP
The chromatogram that impurity obtains after being detached in the compound of inhibitor, as shown in Figure 5, the method that the embodiment of the present invention 4 provides
LC-Z09 impurity (the structure chemical combination shown in formula h in the compound of angiotensin receptor antagonist and nep inhibitor is detached
Object), LC-01 impurity (compound of structure shown in formula o), LC-Z01 impurity (compound of structure shown in formula a), LC-Z17 impurity
(compound of structure shown in formula m), LC-Z02 (compound of structure shown in formula b), the LC-Z03 (chemical combination of structure shown in formula c
Object), LC-Z18 (compound of structure shown in formula n), LC-Z15 impurity (compound of structure shown in formula k), LC-Z04 impurity (formulas
The compound of structure shown in d), LC-Z12 impurity (compound of structure shown in formula i), (change of structure shown in formula f of LC-Z06 impurity
Close object), LC-Z14 (compound of structure shown in formula j), LC-Z08 impurity (compound of structure shown in formula g), LC-Z16 impurity
(compound of structure shown in formula l), LC-Z05 impurity (compound of structure shown in formula e).
According to the method described in above-mentioned technical proposal, the angiotensin receptor antagonist and NEP that are obtained after test separation
Impurity content and yield in the compound sterling of inhibitor；Testing result is the impurity separation side that the embodiment of the present invention 4 provides
The mass content of single impurity is respectively less than in the compound sterling for the angiotensin receptor antagonist and nep inhibitor that method obtains
0.1%.
According to the method described in above-mentioned technical proposal, angiotensin receptor antagonist and NEP provided by the invention are tested
The reproducibility for the method that impurity detaches in the compound of inhibitor, as shown in table 5 and table 6, table 5 is that the present invention is implemented to test result
The method impurity retention time test result that example 4 provides, table 6 are the survey for the method impurity peak area that the embodiment of the present invention 4 provides
Test result.
The method impurity retention time test result that 5 embodiment of the present invention 4 of table provides
The test result for the method impurity peak area that 6 embodiment of the present invention 4 of table provides
Note：A-o is related impurities；LC-03 and LC-2 is main peak；SS is system suitability solution
By table 5 and table 6 it is found that angiotensin receptor antagonist that the embodiment of the present invention 5 provides and nep inhibitor are answered
Closing the method that impurity detaches in object has preferable reproducibility, method accurate and reliable.
Comparative example 1
The angiotensin receptor antagonist and NEP that the embodiment of the present invention 1 is prepared using high performance liquid chromatography
The compound of inhibitor carries out impurity separation, and the chromatography column packing in the high performance liquid chromatography is octadecylsilane key
Silica gel is closed, mobile phase includes the glacial acetic acid aqueous solution (water phase) and acetonitrile (organic phase) that mass concentration is 0.1%, 0.1% ice
The initial volume of aqueous acetic acid and acetonitrile ratio is 45：55, the flow velocity of mobile phase is 1.0mL/min, and column temperature is 35 DEG C, detects wave
A length of 250nm；Mobile phase is set to pass through chromatographic column using the method for gradient elution, the method for the gradient elution is specially：0~
35min, volume content of the water phase in mobile phase are reduced to 10% by 45%, and volume content of the organic phase in mobile phase has
55% increases to 90%；In 35min~45min, volume content of the water phase in mobile phase keeps 10%, and organic phase is in mobile phase
In volume content keep 90%.
After comparative example 1 of the present invention carries out impurity separation to the compound of angiotensin receptor antagonist and nep inhibitor
Obtained chromatogram as shown in fig. 6, the method that Fig. 6, which is comparative example 1 of the present invention, to be provided to angiotensin receptor antagonist and NEP
The chromatogram that impurity obtains after being detached in the compound of inhibitor, it will be appreciated from fig. 6 that the method that comparative example of the present invention 1 provides
Impurity in the compound of separating blood vessel angiotensin receptor antagonist and nep inhibitor, impurity cannot reach baseline point with main peak
From separating effect is poor.
As seen from the above embodiment, answering the present invention provides a kind of angiotensin receptor antagonist and nep inhibitor
The separation method of impurity in object is closed, including：Using high performance liquid chromatography to angiotensin receptor antagonist and nep inhibitor
Pharmaceutical composition carry out impurity separation；Mobile phase in the high performance liquid chromatography includes water phase and organic phase；The water
Include mutually phosphate buffer, trifluoroacetic acid solution or formic acid solution；The organic phase includes organic mixture or acetonitrile, described
Organic mixture includes acetonitrile and methanol；The volume ratio of the water phase and organic phase is (45%~65%)：(55%~35%).
The present invention carries out impurity separation using high performance liquid chromatography, and the mobile phase selected during high performance liquid chromatography includes specific
The water phase and organic phase of ingredient can be effectively separated angiotensin receptor antagonist using this mobile phase and NEP press down
Plurality of impurities in the compound of preparation.
Claims (10)
1. the separation method of impurity in the compound of a kind of angiotensin receptor antagonist and nep inhibitor, including：
Impurity point is carried out to the pharmaceutical composition of angiotensin receptor antagonist and nep inhibitor using high performance liquid chromatography
From；
Mobile phase in the high performance liquid chromatography includes water phase and organic phase；
The water phase includes phosphate buffer, trifluoroacetic acid solution or formic acid solution；
The organic phase includes organic mixture or acetonitrile, and the organic mixture includes acetonitrile and methanol；
The volume ratio of the water phase and organic phase is (45%~65%):(55%~35%)；
The compound of the angiotensin receptor antagonist and nep inhibitor is [3- ((1S, 3R) -1- biphenyl -4- ylmethyls -
3- ethoxy carbonyl -1- butylcarbamoyls) -3 '-methyl -2 ' of propionic acid-(S)-(valeryl { 2 "-(tetrazolium -5- bases) biphenyl -
4 '-ylmethyls } amino) butyric acid] half pentahydrate of trisodium, there is structure shown in Formulas I：
2. separation method according to claim 1, which is characterized in that the molar concentration of the phosphate buffer is
25mmol/L~45mmol/L.
3. separation method according to claim 1, which is characterized in that the volumetric concentration of the trifluoroacetic acid solution is
0.05%~0.15%.
4. separation method according to claim 1, which is characterized in that the volumetric concentration of the formic acid solution be 0.05%~
0.15%.
5. separation method according to claim 1, which is characterized in that the body of acetonitrile and methanol in the organic mixture
Product is than being (50~60)：(50~40).
6. separation method according to claim 1, which is characterized in that water phase in the high performance liquid chromatography and organic
The volume ratio of phase is (50%~60%)：(50%~40%).
7. separation method according to claim 1, which is characterized in that the flow rate of mobile phase in the high performance liquid chromatography
For 0.5mL/min~1.5mL/min.
8. separation method according to claim 1, which is characterized in that the column temperature in the high performance liquid chromatography is 25 DEG C
~45 DEG C.
9. separation method according to claim 1, which is characterized in that the chromatographic column filling in the high performance liquid chromatography
Agent is octadecylsilane chemically bonded silica or eight alkyl silane bonded silica gels.
10. separation method according to claim 1, which is characterized in that the Detection wavelength in the high performance liquid chromatography
For 240nm~260nm.
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