Preparation Of Vancomycin Chiral Stationary Phase Chromatography Packing And Application In The Separation For Drugs

Up to now,the separation and separation of chiral drugs is still a topic worthy of study.In this paper,the 5 μm silica gel purchased from Lanzhou Institute of Chemical Physics and the uniform particle size,monodisperse polymethacrylic resin microspheres(Poly(GMA-co-EDMA))prepared by the research group were used as the matrix.Vancomycin,as a chiral selector,was used for the first time to bond the chiral selector via surface-initiated atom transfer radical polymerization(SI-ATRP),which is different from the traditional bonding method,and successfully prepared three macrocyclic glycopeptide chiral stationary phases(CSP).The analytical method of high performance liquid chromatography(HPLC)was used to test the resolution of the synthesized stationary phase.In order to detect the performance of CSP,we optimized the mobile phase conditions(temperature,flow rate,p H,content of organic phase and content of TEA).The commercialized macrocyclic glycopeptide chiral stationary phase once again proves that vancomycin as a chiral selector has broad application prospects in the field of chiral separation.The contents of this paper are as follows:1.Preparation of vancomycin chiral stationary phase via SI-ATRPThe chromatographic technique based on CSP has been explored for enantioseparation.Herein,poly(glycidyl methacrylate)(poly(GMA))brushes were grafted on the surface of silica gel via SI-ATRP,followed by introduction of vancomycin as chiral selector.The as-synthesized material was characterized by elemental analysis,scanning electron microscopy(SEM),Fourier transform infrared(FT-IR)and thermogravimetric analysis(TGA)proving the formation of vancomycin-immobilized brush.Then the resulting CSP was explored to separate 7racemic drugs(bicalutamide,1-benzyl-5-phenylbarbituric acid,chlorpheniramine maleate,fluoxetine hydrochloride,verapamil hydrochloride,benzoxazocine hydrochloride and isoprenaline hydrochloride)in HPLC.Several factors affecting the enantioseparation performance of vancomycin-immobilized CSP were optimized.Under the optimal conditions,baseline separation of fluoxetine hydrochloride(Rs=2.52)was achieved,which was better than that on commericial Chirobiotic V column,while partly enantioseparation of bicalutamide(Rs=1.01),chlorpheniramine maleate(Rs=0.77),1-benzyl-5-phenylbarbituric acid(Rs=0.67),isoprenaline hydrochloride(Rs=0.73),verapamil hydrochlorid(Rs=0.91)and benzoxazocine hydrochloride(Rs=1.03)were acquired.It was concluded that SI-ATRP is a robust way to fabricate vancomycin-based CSP for enantioseparation.2.Comparison of resolution performance of vancomycin chiral stationary phase and its derivativesVancomycin is a highly efficient glycopeptide antibiotic and a chiral separation material with excellent performance,whose structure contains multiple chiral sites.The change of structure can alter the binding performance between CSP and chiral drugs and transform the chiral resolution ability.Herein,vancomycin-immobilized CSP(V-CSP)and 3,5-dimethylphenyl isocyanate(DMP)derivative CSP(D-V-CSP)were prepared respectively.Both V-CSP and D-V-CSP were characterized through elemental analysis,FT-IR,TGA and SEM,and then explored to separate chiral drugs HPLC.After further optimizing the chromatographic separation conditions such as temperature,p H value,organic phase and flow rate,the enantioseparation of fluoxetine,labetalol,verapami,rosiglitazone,mirtazapine and chlorphenamine maleate were obtained.Through change of the structure for vancomycin to make CSP exhibited different chiral recognition ability.Partial separation of labetalol(Rs=0.682)and mirtazapine(Rs=0.913)was achieved on V-CSP,which was better than that on D-V-CSP.Compared with the stationary phase itself,the derivatized stationary phase will have strong hydrophobic and cavity interactions with the chiral drug,therefore,on D-V-CSP chiral columns,baseline separation of fluoxetine(Rs=2.149),rosiglitazone(Rs=2.050)and chlorpheniramine maleate(Rs=1.815)and partial separation of verapamil(Rs=0.946)were achieved.Based on the above results,both V-CSP and D-V-CSP would successfully be applied in the separation of chiral drugs,and changing the structure of the stationary phase is beneficial to the improvement of chiral recognition ability for specific chiral drugs.3.Preparation and evaluation of chiral stationary phase based on macroporous adsorption resin vancomycinAt present,most of CSP that have been commercialized on the market are based on silica gel with better rigidity.We used polymethacrylic resin microspheres Poly(GMA-co-EDMA)with uniform particle size prepared in the laboratory as the stationary phase matrix.The vancomycin chiral stationary phase(Poly(GMA-co-EDMA)-Vancomycin)was successfully prepared by grafting the vancomycin chiral selective agent on the surface of Poly(GMA-co-EDMA)through the spacer arm.The experimentally obtained fillers were subjected to elemental analysis,SEM,TGA and FT-IR of Poly(GMA-co-EDMA)-Vancomycin material.The final synthesized filler was used as the stationary phase of liquid chromatography,and then the separation performance of the stationary phase was detected and explored by HPLC.In the process of this experiment,we selected five chiral drugs as the substances to be separated and detected.The five chiral drugs were labetalol,rosiglitazone,mirtazapine,nefopam and propranolol.Chromatographic conditions are an important factor affecting the performance of stationary phase separation.By changing the temperature,flow rate,and ratio of organic phase in the mobile phase during the experiment.Because the experiment uses triethylamine salt buffer as the mobile phase,the addition ratio of TEA in the buffer salt solution(TEAA)will also affect the resolution of the stationary phase.The experimental results showed that the resolutions of labetalol(Rs=0.68)and mirtazapine(Rs=0.76)were both <1.The resolution for rosiglitazone(Rs=1.51),nefopam(Rs=1.34)and propranolol(Rs=1.14)were >1.And rosiglitazone was obtained baseline separation in Poly(GMA-co-EDMA)-Vancomycin.Based on the above conclusions,we can prepare polymer stationary phases different from traditional stationary phases for the resolution of chiral drugs.