Preparation And Evaluation Of Urushiol Methacrylate Bonded SiO₂ Stationary Phase For HPLC

Objective As a cultural treasure of China,Traditional Chinese medicines had made great contributions to the survival and health of human beings.However,the problems of the complex chemical composition,unclear mechanism of medicinal action and low purification efficiency in traditional Chinese medicines have limited the further development of themselves.Therefore it is necessary to establish an efficient,simple and feasible method for the separation and purification.High performance liquid chromatography,as a technique for the separation and detection of various components,has been widely used in various analytical fields.Stationary phase as the core of chromatographic separation techniques,its preparation and application have become the research focus and received much attention in recent years of chromatographic research.Among them,the spherical porous silica gel has become the most widely used chromatographic packing carrier because of its large specific surface area,easy modification and high mechanical strength.Based on the above research background,a novel chromatographic stationary phase was prepared and used for the separation and purification of the main active components of traditional Chinese medicine in this study.Methods(1)The mixed urushiol methacrylata was prepared using urushiol and methacryloyl chloride via asubstitution reaction and then coated onto the surface of spherical silica by physical adsorption.The spherical silica was chemically modified with3-methacryloyloxypropyl trimethoxysilane.Then,the urushiol methacrylate bonded silica stationary phase was synthesized via the surface radical polymerization of urshiol methacrylate and the pendant vinyl groups onto the surface of the spherical silica.The stationary phase was characterized by FT-IR,TGA,SEM,TEM and EA.(2)The stationary phases were packed in statinless steel hollow column by the slurry packing method.The stability of the column was evaluated by the peak shape and retention time of the alkylbenzene analyzers.The separation characteristics of chromatographic column were evaluated by Tanaka method.A mixture of alkylbenzene and PAHs were used as probes to evaluate the separation performance and separation mechanism of the column,and compared with the conventional bonded silica gel column C18.(3)The urushiol methacrylate bonded silica column was used for the separation and purification of three active components of traditional Chinese medicine,Gastrodia elata,Evodia rutaecarpa and Heather tree fruit.The separation mechanism of three active components of traditional Chinese medicines on chromatographic column were studied by using Van’t Hoff equation.Results(1)The results revealed that the urushiol methacrylate was successfully immobilized on the spherical silica surface after the surface polymerization reaction.These stationary phases had excellent monodispersity and low toxicity,and the particle size distribution is 2-10μm,the pore size was 6.46 nm,the specific surface is 321.7m~2/g.The stationary phase began to decomposed at 200℃,which showed that it has good thermal stability.(2)The column exhibits typical reversed-phase liquid chromatography performance,and has good fluidity and stability.Compared with C18 column,this column can realize the separation and detection of alkylbenzene and PAHs in a shorter time under the condition of mobile phase with low organic solvent ratio and low flow rate,and the column has good shape selectivity.Separation of PAHs is a spontaneous process,which was driven by enthalpy.(3)After separation and purification of the gastrodin elata extract by column,the purity of gastrodin increased from 17.06%to 93.45%.The separation of the gastrodin elata extract by column was a spontaneous process,which was driven by entropy and enthalpy.The purity of evodiamine increased from 8.46%to 88.12%.The separation of frutus evodiae extract was a spontaneous process,which is driven by enthalpy.And the purity of camptothecin increased from 42.13%to 93.72%.The separation of camptotheca acuminata extract was a spontaneous process,which was driven by both entropy and enthalpy.Conclusion In this paper,a novel stationary phase was prepared using urushiol,which was a green renewable resource,as a raw material.The stationary phase shows a good separation and purification effect on the three active ingredients of gastrodin,evodiamine and camptothecin.Compared with the C18 column,the separation conditions of the urushiol methacrylate bonded silica was safer and more environmentally friendly.This study provides not only a new method for the separation and purification of gastrodin,evodiamine and camptothecin in real samples,but also a new strategy for the preparation of chromatographic stationary phases.It expanded the application of raw lacquer in chromatographic separation materials.