Application Of Microemulsion Liquid Chromatography To Several Proprietary Chinese Medicines And Their Active Ingredients

Microemulsion liquid chromatography is a newly developed chromatographic technique that uses microemulsions as mobile phases for separation and detection.Microemulsions are usually classified as oil-in-water,water-in-oil and bicontinuous.Oil-in-water and water-in-oil microemulsions are usually used in re versed-phase liquid chromatography,among which oil-in-water microemulsions have certain advantages as reversed-phase liquid chromatography.Chinese medicine,as a drug used under the theory of traditional Chinese medicine,is generally derived from natural plants and their processed products,and has a long history of use in China.Due to the complexity of plant active ingredients,problems such as excessive interference and long separation time are encountered in the process of analysis and separation.With the increase of people’s awareness of health care,herbal medicines of the same origin have become the health care plants used by people on a daily basis,so the control of active ingredients of plants has become a topic of concern.In addition,the green separation mode is also gradually becoming the direction that people pay attention to.Microemulsions in which the aqueous phase occupies a large proportion and the proportion of organic solvents is small are in line with the concept of green and environmentally friendly analysis.The purpose of this study is to solve the problems that traditional high performance liquid chromatography requires complicated pre-treatment or setting up gradient procedures when separating substances with large differences in hydrophobicity,and a large amount of organic solvents are used in the analysis process,which damages the health of laboratory personnel as well as the environment.A low-cost,simple,rapid,and green microemulsion chromatographic method was developed for the analysis of complex components in red clover,angelica tonic blood soup,angelica tonic blood oral liquid,and si wu tang.The main research works are as follows:(1)A method was developed for the simultaneous separation of five isoflavones(formononetin,ononin,calycosin,calycosin-7-glucoside,and biochanin A)from red clover in 10 min using microemulsion liquid chromatography.Stable and clear microemulsions were prepared by studying the types and concentrations of surfactants,co-surfactants,and oil phases.The mobile phase pH and column temperature were studied to observe their effects on the separation and analysis,and then the optimal experimental conditions were obtained.The possible separation mechanism of the five isoflavones in red clover by microemulsion liquid chromatography was explored and predicted,and the actual separation results were consistent with the predicted results.The method established in this study is simple,low-cost,green and efficient,and provides an optional method for the rapid separation of red clover isoflavones in less than 10 min without the need to set up a complicated gradient method.(2)Microemulsion liquid chromatography was successfully applied to the separation and detection of six components(ferulic acid,chlorogenic acid,calycosin,calycosin-7-glucoside,ononin and biochanin A)in two herbs(Angelica sinensis and Milkvetch Root)of danggui buxue oral liquid.The types and concentrations of surfactant and co-surfactant as well as the oil phase were examined.The experimental results showed that the best separation conditions for Angelica sinensis Blood Tonic Oral Liquid were 3.5%SDS(w/v),10%n-butanol(v/v),1.0%ethyl acetate(v/v)and 85.5%water(v/v)with pH adjusted to 3.0 by phosphoric acid.The developed method has the advantage of simple,fast,green and efficient.Meanwhile,the six components(ferulic acid,chlorogenic acid,mangosteen,chickpea sproutin A,mullein isoflavone glucoside and mullein isoflavone)in Angelica tonic blood soup were separated under the experimental conditions of 2.5%SDS(w/v),9%n-butanol(v/v),0.9%ethyl acetate(v/v)and 87.6%water(v/v),and the six components in Angelica tonic blood soup were separated rapidly within 12 min without the need to set up a gradient program.(3)A microemulsion phase chromatographic method was developed for the separation of five components(paeoniflorin,acteoside,chlorogenic acid,ferulic acid and ligustilide)from four herbs(Angelica sinensis,Rhizoma chuanxiong,Radix paeoniae alba and Radix rehmanniae preparata)in Siwu Decoction.The types and concentrations of surfactant,co-surfactant and oil phase were investigated,The influence of mobile phase pH,column in temperature and many other chromatographic factors on the analysis of retention was also evaluated,and the most optimal separations conditions were achieved.The possible separation mechanism of the five components in Siwu Decoction was investigated,and the peak order was predicted.The actual results obtained eventually matched with the predicted results and were consistent with the peak emergence pattern of liquid chromatography.This study established a low-cost,environmentally friendly and simple analytical method that can be successfully applied to the determination of the quality of si wu tang.In addition,microemulsion liquid chromatography still has great potential in the analysis of Chinese medicine components.