Environmentally Benign Solventless Preparation Of Bonded Silica Chromatographic Packings

Pharmaceutical analysis plays an important role in new drug research because it’s related to the drug quality and safety.High performance liquid chromatography（HPLC）has been widely used to separate and analysize drugs due to its high sensitivity and selectivity.In HPLC system,chromatographic column is a key factor to determine the separation performance.Chromatographic packing is the“heart”of column.Therefore,it requires good physical and chemical stability,regular and uniform spherical and suitable pore structure.Silica is an inorganic matrix,which exhibits superior chemical stability and mechanical strength.Besides,silanol groups on silica surface can interact with the functional groups to prepare stationary phases.At present,there are mainly three modification methods to prepare bonded silica stationary phases,including organic solvent method,supercritical fluid method and vapor deposition method.Organic solvent method is an traditional method,but it’s harmful to both human health and environment.Although supercritical fluid method can obtain higher bonded density,it needs special facilities with high cost and is inconvenienced to operate.Vapor deposition method refers to the direct reaction between silica and silane reagents at high temperature,which doesn’t involve any liquid solvent.However,it still has some limitations for high boiling point silane.Reversed phase liquid chromatography（RPLC）is widely used separation technique owning to its high efficiency,good reproducibility and compatibility with MS.Octadecyl bonded silica stationary phase(C₁₈)is the most commonly used in RPLC.C₁₈8 commercial column is usually prepared by organic solvent method.Some researchers have used impregnation method to prepare silane film on silica or glass surface in the field of biosensor and DNA microarray,which can obtain neat surface structure.The impregnation method can adsorb silane on silica surface at room temperature.Based on impregnation method,the first aim of this work is to seek a method to prepare C₁₈8 phase by avoiding organic solvent.More and more complex compounds with different polarity are continuously prepared in new drugs research.Therefore,it’s difficult to separate these compounds only with RPLC column.In order to expand the selectivity of chromatographic separation,a mixed-mode chromatography（MMC）has been developed.RPLC is suitable for separating non-polar or weakly polar substances.Hydrophilic interaction liquid chromatography（HILIC）can separate highly polar and hydrophilic compounds.Hence,RPLC/HILIC is a good combination MMC.Cyano group has polarity and dipole-dipole interactions,which has special selectivity and short equilibration time in comparison with silica column.However,commercial cyano columns,which are prepared by organic solvent method,are not widely used due to their poor stability.Thiol-ene click chemistry has developed rapidly in stationary phases preparation due to its high efficiency,high selectivity,mild reaction conditions and it is environmentally friendly method.Therefore,the second aim of this work is to develop a method to prepare RPLC/HILIC cyano packing with higher stability.The results of Tanaka tests have shown that synthesized cyano column not only exhibits MMC chromatographic behavior but also exhibits high shape selectivity.This may be related to properties of mobile phase,separation substances and stationary phase.Linear solvation energy relationships（LSERs）have been widely used to evaluate retention mechanisms of stationary phases.In addition,multiple sites model has been widely used to investigate the effect of residual silanols.Hence,the third aim of this work is to evaluate the synthesized cyano column and reveal the separation mechanism by Tanaka tests,LSERs and multiple sites model.In summary,this thesis proposes a solventless method to prepare C₁₈8 reversed phase and cyano-bonded RPLC/HILIC stationary phase.Solventless method not only has the advantage of environmentally benign,but also increases the bonded density of chromatographic packings.Therefore,this paper carries out three research contents in order to solve above problems.The first part of research is to prepare C₁₈8 reversed phase by impregnation method and vapor deposition method,optimize the preparation conditions and study the chromatographic properties and reproducibility.The second part has prepared cyano-based RPLC/HILIC stationary phase by vapor deposition method and thiol-ene click chemistry reaction.The results have been shown that synthesized cyano column has RPLC/HILIC dual chromatographic properties and higher hydrothermal stability.The third part is the retention mechanism of prepared cyano packing,which has been further revealed its mechanism affected by-S-,CN-and silanol groups.