The Design,Preparation And Application Of High-performance Boronate Affinity Stationary Phase

Cis-diol molecules,such as carbohydrates,nucleosides and glycoproteins have demonstrated great biological significance in glycomics,metabolomics and proteomics.Many of them have been considered as biomarkers for various cancers.However,cis-diol molecules in real samples usually exist in very low abundance along with abundant interfering components.Therefore,the enrichment and separation of targeting cis-diol molecules are critical steps for analysis of these biomolecules.Boronate affinity materials which can form reversible and p H-dependent cyclic esters with cis-diol molecules have been proved to be a facile and efficient way for selective capture and enrichment of cis-diol molecules.Herein,We designed and prepared high-performance boronate affinity stationary phases and studied the separation and enrichment of them.And the research contents are as follows:?1?Developing a facile and efficient way for selective capture and enrichment of cis-diol molecules with high performance is greatly important.In the first work,we modified silica surface with GMA polymer brushes through the SI-ATRP,expanding active binding sites for grafting and improving the binding capacity of boronate affinity materials.The 3-?dimethylaminomethyl?aniline-4-boronic acid was immobilized by the ring-opening reaction of the amino-functionalized Wulff-type boronate acid ligand and the epoxy.And the formation of the hydroxyl groups improved the hydrophilic interaction of boronate affinity materials and reduced the hydrophobic interaction and nonspecific adsorption of cis-diol molecules.FT-IR and elemental analysis was carried out to confirm the successful modification of GMA polymer brushes and 3-?dimethylaminomethyl?aniline-4-boronic acid on the SiO₂.Meanwhile,the pK_a value of Wulff-PBA is 6.7,which lead to the low binding pH??5.5?.And this materals can adsorb cis-diol molecules under acidic condition with high binding capacity for complex samples.?2?Most of existing boronate affinity materials only can achieve the separation of cis-diol and non cis-diol molecules.The lack of second separation for enriching cis-diol molecules,often requires additional separate methods,such as RPLC or HILIC,which increases the analytical steps,and adversely affects the results of the analysis.So the development of boronate affinity materials with secondary separation capability in single column is significant,which can reduce the errors in the replacement of chromatographic columns and the transfer of samples.In the second work,the mixed-mode stationary phase modified with 3-?dimethylaminomethyl?aniline-4-boronic acid and C12 group was synthesized.And FT-IR and elemental analysis was carried out to confirm the successful preparation of stationary phase.The3-?dimethylaminomethyl?aniline-4-boronic acid was modified on the surface of SiO₂and can specifically binds cis-diol molecules.In addition,the C12 groups and amine groups endowed the stationary phase with reversed-phase?RP?and anion-exchange site?AE?,respectively.Alkylbenzenes,nucleosides,polycyclic aromatic hydrocarbons?PAHs?,anilines,sulfonamides and phenols were chromatographed on the column in RP-mode,while inorganic anions were chromatographed in AE-mode.Meanwhile the off-line 2D-LC,for selective capture and separation of urinary nucleosides by switching mobile phases was successfully realized on a single column.?3?Binding pH is a critical factor for boronate affinity.In the third work,4-carbonyl-benzoboroxole was designed and synthesized,and NMR was carried out to conform the successful synthesis of them.It was then combined with the modification effect of polyethyleneimine?PEI?for the preparation of boronate affinity stationary phase.Multitudinous amine groups of PEI might provide enough active binding sites for grafting,which can greatly improve the binding capacity of the materals.And the unreacted amine groups can improve the hydrophilic interaction of boronate affinity materials and reduce the nonspecific adsorption of cis-diol molecules.FT-IR,elemental analysis,SEM and zeta potentials was carried out to confirm the successful modification of PEI and 4-carbonyl-benzoboroxole on the SiO₂.The column showed excellent selectivity,high binding capacity and lowest binding pH.In addition,the column also showed secondary separation capability under acidic conditions and realized separate cis-diol molecules on a single column.