Controlled synthesis of superhydrophilic flower-like hierarchical porous diboronate affinity materials for capturing biomarkers

Abstract

Background

Boronate affinity chromatography represents a powerful analytical technique for the selective separation and enrichment of biomolecules containing cis-diol moieties, including carbohydrates, glycoproteins, and other cis-dihydroxy compounds. While boronate affinity materials (BAMs) have shown promise in glycosylation-based separation and analysis, their practical application is hindered by non-biocompatible binding pH, low enrichment efficiency for low-abundance samples, non-specific adsorption, and limited loading capacity. To address these limitations, this work focuses on developing flower-like hierarchical porous diboronate affinity materials (FHP-DBAMs) with enhanced binding strength, selectivity, and capacity for cis-diol-containing biomolecules.

Results

FHP-DBAM was synthesized via a facile sol-gel method, using tetrahydroxydiboron as a hydrophilic diboronic acid monomer. The electron-withdrawing nature and hydrophilicity of diboronate affinity mechanism enable FHP-DBAM to operate at lower pH values (pH ≥ 5), addressing the biocompatibility issue. DFT and experiment calculations confirm the enhanced cis-diol binding affinity of diboronate affinity mechanism compared with monoboronate affinity mechanism, resulting in a remarkably low dissociation constant (DFT Kd = 6.74 × 10⁻⁵ M, experiment Kd = 9.95 × 10⁻⁵ M) for FHP-DBAM. Furthermore, the unique flower-like hierarchical porous structure provides a high surface area and nanoconfinement effect, significantly boosting target molecule loading capacity and affinity reaction kinetics. Compared to traditional BAMs, FHP-DBAM exhibits over ten times higher loading capacity. As a proof-of-concept, FHP-DBAM successfully captures the biomarker GM₁ in breast cancer cells MCF-7 with high efficiency.

Significance and novelty

This work introduces diboronate affinity mechanism and flower-like hierarchical porous structure as new solution to overcome the limitations of conventional BAMs. FHP-DBAMs achieve lower binding pH, enhanced selectivity, and stronger binding stability through diboronate affinity mechanism. The unique flower-like porous structure maximizes surface area and active sites, addressing low enrichment efficiency and loading capacity. These advancements are critical for the efficient and biocompatible separation of cis-diol-containing biomolecules.