Covalent organic frameworks (COFs) represent a new class of porous crystalline polymers with a diversity of applications. However, synthesis of uniform spherical COFs poses a great challenge. Here, we present size-controllable synthesis of uniform spherical COFs from nanometer to micrometer scale by a facile approach at room temperature. The as-prepared spherical COFs with different sizes exhibited ultrahigh surface area, good crystallinity, and chemical/thermal stability. Multifarious microscopic and spectroscopic techniques were performed to understand the formation mechanism and influencing factors of the spherical COFs. Moreover, the general applicability for room-temperature synthesis of the spherical COFs was demonstrated by varying different building blocks. Spherical COFs, because of the advantageous nature of their surface area, hydrophobicity, and mesoporous microenvironment, serve as an attractive restricted-access adsorption material for highly selective and efficient enrichment of hydrophobic peptides and size exclusion of macromolecular proteins simultaneously. On this basis, the spherical COFs were successfully applied to the specific capture of ultratrace C-peptide from human serum and urine samples. This research provides a new strategy for room-temperature controllable synthesis of uniform spherical COFs with different sizes and extends the application of COFs as an attractive sample-enrichment probe for clinical analysis.

This study reports a facile approach for the synthesis of horseradish peroxidise (HRP)-inorganic hybrid nanoflowers by self-assembly of HRP and copper phosphate (Cu3(PO4)2·3H2O) in aqueous solution. Several reaction parameters that affect the formation of the hybrid nanoflowers were investigated and a hierarchical flowerlike spherical structure with hundreds of nanopetals was obtained under the optimum synthetic conditions. The enzymatic activity of HRP embedded in hybrid naonflowers was evaluated based on the principle of HRP catalyzing the oxidation of o-phenylenediamine (OPD) in the presence of hydrogen peroxide (H2O2). The results showed that 506% enhancement of enzymatic activity in the hybrid nanoflowers could be achieved compared with the free HRP in solution. Taking advantages of the structural feature with catalytic property, a nanoflower-based colorimetric platform was newly designed and applied for fast and sensitive visual detection of H2O2 and phenol. The limits of detection (LODs) for H2O2 and phenol were as low as 0.5 μM and 1.0 μM by the naked-eye visualization, which meet the requirements of detection of both analytes in clinical diagnosis and environmental water. The proposed method has been successfully applied to the analysis of low-level H2O2 in spiked human serum and phenol in sewage, respectively. The recoveries for all the determinations were higher than 92.6%. In addition, the hybrid nanoflowers exhibited excellent reusability and reproducibility in cycle analysis. These primary results demonstrate that the hybrid nanoflowers have a great potential for applications in biomedical and environmental chemistry.

A facile approach for the synthesis of enzyme-inorganic hybrid nanoflowers and its application as an immobilized α-chymotrypsin (ChT) reactor (IMER) for highly efficient protein digestion was described. The hybrid nanoflowers were room-temperature synthesized in aqueous solution using calcium phosphate (Ca3(PO4)2) as the inorganic component and ChT as the organic component. The effects of reaction parameters on the formation of the enzyme-embedded hybrid nanoflowers and its growth mechanism were investigated systematically. By monitoring the reaction of N-benzoyl-L-tyrosine ethyl ester (BTEE), the enzymatic activity of the immobilized ChT was calculated and the results showed 266% enhancement in enzymatic activity. The performance of such a nanoreactor was further demonstrated by digesting bovine serum albumin (BSA) and human serum albumin (HSA), with a stringent threshold for unambiguous identification of these digests, the yielding sequence coverage for nanoflower-based digestion were 48% and 34%, higher than those obtained with the free enzyme. Whereas the digestion time of BSA and HSA in the former case was less than 2 min, about 1/360 of that performed in the latter case (12 h). Furthermore, the residual activity of the nanoflowers decreased slightly even after eight repeated use, demonstrating promising stability. In addition, the hybrid nanoflower-based IMER was applicable to the digestion of complex human sample, showing great promise for proteome analysis.Ca 3 (PO 4 ) 2 -ChT hybrid nanoflowers were synthesized by a facile approach. The nanoflowers exhibited an enhanced enzymatic activity and can be used as an immobilized enzyme reactor (IMER) for highly efficient protein digestion.

A novel kind of immobilized trypsin reactor based on enzyme-inorganic hybrid nanoflowers was first developed and can be applied to ultra fast, highly efficient proteome digestion.