Photocatalytically Patterned TiO₂Arrays for On-Plate Selective Enrichment of Phosphopeptides and Direct MALDI MS Analysis

Abstract

We report the development of photocatalytically patterned TiO(2) arrays for selective on-plate enrichment and direct matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) analysis of phosphopeptides. A thin TiO(2) nanofilm with controlled porosity is prepared on gold-covered glass slides by a layer-by-layer (LbL) deposition/calcination process. The highly porous and rough nanostructure offers high surface area for selective binding of phosphorylated species. The patterned arrays are generated using an octadecyltrichlorosilane (OTS) coating in combination of UV irradiation with a photomask, followed by NaOH etching. The resulting hydrophilic TiO(2) spots are thus surrounded by a hydrophobic OTS layer, which can facilitate the enrichment of low-abundance components by confining a large volume sample into a small area. The TiO(2) arrays exhibit high specificity toward phosphopeptides in complex samples including phosphoprotein digests and human serum, and the detection can be made in the fmole range. Additional advantages of the arrays include excellent stability, reusability/reproducibility, and low cost. This method has been successfully applied to the analysis of phosphopeptides in nonfat milk. The patterned TiO(2) arrays provide an attractive interface for performing on-plate reactions, including selective capture of target species for MALDI-MS analysis, and can serve as a versatile lab-on-a-chip platform for high throughput analysis in phosphoproteome research.

MALDI Profiling and Applications in Medicine

“…However different groups have reported improved signals utilising a further matrix (dihydroxybenzoic acid) in combination with phosphoric acid as a matrix additive or combinations of two matrices (CHCA and 3-hydroxypinolinic acid (3-HPA)) [ 45 , 100 ]. Further approaches to enhance phosphoprotein and peptide analysis by MALDI profi ling have focussed on chemically altering the MALDI plate in order to selectively trap phosphate containing species and these have mainly utilised titanium dioxide as the linker to trap the phosphate group [ 89 ]. Phosphopeptide capture has been achieved on a sintered material before elution and subsequent MALDI profi ling analysis and also by on-plate capture using photocatalytically patterned titanium dioxide distributions on plate using nanoparticles [ 20 , 77 ].…”

Mesoporous TiO2 aerogel for selective enrichment of phosphopeptides in rat liver mitochondria

“…5B. The sensitivity of the on-target enrichment performed by mesoporous TiO 2 nanoparticles is at the low femtomole [21,27,28,31]. The higher sensitivity achieved by TiO 2 aerogel might be contributed to the excellent compatibility of enrichment procedure with MALDI-TOF MS and the large binding capacity of TiO 2 aerogel.…”

“…The limited specific surface area of the reported MOAC materials results in low binding capacity and low sensitivity for phosphopeptide detection. The sensitivity of the on-plate purification approach based on MOAC is at femtomole level [21,27,28,31], which may limit the application of MOAC in phosphoproteome study. Therefore, the demand is high for new materials with high surface area, in order to improve the binding capacity and detection sensitivity for phosphopeptide analysis.…”

Zinc Ion-immobilized Magnetic Microspheres for Enrichment and Identification of Multi-phosphorylated Peptides by Mass Spectrometry

“…21 MOAC also has been intensively applied for the selective enrichment of phosphopeptides based on Lewis acid-base interaction, using titanium, zirconium, aluminum, niobium, and other metal oxides. [22][23][24][25][26] Recently, TiO2 has gained increasing attention and is regarded as one of the most powerful materials for phosphopeptide enrichment, particularly for isolating mono-phosphorylated peptides. 27,28 Consequently, design and synthesis of new materials with high enrichment selectivity for multi-phosphorylated peptides are still demanded.…”