Nanoporous Substrates with Molecular-Level Perfluoroalkyl/Alkylamide Surface for Laser Desorption/Ionization Mass Spectrometry of Small Proteins

Abstract

The rapid detection of biomolecules greatly contributes to health management, clinical diagnosis, and prevention of diseases. Mass spectrometry (MS) is effective for detecting and analyzing various molecules at high throughput. However, there are problems with the MS analysis of biological samples, including complicated separation operations and essential pretreatments. In this study, a nanostructured organosilica substrate for laser desorption/ionization mass spectrometry (LDI-MS) is designed and synthesized to detect peptides and small proteins efficiently and rapidly. The surface functionality of the substrate is tuned by perfluoroalkyl/alkylamide groups mixed at a molecular level. This contributes to both lowering the surface free energy and introducing weak anchoring sites for peptides and proteins. Analyte molecules applied onto the substrate are homogeneously distributed and readily desorbed by the laser irradiation. The organosilica substrate enables the efficient LDI of various compounds, including peptides, small proteins, phospholipids, and drugs. An amyloid β protein fragment, which is known as a biomarker for Alzheimerʼs disease, is detectable at 0.05 fmol μL–1. The detection of the amyloid β at 0.2 fmol μL–1 is also confirmed in the presence of blood components. Nanostructured organosilica substrates incorporating a molecular-level surface design have the potential to enable easy detection of a wide range of biomolecules.

Construction of a ternary component chip with enhanced desorption efficiency for laser desorption/ionization mass spectrometry based metabolic fingerprinting

“…In contrast to those conventional substrates, silica serves as an ideal candidate for LDI chip substrate due to the low thermal conductivity (7.6 W m/ (m•k)), high chemical stability, low background interference, and adjustable pore structure (Zhao et al, 2022). Very recently, Norihiro et al reported nanoporous organosilica membranes with a pore size of ~150 nm as a novel substrate for small protein detection in LDI-MS (Mizoshita et al, 2022). While to achieve a direct and sensitive detection of small metabolites (<5 nm) in bio-fluids, the mesoporous silica substrate needs to be carefully designed to couple with the LDI-MS chip for high sensitivity and selectivity.…”

Perfluoroalkyl Group-Covered Organosilica Films for the Sensitive Detection of Sulfonylurea Herbicides in Laser Desorption/Ionization Mass Spectrometry

“…25 We further extended the tailor-made surface design to detect a trace amount of an amyloid β-protein fragment, which is an important biomarker for Alzheimer's disease. 26 In a prior study, an alkylamide group was introduced into the naphthalimide moiety and a perfluoroalkyl group reacted with silanol, leading to a mixed surface, where opposite functional groups were distributed at the molecular level. The mixed surface achieved the detection of 0.2 fmol μL −1 of amyloid β in the presence of blood-related components.…”

“…Matrix-free LDI-MS has been developed using UV-absorbing solid substrates, such as nanostructured silicon, metal oxide nanoparticles, and carbon materials, where the absorbed laser energy is transferred to analyte molecules for desorption and ionization. Recently, we proposed the use of organosilica films as promising solid substrates. − The matrix-free LDI technique, which is especially useful for analyzing low-molecular-weight compounds, including drugs, peptides, lipids, and agrochemicals, − would be helpful for rapid screening of trace SU herbicide residues in environmental and food analysis.…”

Nanostructured Layer of Silver for Detection of Small Biomolecules in Surface-Assisted Laser Desorption Ionization Mass Spectrometry

“…In addition, the process of radiotherapy in patients was monitored, and a preliminary panel of metabolites in serum was identified [20]. Mizoshita et al designed nanoporous substrates with molecular-level perfluoroalkyl/alkylamide surface for LDI-MS of small proteins [21]. Surface functionality was tuned by mixing perfluoroalkyl/alkylamide groups at a molecular level, and the amyloid fragment, which was suggested as a biomarker of Alzheimer's disease, was detected at a concentration of 0.2 fmol/µL in the presence of other blood compounds [21].…”

“…Mizoshita et al designed nanoporous substrates with molecular-level perfluoroalkyl/alkylamide surface for LDI-MS of small proteins [21]. Surface functionality was tuned by mixing perfluoroalkyl/alkylamide groups at a molecular level, and the amyloid fragment, which was suggested as a biomarker of Alzheimer's disease, was detected at a concentration of 0.2 fmol/µL in the presence of other blood compounds [21]. Palladium nanoparticle-decorated thiol-functionalized metal-organic framework (MOF) nanocomposite was fabricated for the sensitive analysis of oligosaccharides [22].…”