Covalent organic framework film-based laser desorption/ionization mass spectrometry for rapid and sensitive quantification of homocysteine in human serum

Abstract

Rationale: Homocysteine (hcy) is a metabolite in the human body and an important determinant of cardiovascular health. To assist in the assessment of human cardiovascular safety, a rapid and accurate quantitative analysis method must be developed. Laser desorption/ionization mass spectrometry (LDI-MS) has been widely used in biological, chemical, and medical analyses due to its excellent characteristics of high throughput, low sample consumption, and high speed. Here, an LDI-MS method based on covalent organic framework (COF) film was developed for the rapid and sensitive determination of hcy in human serum using an isotope-labeled internal standard.

Methods: We tried to cultivate COFs on indium-tin oxide substrate at room temperature to form a thin film, which was used in LDI-MS. Compared with the traditional organic matrices, the COF film had a clean background and a high signal response for the detection of hcy. In addition, using COF film as the substrate, with a high signal intensity and a clean background, can be helpful to analyze a series of small molecules such as amino acids, bisphenols (Bps), estrogens, and drugs. We evaluated the limit of detection (LOD) and the reproducibility of this method. Finally, the calibration curve was constructed for the quantification of serum hcy using an isotope-labeled internal standard and was applied for the rapid determination of hcy in clinical human serum.

Results: COF film-assisted LDI-MS had a higher response signal and a cleaner background compared to four conventional organic matrices. The LOD of the method for hcy was 0.5 μmol/L, equivalent to 500 fmol. This method exhibited excellent performance for small-molecule compounds, including amino acids, Bps, drugs, and estrogens. The reproducibilities of this method for shot-to-shot and dot-to-dot were determined to be good. This method was applied for the rapid analysis of hcy in clinical human serum, showing good correlation with those obtained by hospital testing.

Conclusion: The COF film-based LDI-MS method had simple sample preparation, short analysis time, clean background, and good reproducibility for hcy analysis. As an important indicator of human health, the detection of serum hcy is of great significance to human health.