Improvement and extension of the application scope for matrix-assisted laser desorption/ionization mass spectrometric analysis-oriented N-alkylpyridinium isotope quaternization

Based on our previous report on N-alkylpyridinium isotope quaternization (NAPIQ) for the analysis of cholesterol and fatty alcohols, we have further developed the NAPIQ method for wider applications in screening various small compounds. The reaction scope and improvement were investigated via the screening of compounds with different reaction sites. The experimental results showed that the NAPIQ strategy was suitable not only for steroids with alcoholic or α,β-unsaturated ketone moieties, but could also be applied to label carbohydrates by changing the reaction solvent. The byproducts from target compounds with multiple groups could be reduced by improving the reaction conditions. The derivatization method significantly improved the detection sensitivity for these compounds in matrix-assisted laser desorption/ionization Fourier transform ion cyclotron resonance mass spectrometric (MALDI-FTMS) analysis. The detection sensitivity and selectivity of the NAPIQ method were higher than conventionally used Girard reagents. Finally, endogenous steroids in urine samples were screened using the NAPIQ method. NAPIQ was proven to be an efficient alternative method for analyzing steroids and carbohydrates, without the need for isotopically coded internal standards.