摘要Sulfonylation is extensively used to label DNA and RNA,assess their interactions,and quantify compo-nents including nucleobases and nucleosides/nucleotides although the sulfonylation sites remain controversial.Here,we systematically investigated the sulfonylation of adenine(ade)and its nucleosides/nucleotides with 5-(dimethylamino)-naphthalene-1-sulfonyl chloride(DNS-Cl),5-(diethylamino)-naphthalene-1-sulfonyl chloride(DEANS-Cl),and 5-((N,N-diethylleucyl)amino)-naphthalene-1-sulfonyl chloride(DELANS-Cl).Detailed spectral analysis with nuclear magnetic resonance(NMR)spectroscopy and high-resolution mass spectrometry(HRMS)showed similar sulfonylation behaviors among the re-agents.For ade,its secondary amine in the imidazole ring(N9H)sulfonylated more readily than the exocyclic amino group(N6H2).For adenosine and its nucleotides,the 2′-OH group in the ribosyl moiety was preferably sulfonylated,whereas the 3′-OH was the preferred site for 2′-deoxyadenosine and its nucleotides.Alkylation and amidation of the aromatic amino group in these 5-amino-naphthalene-1-sulfonyl chlorides did not influence the sulfonylation preferences.This offered a reliable approach and comprehensive details of such sites for ade and its nucleosides/nucleotides.