UV resonance Raman spectra of guanosine and its seven isotope-substituted analogs (2-13 C, 2-15 N, 6-18 O, 7-15 N, 8-13 C, 9-15 N and 1 -13 C) were measured with 257 nm excitation in H 2 O and D 2 O solutions. In-plane vibrations of the guanine ring were selectively enhanced in the UV resonance Raman spectra, and most Raman bands showed significant wavenumber shifts upon isotopic substitution. The observed isotope shifts were used to assign the Raman bands to vibrations of the peripheral sites (N1-H, C2-NH 2 and C6 O), the pyrimidine ring and/or the imidazole ring. Previous assignments for some Raman bands were shown to be inconsistent with the isotopic data and they were revised. Relationships between the vibrational modes and the sensitivities to hydrogen bonding or conformation are discussed for known Raman marker bands. Each hydrogen bond marker arises from a vibration that involves, at least partly, the proton donor or acceptor atom. All the marker bands of glycosidic bond orientation and ribose ring puckering actually involve atomic displacements around the N9-C1 moiety connecting the guanine ring to ribose, permitting vibrational coupling between them. The isotopic wavenumber shifts reported here may be useful in improving the force field for the 9-substituted guanine ring and in interpreting the vibrational spectra of guanine nucleoside and nucleotides.