UV RESONANCE RAMAN STUDIES OF DNA PYRENE INTERACTIONS - OPTICAL DECOUPLING RAMAN-SPECTROSCOPY SELECTIVELY EXAMINES EXTERNAL SITE BOUND PYRENE

Intercalation of pyrene into DNA results in a strong hypochromism of those pyrene electronic absorption bands which overlap the nucleic acid electronic transitions. The resonance Raman cross sections of pyrene excited within these absorption bands are decreased by factors of 10-100-fold. Raman saturation measurements indicate a dramatic increase in the intercalated pyrene nonradiative relaxation rate, which is consistent with fluorescence quenching observations. We demonstrate using the new technique of optical decoupling Raman spectroscopy (ODRS) that the remaining fluorescence derives from pyrenes not intercalated between bases, but most likely bound externally. ODRS involves a double pulse sequence, where the pump laser excites the sample in a spectral region which is selectively absorbed by one component; the pump selectively excites this component out of its electronic ground state. A probe pulse monitors the Raman spectra of the remaining ground-state species in the sample, in this case selectively enriched in intercalated pyrene. The Raman intensities and enhancement patterns of the intercalated pyrene are very different from the external site bound pyrene. This new technique of ODRS permits us to selectively monitor pyrene molecules bound at two different sites of DNA and is a new general method for studying complex systems.