other opportunities. F o r example, s a t u r a t i o n of t h e R a m a n s p e c t r a could be used to selectively remove contributions from particular spe cies. We have demonstrated the use fulness of a technique we invented called R a m a n decoupling spectros copy (11) and have used it to study separately t h e pyrene bound to t h e minor DNA groove in comparison with the pyrene that intercalates be tween bases. The intercalated pyrene has a n extremely short 7 \ lifetime and shows little saturation. In con trast, the minor groove bound pyrene has a relatively long lifetime. A R a m a n difference s p e c t r u m between high and low pulse energy flux excitation displays t h e spectral f e a t u r e s of t h e s a t u r a t i n g minor groove bound pyrene, whereas t h e high-power spectrum is enriched in intercalated pyrene. The decoupling spectroscopy can use a multipulse se quence in which a pump pulse at a particular wavelength saturates one species, whereas a probe pulse inter rogates t h e spectra of those mole cules t h a t remain i n their ground state. We expect this technique to simplify the resonance Raman spec t r a of complicated samples. Other techniques that will become increasingly important in the future include h y p e r - R a m a n scattering (53), where i n t e n s e visible wave length excitation a t v 0 results in scattering at a frequency of 2v 0 -v v , spectra with high S/N, and new vi brations that can be especially sensi tive to environment.
Future developments I n s t r u m e n t a l advances will make U V - R a m a n m e a s u r e m e n t s easier. The intercavity doubled Ar + laser makes the UV-Raman measurement comparable in difficulty to the typical visible-wavelength Raman measure ment. This continuous laser makes it possible to measure solid absorbing samples a n d to focus t h e beam to small diameters, allowing construc tion of a UV-Raman microscope. Because the majority of molecules have UV absorption bands, it is now possible to apply resonance Raman spectroscopy to many problems. The combined application of UV, visible, and near-IR Raman excitation may become common. UV excitation se lects for a small n u m b e r of reso nance-enhanced bands of an analyte and t h e measurements are made with high sensitivity and selectivity. In contrast, with nonresonance visi ble and near-IR excitation, numer ous Raman bands occur with similar intensities for all components in the
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sample in proportion to their concen trations. Thus we will obtain both average and specific information on sample composition. The number of applications of UV-Raman spectros copy is expanding rapidly a s t h e technique finds new practitioners.
Booth
I gratefully acknowledge the contributions of the graduate students, postdocs, and colleagues who accomplished the work I have the pleasure of discussing here. In addition, I am especially indebted to Richard Bormett and John Holtz for helping me with this article and to Richard Bormett and Namjun Cho for allowing me to use their data before publication. I especially thank Foil Miller for helpful conversations and Tom Spiro and Bruce Hudson for allowing me to use their figures. We also gratefully acknowl edge NIH grant 1R01GM30741 for funding this program.
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References (1) Asher, S. A. Anal. Chem. 1993, 65, 5962. (2) Asher, S. Α.; Johnson, C. R. Science 1984,225,311-13. (3) Asher, S. A. Anal. Chem. 1984, 56, 720-24. (4) Johnson, C. B.; Asher, S. A. Anal. Chem. 1984, 56, 2258-61. (5) Asher, S. Α.; Jones, C. M. In New Ap plications of Analytical Techniques to Fossil Fuels; Perry, M.; Retcofsky, H., Eds.; ACS Symposium Series 31; American Chemical Society: Washington, DC, 1986; pp. 170-80. (6) Jones, C. M.; Asher, S. A. /. Chem. Fhys. 1988, 89, 2649-61. (7) Rummelfanger, R.; Asher, S. Α.; Perry, M. B. Appl. Spectrosc. 1988, 42, 267-72. (8) Cho, N.; Asher, S. Α., submitted for publication in/. Am. Chem. Soc. (9) Killough, P. M.; DeVito, V. L.; Asher, S. A. Appl. Spectrosc. 1991, 45, 1067-69. (10) Dudik, J. M.; Johnson, C. R.; Asher, S. A./. Chem. Phys. 1985, 82, 1732-40. (11) Dudik, J. M.; Johnson, C. R.; Asher, S. A./. Phys. Chem. 1985, 89, 3805-14. (12) Williams, K.P.J.; Klenerman, D. / Raman Spectrosc. 1992, 23, 191. (13) Getty, J. D.; Westre, S. G.; Bezebeh, D. Z.; Barrall, G. Α.; Burmeister, M. J.; Kelly, P. B.Appl. Spectrosc. 1992, 46, 620. (14) a. Asher, S. Α.; Ludwig, M.; Johnson, C. B..J. Am. Chem. Soc. 1986, 108, 318697. b. Johnson, C. R.; Ludwig, M.; Asher, S. A. /. Am. Chem. Soc. 1986, 108, 905-12. (15) Ludwig, M.; Asher, S.A./ Am. Chem. Soc. 1988, 110, 1005-11. (16) Sweeney, J.; Asher, S.A. /. Phys. Chem. 1990, 94, 4784-91. (17) Johnson, C. R.; Ludwig, M.; O'Donnell, S.; Asher, S. A. /. Am. Chem. Soc. 1984, 106, 5008-10. (18) Su, C; Wang, Y.; Spiro, T. G. /. Ra man Spectrosc. 1990, 21, 8179. (19) Fodor, S.; Copeland, R.; Grygon, C; Spiro, T. / Am. Chem. Soc. 1989, 111, 5509 (20) Rava, R.; Spiro, T. G. /. Phys. Chem. 1985, 89, 1856. (21) Harada, I.; Takeuchi, H. In Spectros copy of Biological Systems; Clark, R. T.; Hester, R. E., Eds.; John Wiley and Sons: New York, 1986. (22) Hudson, B. S.; Mayne, L. C. In Bio logical Applications of Raman Spectroscopy, Spiro, T. G., Ed.; John Wiley and Sons: New York, 1987; Vol. II.
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ANALYTICAL CHEMISTRY, VOL. 65, NO. 4, FEBRUARY 15, 1993 · 209 A
