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Abstract—A spectrophotometer using the photon‐counting method is described. The sample cuvette is mounted on a table which can be displaced in the X and Y directions, permitting investigation of absorption and self‐absorption processes within the cuvette. Fluorescence detection is performed at right angles to the direction of the exciting beam, the analysis grating being driven by a stepping motor. The apparent spectrum is fed into a multichannel analyser and printed. Corrections are made to the data provided by the printer. Signal‐to‐noise ratio, fluorescence spectra and fluorescence excitation spectra are discussed in more detail for the case of more concentrated solutions. This very high sensitivity device allows room‐temperature investigation of the fluorescent emission from neutral aqueous solutions of nucleic acid bases, nucleosides, nucleotides, dinucleotides etc., quantum yields for these molecules being of the order of 10‐4. A table of minimum concentration values of some commonly used fluorescent probes is given.

SynopsisRaman spectra of AMP, UTP, GMP, and CMP, and of their bromo-derivatives , are reported. They are obtained using excitation wavelengths of 457.9 nm (ionized continuous argon laser) and of 300 nm (tunable pulsed dye laser). Comparison of spectra leads to the following observations: (1) preresonance Raman effects on nucleotides spectra a t 300 nm; (2) resonance Raman effects on bromoderivatives spectra a t 300 nm; (3) in dilute solution (10-4M), shifts and enhancements of Raman lines of the bromo-derivatives with respect to the corresponding lines of nucleotides. On the basis of these comparisons, the assignments of the Raman lines are discussed. This provides the necessary background for the understanding of the properties of selected groups in DNA in dilute solution. The new experimental set-up for measurements of Raman spectra using excitation in the uv region is described. It is specially designed to incorporate the pulsed feature of the excitation laser and for correcting of the instabilities of the source.

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