The need to measure ultralow levels of pharmaceuticals in biological matrices at femtogram and attogram levels presents a significant challenge to bioanalysts. Liquid chromatography has proven to be a versatile and valuable tool for separating analytes from complex biological matrices and fluorescence detection provides both the sensitivity and the selectivity required to measure femtogram and attogram levels of analytes. Solid-state diode lasers have been used primarily in printers, compact disc recorders, and bar code scanners but have recently been adapted for use as light sources for fluorescence detection. Excellent spectral features in the visible and near-infrared regions of the spectrum, together with low cost and ruggedness, make diode lasers an attractive alternative for use as light sources in analytical measurement. Biological matrices demonstrate minimal background signals in the far-red regions of the spectrum which diode lasers emit and diode lasers are among the most stable light sources available. These facts along with expected developments in labeling systems make the potential for use of diode lasers in LC-detection quite promising. This paper reviews characteristics of diode lasers, the properties of potential visible and near-infrared fluorescent probes, instrumental aspects of diode laser fluorometers, and future trends that can be expected in this exciting field of bioanalytical research.

The concentration detection limit was superior to literature reports for detection of fatty acids. The mass detection limit provided approximately an order of magnitude improvement over conventional fluorescence. The reagent is potentially useful for analysis of carboxyl containing analytes at low concentrations.

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This report describes the first application of 5-(bromomethyl)fluorescein (5-BMF) for the quantitation of a pharmaceutically relevant carboxyl-containing analyte in a biological matrix. An analytical method for quantitation of γ-(cholesteryloxy)butyric acid (CBA), a relatively new antitumor agent, in different tissues of Sprague-Dawley rats was developed. 5-BMF was employed to form a stable and spectrally well-characterized conjugate of CBA. The derivatization yield was maximized by optimizing several reaction variables. The conjugate was separated by high-performance liquid chromatography and quantitated by a laboratory-constructed argon ion laser fluorometer. The detection limits for CBA were 4.6 × 10 -9 and 6.34 × 10 -11 M by conventional and laser-induced fluorescence (LIF), respectively. A derivatization limit of detection of 1.85 × 10 -9 M was achieved by LIF for the conjugate. The analytical method was useful for quantitation of CBA in various tissues in the picogram per milliliter range.