Attenuated total internal reflectance infrared microspectroscopy as a detection technique for high-performance liquid...

The use of single reflection attenuated total internal reflection infrared microspectroscopy as a detector for high-performance liquid chromatography (HPLC) is demonstrated. The terminus of the HPLC column is placed at the focus of an ATR infrared microscope, allowing several advantages over other detection techniques. These include the following: (1) the reduction or elimination, or both, of detection cell dead volume, (2) the ability to interrogate a nearly pure aliquot of analyte, and (3) the ability to signal average spectra of the same aliquot (depending on its size) thereby improving the signal-to-noise ratio of the measurement and, concomitantly, the analytical characteristics of the method. Static measurements of succinylcholine chloride in water have shown a detection limit of 0.7 parts per thousand (ppt). Two-microliter injections of succinylcholine chloride in a 5-cm x 1-mm C-18 column with a flow rate of 60 microL/min have shown a detection limit of 1.9 ppt. This analytical concentration corresponds to a mass of 3.8 microg in the injection loop and a mass of 350 fg in the sampling volume. The potential of this detector for HPLC is demonstrated, and future improvements are also discussed.