ABSTRACT
After development, the layer is removed from the chamber, and the mobile-phase solvents are evaporated in a well-ventilated area in ambient air, with warm or hot air from a hair dryer, or in an oven. The evaporation process must not cause loss of volatile solutes from the layer or be carried out at a temperature that is high enough to cause compound decomposition. In some cases, drying is carried out using nitrogen flow to preclude oxidation of air-sensitive solutes. Zones are then detected by various means. Colored substances may be viewed in daylight without any treatment. Detection of colorless substances is simplest if compounds show self-absorption in the short-wave ultraviolet (UV) region (254 nm) or if they can be excited to produce fluorescence by short-wave and/or long-wave (366 nm) UV radiation. Otherwise, detection can be achieved by means of chromogenic reagents (producing colored zones) or fluorogenic reagents (producing fluorescent zones), or by biological methods. In the ideal case, the detection reagent should produce high-contrast, high-sensitivity zones that are stable (unless the method is nondestructive or reversible) and proportional to the quantity present on the layer (for quantitative evaluation). A specialized approach for direct, nondestructive detection involves the use of autoradiography, fluorography, spark chamber, or scanning techniques to measure radioactively labeled solutes (see Chapter 13). Typical detection limits for TLC and HPTLC are shown in Table 2.1. The layer serves as a storage medium for the separated zones, which can be examined as long as necessary to obtain the maximum amount of information concerning the chromatograms. The freedom from time restraints and the ability to use a variety of techniques for detection (viewing under short and long UV
FIGURE 8.1 Cabinet with removable lamp for viewing TLC plates under 254nm and 366-nm UV light. (Photography courtesy of Analtech.)
light, chromogenic and fluorogenic reagents, and biological methods) and identification (UV absorption, FTIR, Raman, and mass spectrometry, Chapter 9) in sequence are among the greatest advantages of TLC.
