Indirect fluorometric detection techniques on thin layer chromatography and effect of ultrasound on gel electrophoresis
Thin-layer chromatography (TLC) is a broadly applicable separation technique. It offers many advantages over high performance liquid chromatography (HPLC), such as easily adapted for two-dimensional separation, for "whole-column" detection and for handling multiple samples, etc. However, due to its draggy development of detection techniques comparing with HPLC, TLC has not received the attention it deserves. Therefore, exploring new detection techniques is very important to the development of TLC. It is the principal purpose of this dissertation to present a new detection method for TLC--indirect fluorometric detection method. This detection technique is universal, sensitive, nondestructive, and simple. This will be described in detail from Sections I through Section V;Section I and III describe the indirect fluorometric detection of anions and nonelectrolytes in TLC. They are based on the displacement of fluorophore by the sample molecule, which cause a decrease of fluorescence. The negative signals can be monitored, i.e., analytes can be detected indirectly;In Section II, a detection method for cations based on fluorescence quenching of ethidium bromide is presented. Cations such as Cu[superscript]2+, Cr[superscript]3+ and Hg[superscript]+ can be separated on silica gel TLC plate and detected by quenching the red fluorescence of ethidium bromide;In Section IV, a simple and interesting TLC experiment is designed, three different fluorescence detection principles are used for the determination of caffeine, saccharin and sodium benzoate in beverages;A laser-based indirect fluorometric detection technique in TLC is developed in Section V. The new technique can be easily used for quantitative measurements because of its two-dimension scanning capability. With this technique a detection limit of 6 pg can be achieved;Section VI is totally different from Sections I through V. An ultrasonic effect on the separation of DNA fragments in agarose gel electrophoresis is investigated. It is found that the ultrasound improves the separation of DNA fragments without degrading them, a phenomenon that is totally different from that of ultrasonic scission.