developed by Yang et al. [14] developed and involved the synthesis of their fluorescent derivatives, which were formed by reacting them with an electrophilic fluorogenic reagent, 4-(N-chloroformethyl-N- methylamino-7-N- N–dimethylaminosulfonyl-2, 1, 2–benzoxydiazole (DBD- COCl). These derivatives show intense fluorescence at long excitation wavelengths (450Ex and 560Em nm) and thus significantly reduce the level of detection. It was found that Propranolol, Metoprolol and Atenolol, three very commonly used b blockers, reacted directly with reagent forming 1:1 adducts by reaction with the secondary amino group of the b-blockers. The DBD-COCl reagent reacted readily with the drugs under mild conditions, with no catalyst, and the reaction was relatively fast and was complete in about 5 min. The initial separation was carried out on a TSKgel ODS- 80T column, 15 cm long, 4.6 mm I.D., packed with 5 mm particles. The solvent gradient used to

of 0.5 ml/min., the separation ratio was 1.33. The mobile phase used for the separation of DBD–Ate was methanol, also at a flow rate of 0.5 ml/min., the separation ratio being 1.53. The excitation wavelength was 450 nm and the emission wavelength was 560 nm. The fluorescent derivatives were found to be stable at 4˚C for over 1 week. Courtesy of the Royal society of Chemistry, Ref. [12]   Figure 49 The Separation of Derivatized Metoprolol and Atenolol at High Sensitivity

nbsp; Courtesy of the Royal Society of Chemistry, Ref. [14]   Figure 48 The Separation of Derivatized Propranolol (Pro) Metroprolol (Met) and Atenolol (Ate

nbsp; The detection limits at a signal-to-noise ratio of 3 were 50 fmol for both (S)- and (R)-Propanolol, 12 and 17 fmol for (S)- and (R)-Metroprolol respectively and 15 and 20 fm for (S)- and (R)-Atenolol respectively.   The cyclodextrin based stationary phases are some of the more popular and effective chiral stationary phases presently available. One of their distinct advantages lies in their unrestricted and successful use with all types of solvent. In particular, they can be used very effectively in the reversed phase mode (a method of development that is not possible with some other chiral stationary phases) as well as being very effective in a normal phase