Quantitative thin-layer chromatographic analysis of ergotamine tartrate and caffeine in the nanogram range

Direct, quantitative, thin-layer chromatographic methods for the determination of ergotamine tartrate and caffeine in the presence of each other in blood and in pharmaceutical preparations are described. The blood is centrifuged, the plasma decanted from the coagulum and deproteinized with acetone-methanol. After removal of the solvent mixture, the active ingredients are extracted from the remaining aqueous solution with chloroform. In the case of the pharmaceutical preparations, the active ingredients are also extracted with chloroform or methanol. Ergotamine tartrate and caffeine are separated on pre-coated silica gel 60 F254 plates and measured directly on the thin-layer plate, ergotamine tartrate being determined by the fluorescence method, excitation wavelength 365 nm, at lambdamax.=450 nm and caffeine by the reflactance method at lambdamax.=274 nm. The analytical methods are suitable for the bioavailability studies of these drugs in blood as they enable the determination of 20 ng ergotamine tartrate with a coefficient of variation of 6.7% and the determination of 200 ng caffeine with a coefficient of variation of 6.1%. The methods have also proved useful in the determination of the active ingredients of drug forms such as tablets and suppositories and can be well reproduced with a maximum coefficient of variation of 4.9% for ergotamine tartrate and 3.5% for caffeine.