Diphenylcarbazone reagent

Barbiturate metabolites are more heavily colored by the mercury(I) nitrate reagent (exception allobarbital), while unaltered barbiturates react more sensitively to the mercury(II)-diphenylcarbazone reagent (q.v.) [1]. 

Cover columns 3 to 8 with a glass plate and spray columns 1 and 2 with mercuric chloride-diphenylcarbazone reagent White spots on a violet background indicate the presence of barbiturates or related compounds (e.g. glutethimide). Cover columns 1 and 2 and 5 to 8 with glass plates. 

Mercuric chloride-diphenylcarbazone reagent. Blue spots are given by sulphonamides. 

Thin-layer Chromatography. System TD—Rf 41 system TE—Rf 31 system TF—Rf 61 system TH—Rf 51. Mercuric chloride-diphenylcarbazone reagent, positive mercurous nitrate spray, black Zwikkefs reagent, pink.)... 

Thin-layer Chromatography. System TD—Rf 05 system TE—Rf 02 system TF—Rf 02. Ferric chloride solution, yellow mercuric chloride-diphenylcarbazone reagent, violet, turns pink on heating acidified potassium permanganate solution, yellow Van Urk reagent, yellow, fades on heating.)... 

Thin-layer Chromatography. Sy.stem TA—Rf 52 system TB—Rf 07 system TC—Rf 62 system TD—Rf 43 system TE—Rf 61 system TF—Rf 47. (Dragendorff spray, positive FPN reagent, pale yellow Marquis reagent, very weak pale yellow mercuric chloride-diphenylcarbazone reagent, blue, on heating turns pale pink acidified potassium permanganate solution, positive.)... 

Thin-layer Chromatography. System TA—Rf 58 system TD—Rf 00 system TE—Rf 00 system TF—Rf 00 system TL—Rf 00 system TM—Rf 00. (Location under ultraviolet light, violet fluorescence mercuric chloride-diphenylcarbazone reagent, blue acidified potassium permanganate solution, positive Van Urk reagent, blue.)... 

In this case, the TLC system most commonly employed uses silica gel plates and a mobile phase of ethyl acetate/methanol/25% ammonia (85 10 5, by volume). The plates are prepared and the chromatogram developed in the standard way. After development, the plate is removed from the mobile phase, the solvent front marked, and the plate dried. Visualization of barbiturates is best achieved by the use of a mercuric chloride-diphenylcarbazone reagent. The latter is prepared as two component solutions, i.e. (i) 0.1 g of diphenylcarbazone in 50 ml of methanol, and (ii) 0.1 g of mercuric chloride in 50 ml of ethanol. These solutions should be freshly prepared and mixed just before use. The presence of barbiturates will give rise to blue-violet spots on a pink background when using this reagent system. 

---