Studies on Bilirubinoid-Azopigments Part Ⅰ: Paper Chromatography on Bilirubinoid-Azopigments

Azopigments, yielded after the addition of diazotised sulphanilic acid to the each of bilirubin, mesobilirubin and bilirubindimetbylester, have been studied by paper partition chromatography if there were some differences in Rf on the chromatograms among these azopigments. The following results were obtained; Regarding developing solvents, methylethylketone-n-propanol-water (15:5:6), n-propanol-water (7:3), n-propanol-acetic acid-water (4:1:5) and saturated n-butanol with 10% citric acid were found as giving the most adequate partition of these samples from the chromatographies done with uses of more than forty kinds of solvents. Using these developing solvent systems, the distinct diferences in the partition were found among these three bilirubinoid-azopigments. In developing process on the chromatography of these azopigments, a reddish-violet spot changed its color partially to blue, and was finally divided into two spots of red and blue. This indicated the coexistence of two kinds of the azopigments in the crude samples. By leaving or heating the paper after developing the chromatography, a yellow spot appeared with the Rf common to these three azopigments, and it was possibly throught that this spot resulted from excess amount of the diazonium salt remained in the reacting solution.