Comparison of Natural Crystalline Urobilin (Stercobilin) with that Obtained in Vitro from Mesobilirubinogen.

Abstract

Fischer isolated mesobilirubinogen following an amalgam reduction of bilirubin. Soon afterwards he and Meyer-Betz obtained crystalline urobilinogen from the urine and proved its identity with mesobilirubinogen. There is little doubt that this is the chromogen of natural urobilin or stercobilin, which recent studies, have proven identical. It was therefore expected, after allowing crystalline mesobilirubinogen to stand in the air and light until the Ehrlich reaction disappeared and urobilin characteristics had become intense, that a crystalline urobilin could be isolated which would be identical with that already obtained from urine and feces. However, it appears that the natural formation of urobilin from urobilinogen is more complex than had been believed. Crystalline mesobilirubinogen (urobilinogen) was isolated from bilirubin according to H. Fischer's method. It was dissolved in glacial acetic acid and remained exposed to the ordinary light and temperature of the laboratory for 3 weeks. After this time the Ehrlich reaction was only very faint. The solution now possessed a dark brown color, exhibited intense urobilin absorption and strong green fluorescence with alcoholic zinc acetate. This solution was poured into 3 volumes of chloroform, and from now on the procedure leading to isolation of the crystalline substance was the same as previously described for isolation of urobilin from urine, and stercobilin from feces., (CHCl3 → H2O → HCl → CHCl3 → petroleum ether, ppt. → hot CHCl3, crystallization.) In this way a crystalline urobilin has been isolated twice in small amounts. After repeated recrystallization from hot chloroform the melting point of the crystals was not sharp at 155-158°C. Although this substance exhibits intense green fluorescence in alcoholic zinc acetate solution, as well as a “urobilin” spectrum it differs in a number of respects from natural urobilin or stercobilin.