[Determination of serum bilirubin fractions using gel-filtration]

Serum bilirubin has been fractionated by means of a gel-filtration technique, on Bio-Gel P-10 columns and with caffeine solution as eluent. When sera containing high concentration of direct-reacting bilirubin were applied to the column, a bilirubin fraction was eluted with the protein. This bilirubin fraction is not set free from protein in the presence of urea, guanidine and sodium dodecyl sulfate; moreover it exhibits peculiar behaviour in the direct and indirect diazo-reaction and in enzymatic oxidation by bilirubin-oxidase as well. On the basis of these data, such bilirubin fraction has been identified with the previously described delta-bilirubin. A method for the quantitative measurement of delta-bilirubin, based on gel-filtration separation followed by diazo-reaction, has been worked out. Results by this method satisfactorily compared with the Kodak Ektachem method.