Use of Bilirubin Standards.

A stable stock solution of bilirubin in chloroform may be used to standardize quantitative determination of bilirubin in serum as “azobilirubin.” The molar absorptivity of azobilirubin (560 mj, 1 cm.) is 70,300 ± 280 (mean ± I S.D.) when determined from chloroform-methanol standards treated with a modified van den Bergh reagent in the Malloy and Evelyn procedure for total bilirubin. The absorptivity of azobilirubin prepared from bilirubin in serum standards is essentially the same. The molar absorptivity of bilirubin in serum (460 m, 1 cm.) is 49,100 ± 690 (mean ± 1 S.D.). Manufacturers of standard bilirubin should use the molar constants as criteria of quality. Two of three preparations differed by 6.8 and 8.5% from the molar absorptivity found in standards made by the authors. Reconstituted commercial standards are stable for 3 days when stored at 5#{176}.