Serum bilirubin fractionation using multilayer film method in hepatobiliary diseases in comparison with high performance liquid chromatography

Summary

The Ektachem multilayer film method (Ektachem) and high performance liquid chromatography (HPLC) were employed to fractionate and evaluate serum bilirubin species in 45 serum samples. The false-positive or false-high levels of bilirubin close-bonded with albumin (i.e. the delta bilirubin fraction (Bδ)) was obtained by Ektachem in sera of cases with normal bilirubin concentration and cases with unconjugated hyperbilirubinemia when compared with the results of HPLC. In the sera of cases with conjugated hyperbilirubinemia, Ektachem gave comparable levels of total bilirubin (TB), and unconjugated bilirubin (Bu) to those of HPLC, but underestimated conjugated bilirubin (Bc) and slightly overestimated Bδ.

To investigate the clinical significance of Bδ in 113 cases of various hepatobiliary diseases with conjugated hyperbilirubinemia, the ratios of Bδ to TB (Bδ/TB) and to directly-reacting bilirubin fractions (Bδ/(Bc + Bδ)) and that of Bc to Bδ (Bc/Bδ) were calculated based on the results of Ektachem and compared with each other during the course of jaundice. The mean Bδ/TB was below 40% in various hepatobiliary diseases but became as high as approximately 60% in the convalescence stage. The mean Bδ/(Bc + Bδ) was below 50% in acute hepatitis (the serum bilirubin-elevating stage) and obstructive jaundice, and it increased to above 80% in the recovery stage. In decompensated liver cirrhosis and intrahepatic cholestasis the mean Bδ/(Bc + Bδ) was about 60%, indicating continuous backflow of Bc from liver cells. The changes in Bδ/(Bc + Bδ) were much greater than in Bδ/TB. The serum alkaline phosphatase did not correlate with Bδ/(Bc + Bδ). Bc/Bδ tended to show greater change than Bδ/(Bc + Bδ). Both Bδ/(Bc + Bδ) and Bc/Bδ seemed to be good indicators for the diagnosis of the stages of jaundice in liver diseases.