Delta Bilirubin: The Fourth Fraction of Bile Pigments in Human Serum

Abstract

Delta bilirubin is a strong, possibly covalent complex between bilirubin and an albumin-like protein, found in the serum of most jaundiced adults. The bili-albumin complex reacts directly diazo-positive. An examination of its azo pigments indicates that only one half of the tetrapyrrole is linked to the protein, presumably via an amide bond formed between a propionic acid side chain of bilirubin and an amine group (e.g., E-NH, of a lysine) in the albumin backbone; the other (distal) propionic acid chain is largely unesterified. Preliminary analyses of the peptides generated by chemical or enzymic cleavage of the delta fraction suggest a bilirubin bonding region within positions 128-297 of albumin and another involving positions 298-585. Thus, delta bilirubin may represent the first true human biliprotein. The implications of this biliprotein in both the fundamental and applied aspects of bile pigment research are discussed.

Enzymatic assay for conjugated bilirubin (Bc) in serum using bilirubin oxidase (BOD)

“…Recently, the fractional assay of bilirubins has been widely used as a clinical assay test using the HPLC method or the Ektachem (now called Vitros) system, because the significance of clinical findings with β-, γ-and δ-fractions has been reported by several groups (1,(5)(6)(7)(8).…”

“…Bδ, expressed as a percentage for T-Bil, always fails to rise above 30 to 40% during unresolved hepatic failure (6). In such cases, Bc often remains above 50% of T-Bil until or near the death of the patient (1).…”

“…However, Bu and Bδ react in small amounts with diazo reagent in direct assay, and some observations have been also reported with other methods such as the BOD and vanadate methods (1)(2)(3)(4). In terms of clinical significance, values of Bc and Bδ are better indicators of liver function than are the values of D-Bil, T-Bil, and hepatic enzymes such as AST and ALP (1,(5)(6)(7)(8).…”

Canalicular and Sinusoidal Disposition of Bilirubin Mono- And Diglucuronides in Sandwich-Cultured Human and Rat Primary Hepatocytes

“…The calibration curve was prepared using B as standard. Since there is no significant difference between the absorbance spectra of B and BMG or BDG (Wu, 1983;Brower et al, 2001), this calibration was used for the determination of BMG and BDG concentration.…”

Differences Between Enzymatic and Diazo Methods for Measuring Direct Bilirubin

“…Figure la shows four bilirubin subtractions detected on HPLC: α (unconjugated bilirubin), β (monoconjugated bilirubin), γ (biconjugated bilirubin) and δ (δ bilirubin (10)(11)(12), which is a covalently albumin-bound bilirubin). Their retention times (mean ± SD) were 34.7 ± 0.1 min, 29.3 ± 0.1 min & 28.6 ± 0.1 min (biphasic), 24.9 ± 0.1 min, and 20.2 ± 0.3 min, respectively.…”