Bilirubin and reperfusion injury: all bilirubins are not equal.

In a recent letter, Manzinate et al. reported that pretransplant “bilirubin” concentrations do not correlate with early reperfusion injury after liver transplantation (1). The data were interpreted as showing that bilirubin does not protect against ischemic reperfusion injury in humans as it has been reported to do in animal experiments (2). However, the data presented may have little or no bearing on the protective or antioxidant effects of bilirubin and are not necessarily inconsistent with the prior reports. That is because the “bilirubin” measurements reported do not necessarily reflect the actual concentrations of unconjugated bilirubin. By referring to bilirubin in the singular, the letter glossed over the crucial fact that the blood of the pretransplant patients would have contained a large number of different bilirubinoid species with different chemical and biological properties. These would include unconjugated bilirubin, three bilirubin glucuronides, many isomers of the glucuronides formed by acyl migration, and various species of bilirubin covalently bound to albumin (3–5). Though all of these might have antioxidant properties, only one of them, unconjugated bilirubin, is a highly lipophilic antioxidant (6, 7) and it is this particular biosynthetic chemical that is postulated to have protective effects and has been implicated in previous studies (2, 7, 8). The specific concentrations of unconjugated bilirubin in the patients were not reported and may well have been low in many of them. Even if a concentration could be calculated from, say, direct and total bilirubin measurements, it would not be reliable unless the blood samples had been collected and manipulated under conditions that preclude the facile esterase hydrolysis of bilirubin conjugates to unconjugated bilirubin. Because of the likely complexity of the bilirubinoid species in the blood of the pretransplant patients, a highly discriminating analytical technique, such as highpressure liquid chromatography, and careful sample handling would have been required to accurately measure the true content of unconjugated bilirubin IX . Whether bilirubin (i.e., 4Z,15Z-bilirubin IX ; Chemical Abstracts Number 635-65-4) really does have significant protective effects in vivo or in vitro for preventing hepatic inflammation and injury remains an interesting and open question, but the report by Manzinate et al. (1) is largely irrelevant to the issue because it depended on measurements that did not distinguish the different bilirubinoid species that may have been present in their samples. There is a longstanding convention that the word “bilirubin”, used without qualifiers, refers specifically to the 4Z,15Z isomer of bilirubin IX (9), the particular lipophilic pigment produced by catabolism of heme. It might avoid potential ambiguity if investigators were to follow that convention. Using the term “bilirubin,” in the singular, to refer collectively to several different kinds of bilirubins can be confusing and misleading. Antony F. McDonagh Division of Gastroenterology and The Liver Center University of California San Francisco San Francisco, CA Address correspondence to: Antony F. McDonagh, Ph.D., Division of Gastroenterology, Room S-357, Box 0538, University of California San Francisco, 513 Parnassus Ave, San Francisco, CA 94143-0538. E-mail: tony.mcdonagh@ucsf.edu Received 27 February 2007. Revision requested 22 March 2007. Accepted 2 April 2007. Copyright © 2007 by Lippincott Williams & Wilkins ISSN 0041-1337/07/8312-1658 DOI: 10.1097/01.tp. 0000266996.25289. 6c