Controversies in bilirubin biochemistry and their clinical relevance.

Despite a century of research, several clinically relevant areas of bilirubin biochemistry remain controversial, poorly understood, or unrecognized. These include: (i) The structure and molecularity of bilirubin under physiological environments such as membranes, brain tissue and when bound to proteins. Related to this is the large number of structurally different bilirubin species that may occur in blood under pathological conditions and their potential effects on measurements of bilirubin and free bilirubin. (ii) The mechanism of phototherapy, the neurotoxicity of the photoisomers produced and their influence on measurements of bilirubin and free bilirubin. (iii) The role of membrane transporters in the passage of unconjugated bilirubin across the placenta, intestine, vascular epithelium, blood-brain barrier, and into the liver. (iv) Biochemical mechanisms of bilirubin toxicity, pharmacologic prevention of kernicterus, the contribution of bilirubin to antioxidant defenses, and the practical value of free bilirubin measurements for identifying infants at most risk of kernicterus.

Role of bilirubin as antioxidant in neonatal jaundice and effect of ethanolic extract of sweet lime peel on experimentally induced jaundice in rat.

Bilirubin above a threshold level is toxic to human system and is excreted in urinary and through gastrointestinal tract. The role of bilirubin as antioxidant is debatable. This paper aims at elucidating the role of bilirubin as an antioxidant in neonatal jaundice patients. It is observed that bilirubin up to 6 mg/dl in blood acts as an antioxidant and above 12.5 mg/dl is strongly prooxidant. Phototherapy is the accepted therapeutic management of neonatal jaundice and has been shown to enhance the oxidative stress. Approaches have been taken to formulate a herbal medication which will reduce bilirubin level in the neonates without inducing additional damages. The ethanolic extract of sweet lime peel, administered orally at a dose of 72 microg is found to reduce the oxidative stress in erythrocytes of phenylhydrazine-induced jaundiced rats treated with phototherapy.

Relationships between serum (ZZ)-bilirubin, its subfractions and biliverdin concentrations in infants at 1-month check-ups.

In 150 infants, including those with breast milk jaundice, who were brought to our hospital for their 1-month check-ups, the serum concentrations of (ZZ)-bilirubin, its subfractions and biliverdin were measured by high-performance liquid chromatography and the relationships among them investigated. (ZZ)-Bilirubin was found to have the highest serum concentration, followed by (ZE)-bilirubin, accounting for 14.0 (geometric mean) % of (ZZ)-bilirubin. Biliverdin had a serum concentration of 0.95% of (ZZ)-bilirubin. There was only a small amount of total (di- and mono-) glucuronosyl bilirubin, 0.42% of (ZZ)-bilirubin. (ZE)-Bilirubin, (EZ)-bilirubin, (EZ)-cyclobilirubin. biliverdin, diglucuronosyl bilirubin and monoglucuronosyl bilirubin (C-8 and C-12) showed positive logarithmic correlations with (ZZ)-bilirubin (R2=0.16 or above, P<0.05). (ZE)-Bilirubin showed a significant positive logarithmic correlation with (ZZ)-bilirubin (R2=0.863, P<0.0001). Furthermore, (EZ)-cyclobilirubin, the most important photoisomer in phototherapy for neonatal hyperbilirubinaemia, was detected in very small amounts in approximately half of the neonates (84 of 150) when they were in conditions of only weak ambient light. The relationship between total glucuronosyl bilirubin and (ZZ)-bilirubin concentrations fitted a model of saturation kinetics of bilirubin UDP-glucuronosyltransferase.

Formation of propentdyopents and biliverdin, oxidized metabolites of bilirubin, in infants receiving oxygen therapy.

BACKGROUND: Bilirubin has antioxidative effects. When bilirubin reacts with reactive oxygen species, oxidized metabolites of bilirubin are formed, such as biliverdin and propentdyopents. A decrease in serum bilirubin concentration and an increase in serum and urinary oxidized metabolites of bilirubin may indicate the protective action of bilirubin against reactive oxygen species. METHODS: In the in vitro study, we measured the oxidative products of bilirubin formed through the action of O2- by the xanthine-xanthine oxidase system, either as free bilirubin or bilirubin-human serum albumin complex. In the clinical investigation, serum concentrations of (ZZ)-bilirubin (4Z, 15Z-bilirubin), the subfraction and biliverdin, and urinary propentdyopent absorption, were measured in blood and urine samples, respectively, collected from 30 5-day-old neonates with birth weights of 1500-3624 g who had been hospitalized at the Ehime Prefectural Hospital and who had not undergone phototherapy. RESULTS: In the in vitro study, a significant formation of propentdyopents was observed in aqueous solution. A statistically significant correlation was found between serum (ZZ)-bilirubin concentration and serum biliverdin concentration (r = 0.82, P < 0.0001), but not between serum (ZZ)-bilirubin concentration and urinary propentdyopent absorption. Serum (ZZ)- and serum (ZE)-bilirubin and biliverdin concentrations, and urinary propentdyopent absorption were compared between the groups with and without oxygen therapy. No significant differences were found in serum (ZZ)-bilirubin, serum (ZE)-bilirubin and biliverdin concentration, urinary propentdyopent absorption, serum biliverdin/serum (ZZ)-bilirubin, or urinary propentdyopent absorption/serum (ZZ)-bilirubin. Neither a decrease in serum bilirubin concentration nor an increase in serum biliverdin concentration and urinary propentdyopent absorption after oxygen therapy were demonstrated in the present study. CONCLUSIONS: In the in vitro study, we demonstrated for the first time that propentdyopents were produced from (ZZ)-bilirubin by the xanthine-xanthine oxidase system but biliverdin was not. In the in vivo study, serum biliverdin concentration and urinary propentdyopent absorption seem to have a different relationship to serum (ZZ)-bilirubin concentration in sick and early neonates.

Metabolism of bilirubin and riboflavin in the course of phototherapy for hyperbilirubinaemia in the newborns.

The study of bilirubin photochemical degradation in the presence of riboflavin has shown that, in this case, the primary process is photooxidation during which riboflavin acts as a receptor and bilirubin as a donor of electrons. The reaction proceeds under anaerobic conditions in an equilibrated manner, under aerobic ones as a catalytic process during which the catalyst, i. e., riboflavin is regenerated. The primary intermediate product is biliverdin in vitro, later this is further broken down. For the purpose of studying the mechanism of this process liquid chromatography and computerized technique based on solving nonlinear differential equations have been used. It appears that during phototherapy in the newborn infant there develops a decrease of the blood level of riboflavin reaching up to hypovitaminotic values and a transient biliverdin level elevation. Oral riboflavin administration may maintain this level within physiological range and at the same time shorten the necessary duration of phototherapy. In severe cases of hyperbilirubinaemia a shift of the ratio between flavin-adenin-dinucleotide and free riboflavin, as compared to the physiological state, has been recorded. The results of this work are discussed in the light of our current knowledge concerning the mechanisms of bilirubin breakdown during prototherapy.

Hyperbiliverdinemia in the bronze baby syndrome.

The bronze baby syndrome is an unusual complication of phototherapy for hyperbilirubinemia in the neonate. The pigment or pigments responsible for the discoloration in this syndrome have not yet been identified. Suspected pigments include photodegradation products of bilirubin and copper-porphyrins. We present here the case of a neonate with bronze baby syndrome whose serum had increased spectral absorbance in the region of maximum absorbance for biliverdin. We suggest that biliverdin pigments may also contribute to the "bronze" color associated with this syndrome.

Kinetics of the formation of biliverdin during the photochemical oxidation of bilirubin monitored by column liquid chromatography.

In the photochemical oxidation of bilirubin, biliverdin is formed as the primary product and is further degraded. This photooxidation is especially significant in the presence of riboflavin. Column liquid chromatography was used to monitor the kinetics of this reaction. The biliverdin concentration amounts to a maximum of ca. 38% of the total loss of bilirubin in experiments in vitro. It is probable that this mechanism is also operative during phototherapy. The formation of a product of the photooxidation of biliverdin that has not yet been identified has been observed; the product behaves as a dimer. A method for the determination of biliverdin in the blood of newborn infants has been developed. It has been found that the biliverdin content increases during hyperbilirubinaemia.