BILIRUBIN, RESERVE ALBUMIN FOR BINDING OF BILIRUBIN AND pH IN PLASMA DURING PHOTOTHERAPY (ORDINARY AND DOUBLE LIGHT) OF TERM NEWBORN INFANTS

ABSTRACT. Ebbesen, F. (Department of Neonatology, Rigshospitalet, Copenhagen, Denmark). Bilirubin, reserve albumin for binding of bilirubin and pH in plasma during phototherapy (ordinary and double light) of term newborn infants. Acta Paediatr Scand, 70:223, 1981. –Forty-five term newborn infants with uncomplicated hyperbilirubinaemia were treated continuously with phototherapy for 24 hours. Twenty-eight infants received double light treatment and 17 infants ordinary phototherapy. During both treatments a significant decrease in the serum unconjugated bilirubin concentration, a significant increase in the serum reserve albumin concentration for binding of bilirubin determined by the [¹⁴C] MADDS method, and a significant decrease in the index of serum bilirubin toxicity occurred. The changes in these parameters were significantly greater during the double light treatment than during the ordinary phototherapy. During the treatment the fall in index was constant. No significant change in plasma pH was seen. Thus, the study gives further evidence that the risk of bilirubin encephalopathy is reduced by phototherapy and that double light treatment is in the respect superior to ordinary phototherapy. Prior to phototherapy the molar ratio in serum of unconjugated bilirubin plus reserved albumin for binding of bilirubin to albumin was only 0.60, on average, and during the treatment the increase in the serum reserve albumin concentration was less than the decrease in the serum bilirubin concentration. This can be explained either by the presence in infant serum of an unknown ligand interfering competitively or allosterically in the binding of MADDS and bilirubin to albumin, or by the existence of a foetal albumin with a lower affinity for MADDS than adult albumin.     

Skin colour and bilirubin in neonates.

The correlation between the yellow colour of the skin and serum bilirubin concentration, reserve albumin concentration, and pH was investigated in 76 icteric neonates. Significant linear correlation existed between yellow colour of the skin and serum bilirubin concentration, reciprocal of the reserve albumin concentration, and the squared hydrogen ion concentration. Furthermore, the basic yellowness of the skin at birth correlated linearily with the yellow colour of the skin measured when the child became jaundiced. The results support the proposed hypothesis that bilirubin is transferred from plasma to skin through two different mechanisms: (a) leakage of bilirubin-albumin complexes into extravascular spaces and (b) precipitation of bilirubin acid in phospholipid membranes. The latter mechanism suggests that measurement of the yellow colour of the skin may be a better predictor of brain damage than the serum bilirubin concentration and thus be of clinical utility. Measurement of the yellow colour of the skin as a method of obtaining serum bilirubin concentration is unreliable.     

Albumin binding properties in relation to bilirubin and albumin concentrations during the first week of life

In 19 non-jaundiced and 22 jaundiced neonates, the serum albumin and bilirubin concentrations were measured during the first week of life. Some of the neonates were followed longitudinally. The albumin binding properties were evaluated by determining the reserve albumin concentration for monoacetyldiaminodiphenyl sulphone (MADDS), a deputy ligand for bilirubin. The reserve albumin concentration for MADDS increased with postnatal age. The reason for this increase is still unexplained. There was an inverse relation between the bilirubin and the reserve albumin concentrations, but when the bilirubin concentration increased by 1 mumol/l, the reserve albumin concentration for MADDS decreased by only 0.2 mumol/l. This shows that the reserve albumin concentration for MADDS does not give a direct measure of the bilirubin binding ability of the serum albumin molecule. In spite of this, it is still possible that a low reserve albumin concentration for MADDS is a risk factor for bilirubin encephalopathy.     

Albumin Binding Properties in Relation to Bilirubin and Albumin Concentrations during the First Week of Life

ABSTRACT. In 19 non-jaundiced and 22 jaundiced neonates, the serum albumin and bilirubin concentrations were measured during the first week of life. Some of the neonates were followed longitudinally. The albumin binding properties were evaluated by determining the reserve albumin concentration for monoacetyldiaminodiphenyl sulphone (MADDS), a deputy ligand for bilirubin. The reserve albumin concentration for MADDS increased with postnatal age. The reason for this increase is still unexplained. There was an inverse relation between the bilirubin and the reserve albumin concentrations, but when the bilirubin concentration increased by 1 μmol/l, the reserve albumin concentration for MADDS decreased by only 0.2 μmol/l. This shows that the reserve albumin concentration for MADDS does not give a direct measure of the bilirubin binding ability of the serum albumin molecule. In spite of this, it is still possible that a low reserve albumin concentration for MADDS is a risk factor for bilirubin encephalopathy.     

COMPARISON BETWEEN TWO PREPARATIONS OF HUMAN SERUM ALBUMIN IN TREATMENT OF NEONATAL HYPERBILIRUBINAEMIA

ABSTRACT. Thirty-six newborn infants with normal birth weights and with uncomplicated hyperbilirubinaemia, treated with light, were studied. At onset of phototherapy the infants received intravenously 1 g human serum albumin (HSA) per kg body weight as a 9 % solution. Two different preparations of HSA were used and compared. One of these, HSA_I, contained sodium caprylate and N-acetyltryptophan, 5 mmol/1 of each, as stabilizers. HSA_II contained only caprylate, 5 mmol/1. Nineteen infants received HSA_I and seventeen infants HSA_II. The reserve albumin for binding of bilirubin, measured by the [¹⁴C] MADDS method, was low in both preparations in vitro. During the infusion, the serum concentrations of albumin and reserve albumin increased and the serum unconjugated bilirubin concentration decreased, resulting in a fall in the index of plasma bilirubin toxicity in all infants. After completion of the infusion, the serum concentrations of albumin and reserve albumin declined, and a slight rise in index occurred. The increase in the serum reserve albumin concentration was markedly higher during infusion of HSA_II than of HSA_I. It is concluded that infusion of both HSA preparations during phototherapy provides an immediate protection against bilirubin encephalopathy. HSA_I is inferior to HSA_II, probably due to its content of N-acetyltryptophan.     

EFFECT OF EXCHANGE TRANSFUSION ON SERUM RESERVE ALBUMIN FOR BINDING OF BILIRUBIN AND INDEX OF SERUM BILIRUBIN TOXICITY

ABSTRACT. Ebbesen F. (Department of Neonatology, Rigshospitalet, Copenhagen, Denmark). Effect of exchange transfusion on serum reserve albumin for binding of bilirubin and index of serum bilirubin toxicity. Acta Paediatr Scand, 70:643,.–Seventeen newborn infants, who received their first exchange transfusion due to hyperbilirubinaemia and/or rhesus haemolytic disease, were studied. The exchange transfusions were performed with fresh, citrated blood. During the exchange transfusion a marked increase in the serum reserve albumin concentration for binding of bilirubin measured by the [^,4C]-MADDS method was observed, followed by a smaller decrease after the transfusion. Plasma pH increased both during and after the exchange transfusion. During the exchange transfusion a drastic fall in index of serum bilirubin toxicity was observed, followed by a smaller increase after the transfusion. Citrate was not found to interfere in the binding of bilirubin to albumin. The results are in agreement with the clinical finding that an exchange transfusion performed with fresh, citrated blood effectively reduces the risk of bilirubin encephalopathy. The ratio in serum of binding albumin, i.e. bilirubin plus reserve albumin, to total albumin failed to be increased by the exchange transfusion, and a decrease occurred after the transfusion. These findings indicate the presence in infant serum of non-binding albumin. Donor albumin with intact binding potential is partly transformed into the non-binding variety in the course of one hour after the transfusion. In the most severely rhesus sensitized infant a drastic decline of the serum albumin binding capacity was seen during the first day of life.     

Reduced Albumin Binding of MADDS–a Measure for Bilirubin Binding–in Sick Children

ABSTRACT. The reserve albumin concentration for binding of MADDS (monoacetyldiaminodiphenyl sulphone) in plasma is used as a measure of the reserve albumin concentration for binding of unconjugated bilirubin. The aim of the present study was to investigate whether a reduction in the reserve albumin concentration for binding of MADDS could exist in sick children after 5 months of age, where the bilirubin binding properties of the albumin has reached the adult level. The material included 75 children, 1-15 years of age with mild infections, servere bacterial infections, acute viral hepatitis, chronic hepatic diseases or uraemia, and a control group of 22 healthy children. The reserve albumin concentration was significantly lower in children with severe bacterial infections, acute viral hepatitis, and uraemia, than in healthy children ( p <0.01), while the reserve albumin concentration in children with mild infections and chronic hepatic diseases did not differ significantly from that of the control group ( p > 0.05). The total albumin concentration in plasma in either of the groups of sick children did not differ significantly from that of the healthy children. The plasma concentration of unconjugated bilirubin was elevated in the group of children with acute viral hepatitis, but not enough to influence the concentration of reserve albumin for binding of MADDS to a significant degree. The reserve albumin concentration was significantly lower in children with acute viral hepatitis than in children with severe bacterial infections ( p <0.05).     

Reduced albumin binding of MADDS--a measure for bilirubin binding--in sick children

The reserve albumin concentration for binding of MADDS (monoacetyldiaminodiphenyl sulphone) in plasma is used as a measure of the reserve albumin concentration for binding of unconjugated bilirubin. The aim of the present study was to investigate whether a reduction in the reserve albumin concentration for binding of MADDS could exist in sick children after 5 months of age, where the bilirubin binding properties of the albumin has reached the adult level. The material included 75 children, 1-15 years of age with mild infections, severe bacterial infections, acute viral hepatitis, chronic hepatic diseases or uraemia, and a control group of 22 healthy children. The reserve albumin concentration was significantly lower in children with severe bacterial infections, acute viral hepatitis, and uraemia, than in healthy children (p less than 0.01), while the reserve albumin concentration in children with mild infections and chronic hepatic diseases did not differ significantly from that of the control group (p greater than 0.05). The total albumin concentration in plasma in either of the groups of sick children did not differ significantly from that of the healthy children. The plasma concentration of unconjugated bilirubin was elevated in the group of children with acute viral hepatitis, but not enough to influence the concentration of reserve albumin for binding of MADDS to a significant degree. The reserve albumin concentration was significantly lower in children with acute viral hepatitis than in children with severe bacterial infections (p less than 0.05).     

The Serum Reserve Albumin Concentration for Monoacetyldiaminodiphenyl Sulphone and Auditory Evoked Responses during Neonatal Hyperbilirubinaemia

ABSTRACT. Auditory brainstem evoked responses (ABR) were recorded in 9 neonates with hyperbilirubinaemia. Pathological recordings were found in two children showing absence of waves and prolonged latencies. There was no correlation between latencies to waves and the total serum bilirubin concentration. The serum reserve albumin concentration for monoacetyldiaminodiphenyl sulphone (MADDS) was, however, inversely related to the latencies in the ABR recordings. Our findings suggest that the binding properties of serum albumin contribute to the risk of bilirubin toxicity and that, in this study, the reserve albumin concentration for MADDS seemed to be of Heater significance than the total bilirubin concentration.     

LOW RESERVE ALBUMIN FOR BINDING OF BILIRUBIN IN NEONATES WITH DEFICIENCY OF BILIRUBIN EXCRETION AND BRONZE BABY SYNDROME

ABSTRACT. The plasma reserve albumin concentration for binding of bilirubin was found to be low in four newborn infants with deficiency of bilirubin excretion, of whom two had the bronze baby syndrome. Thus, the risk of bilirubin encephalopathy was increased. Also the ratio of binding fraction of albumin, i. e. unconjugated bilirubin plus reserve albumin, to total albumin was low. Possible causes of the low reserve albumin concentration and the ratio are discussed.     

The serum reserve albumin concentration for monoacetyldiaminodiphenyl sulphone and auditory evoked responses during neonatal hyperbilirubinaemia

Auditory brainstem evoked responses (ABR) were recorded in 9 neonates with hyperbilirubinaemia. Pathological recordings were found in two children showing absence of waves and prolonged latencies. There was no correlation between latencies to waves and the total serum bilirubin concentration. The serum reserve albumin concentration for monoacetyldiaminodiphenyl sulphone (MADDS) was, however, inversely related to the latencies in the ABR recordings. Our findings suggest that the binding properties of serum albumin contribute to the risk of bilirubin toxicity and that, in this study, the reserve albumin concentration for MADDS seemed to be of greater significance than the total bilirubin concentration.     

Relationship of unbound bilirubin concentration to reserve albumin-binding concentration for bilirubin in human neonatal plasma

We examined the relationship of plasma unbound bilirubin concentration and reserve albumin-binding concentration for bilirubin in a sample of 545 neonates. Plasma unbound bilirubin concentration and total bilirubin-binding concentration were determined with the peroxidase assay. Contrary to published reports, we found that plasma unbound bilirubin concentration and plasma reserve albumin for bilirubin-binding concentration are highly correlated (r = -0.706; p less than 0.001) and that the relationship between these two parameters is dependent upon the total bilirubin-binding concentration. That these measured parameters correlate in the same manner as predicted for free bilirubin and free albumin by the law of mass action, suggests that these measurements may be meaningful.     

THE RELATIONSHIP BETWEEN SERUM BILIRUBIN AND RESERVE ALBUMIN FOR BINDING OF BILIRUBIN DURING PHOTOTHERAPY OF PRETERM INFANTS

ABSTRACT. Ebbesen, F. (Department of Neonatology, Rigshospitalet, Copenhagen, Denmark). The relationship between serum bilirubin and reserve albumin for binding of bilirubin during phototherapy of preterm infants. Acta Paediatr Scand, 70:405, 1981.–Thirty-four preterm newborn infants suffering from uncomplicated hyperbilirubinaemia were studied. The infants received ordinary phototherapy continuously during 48 hours. The serum unconjugated bilirubin concentration decreased significantly during the treatment, and a significant correlation between the changes in the serum bilirubin concentration and the changes in the serum reserve albumin concentration for binding of bilirubin measured by the [¹⁴C]MADDS method was found. The regression coefficients were -0.50 and -0.48 after 24 and 48 hours of treatment, respectively. Thus, it can be concluded that the risk of bilirubin encephalopathy is reduced by phototherapy in preterm infants.     

Ceftriaxone effect on bilirubin-albumin binding

The effect of ceftriaxone on bilirubin-albumin binding was measured in vitro using the peroxidase method with human serum albumin and a dialysis rate method with adult and newborn serum. Ceftriaxone competes with bilirubin for binding to human serum albumin; the displacement constant is 1.5 X 10(4) L/mol. Therapeutic levels of ceftriaxone decrease the reserve albumin concentration in newborn serum by 39%. These results indicate that ceftriaxone may increase the risk of bilirubin encephalopathy in jaundiced premature infants.     

EFFECTS OF STABILIZERS IN PREPARATIONS OF HUMAN SERUM ALBUMIN ON THE SERUM BILIRUBIN IN GUNN RATS

ABSTRACT. Commercially available preparations of human serum albumin (HSA) containing stabilizers (i.e. 16 mmol/I Na caprylate plus 16 mmol/I Na N-acetyl- dl -tryptophan) were injected either s.c., i.p. or i.v. into homozygous infant Gunn rats. 30 min and 3 hours after s.c. injection, a serum bilirubin decline which surpassed dilution by the injected volume could be ascertained. It was mainly caused by N-acetyl- dl -tryptophan since s.c. injections of appropriate amounts of this substance alone or a mixture of both components of the stabilizer without HSA brought about similar results. HSA without these stabilizers had not such an effect. It is postulated that under these conditions Na N-acetyl- dl -tryptophanate displaced bilirubin from albumin bonds. It became obvious that after s.c. injection equilibration of HSA between skin and plasma was delayed, whereas Na N-acetyl- dl -tryptophan was rapidly transported to the blood. As for Na caprylate, a displacing effect of short duration could not be excluded by the experimental arrangement used, since the metabolism of the substance in the rat is very fast. When HSA and the stabilizers entered the plasma simultaneously (i.v. injection) no effect on serum bilirubin concentration could be proved 30 min and 3 hours later. All the bilirubin and the Na N-acetyl- dl -tryptophan present in the plasma at that time can be bound to the large amount of albumin which is directly given into the circulation of the animal. 30 min after i.p. injection of HSA preparations containing stabilizers a small decrease of serum bilirubin concentration could be recognized. It was less pronounced and less persisting than after s.c. injection. Probably equilibration of HSA between peritoneum and plasma went on faster than between skin and plasma. Only for a short period a lack of albumin binding sites in the plasma of the rat pointed to a surplus of Na N-acetyl- dl -tryptophan.     

POSTNATAL CHANGES IN THE ABILITY OF PLASMA ALBUMIN TO BIND BILIRUBIN

ABSTRACT. The plasma concentrations of total albumin, unconjugated bilirubin and reserve albumin for bilirubin binding were determined in 407 healthy infants of various age up to eight days. The albumin reserve was measured using monoacetyldiaminodiphenyl-sulfone (MADDS) as a deputy ligand for bilirubin. The fraction of albumin capable of binding bilirubin was calculated as the sum of the concentrations of bilirubin and reserve albumin, divided by the total albumin concentration. Our data showed that this fraction was low (average 0.36) and did not change during the first 24 hours of life, and in this period it was independent of the maturity of the infant, as expressed by its birth weight or gestational age. From about 24 hours of life, the fraction began to increase. This increase came to an end about 60 hours after birth, and no further changes were seen during the following five days. The level of the bilirubin-binding fraction reached 60 hours after birth was related to the maturity of the infant: It increased with increasing birth weight up to 3000 g and with increasing gestational age up to 275 days, when on an average it was about 0.58. The fraction of binding albumin was independent of the sex.     

Postnatal changes in the ability of plasma albumin to bind bilirubin

The plasma concentrations of total albumin, unconjugated bilirubin and reserve albumin for bilirubin binding were determined in 407 healthy infants of various age up to eight days. The albumin reserve was measured using monoacetyldiaminodiphenyl-sulfone (MADDS) as a deputy ligand for bilirubin. The fraction of albumin capable of binding bilirubin was calculated as the sum of the concentrations of bilirubin and reserve albumin, divided by the total albumin concentration. Our data showed that this fraction was low (average 0.36) and did not change during the first 24 hours of life, and in this period it was independent of the maturity of the infant, as expressed by its birth weight or gestational age. From about 24 hours of life, the fraction began to increase. This increase came to an end about 60 hours after birth, and no further changes were seen during the following five days. The level of the bilirubin-binding fraction reached 60 hours after birth was related to the maturity of the infant: It increased with increasing birth weight up to 3000 g and with increasing gestational age up to 275 days, when on an average it was about 0.58. The fraction of binding albumin was independent of the sex.     

The increase of yellow skin colour beyond that of serum bilirubin: A proposed indicator of risk for bilirubin encephalopathy in the newborn

Forty-seven newborn infants with 1 min Apgar score < 7 were studied. On the third postnatal day the following measurements were made: yellow skin colour, serum bilirubin concentration, reserve albumin concentration and plasma pH. Given the bilirubin concentration and the regression curve between the yellow skin colour and the bilirubin concentration, Δ-TcB was calculated as the difference between measured yellow skin colour and the expected yellow skin colour. There was a negative correlation between Δ-TcB and Apgar score (P = 0.003), pH (P = 0.026) and reserve albumin concentration (P = 0.045). Fourteen of the included newborns had central nervous system symptoms in the days just following birth. A tendency towards higher Δ-TcB was noted in this group (P = 0.08). The results suggest that further study of Δ-TcB determination as a tool in the assessment of the icteric newborn infant is justified.     

The risk of bilirubin encephalopathy, as estimated by plasma parameters, in neonates strongly suspected of having sepsis

The study comprises 18 mature newborns, strongly suspected of having sepsis, and a control group of 18 mature, healthy newborns with the same postnatal age. The object of the investigation was to compare the risk of development of bilirubin encephalopathy between the two groups, as estimated by plasma parameters. The sepsis group had significantly lower reserve albumin concentration for binding of MADDS ( p <0.01) and significantly lower total albumin concentration ( p <0.01). No significant differences were observed in unconjugated bilirubin concentration and plasma pH. It is suggested that mature newborns with sepsis have a slightly increased risk of developing bilirubin encephalopathy.     

Comparison of three methods evaluating the saturation of serum bilirubin transporters in children with type I Crigler Najjar disease

The assessment of reserve bilirubin binding capacity of serum in 12 children with type I Crigler-Najjar syndrome was performed by 3 biochemical methods. Two of them use chromatography techniques: thin layer on Sephadex or high performance liquid; they allow the determination of the saturation rate of all serum bilirubin carriers. The third method only evaluates the major carrier, albumin, using the molar bilirubin-albumin ratio. An identical correlation coefficient of 0.90 has been obtained when comparing with the chromatographic method on thin layer, respectively 20 determinations using high performance liquid chromatography and 122 measurements of the bilirubin/albumin ratio. In practice, we advise a first, simple and rapid determination of the bilirubin/albumin ratio in case of a sudden aggravation of the jaundice in the children with Crigler-Najjar type I disease. The chromatographic methods will be used in a second step, or in order to survey the phototherapy treatment that these children receive.