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.     

BILIRUBIN DISPLACING EFFECT OF STABILIZERS ADDED TO INJECTABLE PREPARATIONS OF HUMAN SERUM ALBUMIN

Abstract Stabilizers added to preparations of human serum albumin before heat treatment were tested for bilirubin displacing effect, using the peroxidase method. It was found that N-acetyltryptophan and sodium caprylate displace bilirubin from its complex with human serum albumin in vitro. The quantitative findings were used for a rough estimate of the effect of these substances on the free bilirubin concentration in blood plasma, expected when stabilized albumin preparations are given intravenously for prevention of kernicterus. The calculated effect is a delay of the decrease of free bilirubin concentration, or even a temporary increase. Sodium mandelate displaces less strongly.     

ALBUMIN ADMINISTRATION COMBINED WITH PHOTOTHERAPY IN TREATMENT OF HYPERBILIRUBINAEMIA IN LOW-BIRTH-WEIGHT INFANTS

ABSTRACT. Ebbesen, F., and Brodersen, R. (Department of Neonatology, Rigshospitalet, Copenhagen and Institute of Medical Biochemistry, Aarhus, Denmark). Albumin administration combined with phototherapy in treatment of hyperbilirubinaemia in low-birth-weight infants. Acta Paediatr Scand, 70:649,.–Fifty-nine jaundiced light treated newborn infants with low birth weight were studied. At onset of phototherapy 30 infants received 1 g human serum albumin per kg body weight as a 9 % solution containing sodium caprylate and N-acetyltryptophan as stabilizers. 29 infants did not receive human serum albumin and served as controls. Blood samples were taken before initiation of the therapy and again 24 and 48 h thereafter, and the following determinations were made: Serum concentrations of unconjugated bilirubin, albumin, reserve albumin for binding of bilirubin by the [^l4C]-MADDS method, packed cell volume and pH. Before infusion of albumin it was found that the binding fraction of serum albumin, i.e. the sum of the serum concentrations of bilirubin-albumin and reserve albumin, constituted about half of the total serum albumin concentration. The other half was non-binding, in agreement with previous findings in neonates. The effect of albumin therapy was mainly an unexpected increase of the non-binding fraction of serum albumin, while the increase of the serum reserve albumin concentration was small and the concentration of bilirubin-albumin was not changed.     

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.     

Bilirubin displacing effect of stabilizers added to injectable preparations of human serum albumin

Stabilizers added to preparations of human serum albumin before heat treatment were tested for bilirubin displacing effect, using the peroxidase method. It was found that N-acetyltryptophan and sodium caprylate displace bilirubin from its complex with human serum albumin in vitro. The quantitative findings were used for a rough estimate of the effect of these substances on the free bilirubin concentration in blood plasma, expected when stabilized albumin preparations are given intravenously for prevention of kernicterus. The calculated effect is a delay of the decrease of free bilirubin concentration, or even a temporary increase. Sodium mandelate displaces less strongly.     

RESERVE ALBUMIN AND BILIRUBIN TOXICITY INDEX IN INFANT SERUM

ABSTRACT. Reserve albumin concentration (the concentration of albumin available for binding of unconjugated bilirubin) was determined in 95 sera from 76 subjects by dialysis with ¹⁴C-monoacetyl diamino diphenyl sulfone (MADDS). An index, I of bilirubin toxicity in the plasma was calculated for each subject, based on the bilirubin and reserve albumin concentrations, the affinity of bilirubin for serum albumin, and the pH-dependent solubility of bilirubin in the plasma. The values of reserve albumin and of I varied significantly with gestational age, clinical condition (whether sick or well), and serum bilirubin level. The value of reserve albumin was decreased and I was increased in association with clinical factors (e. g., hyperbilirubinemia, hypoxia, acidosis, or sepsis) recognized as increasing the risk for bilirubin encephalopathy. The lowest values of reserve albumin and the highest values of I were found in the least mature and sickest infants.     

EFFECT OF PHOTOTHERAPY AND EXCHANGE TRANSFUSION ON PRIMARY BILE ACIDS IN THE SERUM OF HYPERBILIRUBINAEMIC NEWBORNS

ABSTRACT. Serum primary bile acid (cholic (CA) and chenodeoxycholic (CDCA) acid) concentrations were measured in 14 preterm and 11 full-term hyperbilirubinaemic newborns at the beginning and end of, and 24 and 72 hours following phototherapy. Only in the preterm newborns with gestational ages of 35-38 weeks there was a significant decrease of mean serum bile acid concentrations which could be shown 72 hours after the beginning of phototherapy. It can be hypothesized that the decrease was a result of a direct effect of light on the excretory liver function. Serum CA and CDCA concentrations were also measured in 5 hyperbilirubinaemic newborns at the beginning and end, and 24, 48 and 72 hours after the end of exchange transfusion. Exchange transfusion caused a clear immediate decrease in the mean serum primary bile acid concentrations. However, on day 2 after exchange transfusion the mean serum concentration of CA was about 150% and that of CDCA about 110% of the initial values. The most hyperbilirubinaemic newborns had extremely high primary bile acid serum concentrations before therapy. As bile acids compete with bilirubin for albumin binding it should be considered whether high bile acids in the serum of hyperbilirubinaemic newborns presuppose exchange transfusions.     

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.     

STUDIES ON PHOTOTHERAPY IN NEWBORN INFANTS

ABSTRACT: Windorfer, A., Faxelius, G. and Boréus, L. O. (Departments of Clinical Pharmacology and Paediatrics, Karolinska Hospital, Stockholm, Sweden). Studies on phototherapy in newborn infants: Influence on protein binding of bilirubin and salicylate and on activity of acetylsalicylic acid esterase. Acta Paediatr Scand 64:293, 1975.– Phototherapy of newborn infants with hyperbilirubinemia was shown to result in an increase in hematocrit values and in the activity of the erythrocyte enzyme acetylsalicylic acid esterase. The elevation of the enzyme activity also could be produced in light-treated rabbits and in vitro after illumination of blood from adult volunteers. The binding of bilirubin to serum albumin and of salicylate to plasma proteins did not alter, nor did the concentrations of albumin or total proteins in plasma. It is concluded that light does not increase the unbound fraction of bilirubin in blood.     

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.     

Enhancement of the phototherapy effect by albumin in Gunn rats. Investigations on the optimal albumin dose

In pharmacokinetic studies we proved that the efficacy of phototherapy in jaundiced Gunn rats was enhanced by administration of human albumin. The albumin injections provoked a bilirubin shift from extravascular compartments into the circulation.In this study, calculations on the optimal albumin dose were carried out. After s.c. injections a good effect could be ascertained with doses between 2 and 4 g/kg body weight. After i.v. injections effective plasma concentrations could be maintained for an acceptable time by about 2 g albumin/kg body weight. When overdoses were given, a considerable amount of albumin was eliminated very quickly—especially after i.v. injections with the rapid rise of the plasma level. Clinical implications are discussed.     

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.     

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.     

D-PENICILLAMINE, A NON-BILIRUBIN-DISPLACING DRUG IN NEONATAL JAUNDICE

Abstract. Brodersen, R., Lakatos, L. and Karmazsin, L. (Institute of Medical Biochemistry, University of Aarhus, Denmark, and Pediatric Clinic, University Medical School, Debrecen, Hungary). D-penicillamine, a non-bilirubin-displacing drug in neonatal jaundice. Acta Paediatr Scand, 69:31, 1980.—D-penicillamine, a drug used clinically for the treatment of neonatal hyperbilirubinaemia, was tested for interference with the binding of bilirubin to human serum albumin by three methods: 1) The peroxidase technique, investigating the effect of D-penicillamine on the equilibrium concentration of unbound bilirubin in a solution containing a molar excess of albumin; 2) the MADDS method, measuring the concentration of vacant bilirubin binding site on albumin in a solution of pure albumin, or infant blood serum, with added D-penicillamine; and 3) injection of D-penicillamine into Gunn rats and determination of any decrease of plasma biiirubin which would be caused by displacement of the pigment. Results were negative in all cases. Quantitatively, the doses of D-penicillamine used clinically cannot displace bilirubin from its binding to albumin. The ameliorating effect on hyperbilirubinaemia in the newborn must be due to some other mechanism     

Effect of Sulfisoxazole on Bilirubin-Albumin Binding in Gunn Rats

Hyperbilirubinemic Gunn rats were used to study sulfisoxazole-mediated displacement of bilirubin from albumin. Bilirubin not bound to albumin or unbound bilirubin, which can enter the brain and cause damage, was measured by the automated peroxidase micromethod using the UB-Analyzer. When sulfisoxazole was added to the rat serum in vitro, it showed measurable displacing competition with bilirubin for the binding sites on albumin. The bilirubin titration curves of hyperbilirubinemic sera, after injection of sulfisoxazole, showed a shift to the left indicating that the drug effectively lowered the capacity of serum albumin to bind to bilirubin. The bilirubin binding affinity of the first binding site on the albumin decreased with the administration of 25 mg/kg or more of sulfixazole. The serum concentration of both total bilirubin and unbound bilirubin decreased after the injection of sulfisoxazole suggesting their diffusion into extravascular compartments.     

高胆红素血症新生大鼠脑组织Na~+-K~+-ATP酶活力的研究

目的：探讨胆红素神经毒性的作用环节,研究胆红素脑病发病机理。方法：72只新生7d SD大鼠随机分为对照组(C组)和实验组(T组),T组又根据腹腔注射胆红素剂量的依次增加分为T1,T2,T3,T4和T5组。检测各组脑组织和血清中胆红素含量,定磷法测定脑组织中Na+ K+ ATP酶(Na+ K+ ATPase)活力。结果：给药后4 h和8 h均随着腹腔注射胆红素量的增加,血清总胆红素浓度、脑组织内胆红素含量逐渐增加,脑组织Na+ K+ ATPase活力则逐渐降低,除T1组外余各组与对照组相比差异均有显著性(P0.05),但脑组织内胆红素含量、Na+ K+ ATPase活力除T1组外各组4 h,8 h之间差异有显著性(P0.05),脑组织内胆红素含量与Na+ K+ ATPase活力呈负相关(r=-0.86,P<0.01)。结论：脑组织内胆红素对Na+ K+ ATPase活力有抑制作用。     

儿童初发急性淋巴细胞白血病的血清前白蛋白测定

通过对前白蛋白 (PA)的检测 ,了解其作为急淋对肝浸润的敏感指标。本文对 2 2例患儿在治前与其中完全缓解后的 1 7例患儿和 30例健康儿童对照组 ,通过免疫比浊法测定 PA,并检测其它肝功能值。结果显示 :2 2例治前患儿 ,9例 PA下降 (与对照组 PA t检验 P<0 .0 1 ) ,3例白蛋白降低 ,1例谷丙、6例谷草转氨酶升高 ,1例总胆红素上升 ;治前 PA异常的例数高于肝功能其它值异常的例数。其中完全缓解后的1 7例患儿 ,PA及肝功能其它值则均在正常范围。因此 ,通过 PA的检测 ,发现治前急淋对肝浸润时 ,PA的变化远较肝功能其它值的变化为明显     

The effect of bilirubin on the surface charge and aggregation tendency of platelets in cord blood

Unbound unconjugated bilirubin markedly increased the negative electrophoretic mobility of washed platelets from cord blood, even at concentration as low as 1.5 mg/dl. The increase in negativity could be abolished by washing the platelets. Aggregation of platelets was observed in parallel with the increase in negativity. These actions of bilirubin required calcium ions, which could not be replaced by magnesium ions. The results suggest attachment of the negative bilirubin molecules onto the platelet surface, probably through calcium ions, leading to platelet aggregation.Plasma components inhibited the actions of bilirubin on platelets up to 19.5 mg/dl bilirubin concentration.The effect of bilirubin on platelets was also investigated in the presence of albumin. When the saturation of albumin with bilirubin was exceeded, platelet negativity increased. However, the fraction of free bilirubin exceeding the albumin saturation point has not the same effect as truly free bilirubin at that concentration in the complete absence of albumin. The results indicate that albumin molecules loosly adsorbed onto the surface may protect the platelets against the attachment of free bilirubin. This protection, however, might be impaired by acidosis, which is frequently combined with hyperbilirubinaemia in the sick newborn. It is suggested that bilirubin could contribute to the haemostatic abnormalities in sick babies by acting on the platelet surface.     

Albumin in Management of Neonatal Hyperbilirubinaemia

The effect of albumin priming on the plasma volume, intravascular bilirubin, and HBABA-binding capacity during the subsequent 2 to 4½ hours before exchange transfusion was studied in 7 jaundiced neonates with no haemolytic disease. After priming with albumin at a dose of 1·75 g/kg body weight as a 10% solution, there was a marked increase in plasma volume as well as total intravascular bilirubin and HBABA-binding capacity. However, the serum bilirubin concentration fluctuated only very slightly, largely because of the dilution effect of the expanded plasma volume. Both albumin and bilirubin gradually diffused out of the intravascular compartment again, but at the end of the observation period there was still a net gain of both.In another 20 neonates, the efficiency of exchange transfusion in removing bilirubin was compared. The efficiency was decreased by early albumin priming and enhanced by enriching the donor''s blood with albumin.It is concluded that albumin offers immediate and short-term protection against bilirubin toxicity, and albumin-priming should be useful in situations where the babies are admitted with high bilirubin levels and blood is not immediately available for exchange transfusion. Because of its effect on the plasma volume, albumin is not recommended for babies who are already hypervolaemic. If albumin is used to increase the efficiency of exchange transfusion, it should be given together with donor''s blood or shortly before the procedure.     

Bilirubin-albumin-binding function of 2 human albumin preparations (placental and plasma). Comparison of their efficacy in the icteric premature infant

Two albumin preparations obtained by Cohn fractionation of either plasma of blood donors (plasmatic albumin) or human placental blood (placental albumin) were studied in vitro and in vivo regarding their bilirubin-binding function. Analysis of this function during the industrial processing of the two preparations indicated that alcoholic fractionation and, to a lesser extent, stabilizers, were responsible for the decrease of (a) the association constants between albumin and bilirubin, (b) bilirubin-binding capacity of albumin. Unexpectedly, improvement of bilirubin-binding parameters was observed after the final heating stage. Stabilizers were reversibly bound as suggested by a further improvement of binding function seen after a brief contact of the preparations with red blood cells. The changes were similar for the two preparations. Fifty-one sick premature hyperbilirubinemic neonates were randomly infused either with placental or plasmatic albumin (1.5 g/kg). Albuminemia, bilirubinemia, erythrocytic bilirubin, unbound bilirubin (peroxidase method) were evaluated before and 3 hours after infusion. Improvement of bilirubin-binding parameters was frequently observed but without clear-cut relation with change in bilirubin/albumin molar ratio. No difference was noted between the two albumin preparations. In spite of a decrease of their association constants with bilirubin, the two albumins retained a high binding potency for bilirubin in vivo.