Determination of bilirubin in liver homogenate with diazotized p-lodoaniline

Mertens, Bruno B.E.; Vijver, M. Van de; Heirwegh, K. P. M.

doi:10.1016/0003-2697(72)90080-2

Cited by 12 publications

(3 citation statements)

References 9 publications

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“…Bilirubin concentration was measured in a 20% liver homogenate, prepared in a sucrose (0.25 mol/l) and EDTA (1 mmol/l) solution, by the method of Mertens, Van de Vijver & Heirwegh (1972), with the diazonium salt of piodoaniline, and the total hepatic content was expressed per l00g body weight. The protein concentration of liver homogenates was estimated by the method of Lowry, Rosebrough, Farr & Randall (195 1) and hepatic cytochrome P-450 content assayed according to the method detailed by Black, Perrett & Cartter (1973).…”

Section: Methodsmentioning

confidence: 99%

The Role of Dietary Lipid in the Regulation of Unconjugated Hyperbilirubinaemia in Gunn Rats

1979

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1. The purpose of this study was to characterize the mechanisms of diet-induced hyperbilirubinaemia in Gunn rats with emphasis on the role of lipids, and to examine their relationship with regard to fasting hyperbilirubinaemia. 2. A lipid-free normocaloric diet produced a threefold increase in plasma bilirubin concentration (baseline 109.4 mumol/l), which was maximal by 10 days and thereafter remained constant. The level of hyperbilirubinaemia attained was not influenced by fasting or phenobarbitone, and returned to baseline concentration within 10 days of resuming a normal diet. 3. Determination of hepatic bilirubin showed that the magnitude of the hepatic bilirubin pool was increased by the lipid-free diet but was unchanged by fasting. Hepatic ligandin concentrations were comparable in fasted Gunn rats and those fed normal or lipid-free diets, although total hepatic ligandin was reduced in the fasted animals. 4. The hyperbilirubinaemic effect of the lipid-free diet was largely reversed by the inclusion of 10% lipid in the diet and was affected to a lesser extent by 5% lipid. Similar reductions in plasma bilirubin concentration were observed with a variety of other lipids (10%), regardless of their fatty acid chain length or degree of saturation. 5. In fasting animals a direct correlation was observed between plasma bilirubin and free fatty acid concentrations and insulin levels were greatly depressed, whereas in those fed on the lipid-free diet no significant changes were evident in plasma concentrations of free fatty acids or insulin. 6. Plasma bilirubin concentration was unrelated to alterations in plasma triglycerides produced by the administration of clofibrate. However, an unexplained decrease in plasma bilirubin (40%) without a significant change in triglycerides was noted when clofibrate was added to the lipid-free diet. 7. Analysis of kinetic data obtained from [14C]bilirubin clearance studies revealed that hyperbilirubinaemia associated with the lipid-free diet reflected a marked reduction (60%) in plasma clearance with no change in bilirubin turnover. This was accompanied by a relative redistribution of bilirubin from the extravascular pool to the plasma pool. 8. Although these studies indicate that fasting and the withdrawal of dietary lipid have some similar effects on bilirubin metabolism, it seems likely that different mechanisms are responsible for the hyperbilirubinaemia.

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Section: Methodsmentioning

confidence: 99%

The Role of Dietary Lipid in the Regulation of Unconjugated Hyperbilirubinaemia in Gunn Rats

1979

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“…Bilirubin was estimated in serum by the technique of Michaelsson, Nosslin, and Sjohn (1965) or by the method of Van Roy, Meuwissen, De Meuter, and Heirwegh (1971), modified by the addition of the anti-oxidant 2,6-di-tert-butyl-p-cresol (Mertens, Van de Vijver, and Heirwegh, 1972). The latter method was also used for bilirubin estimations in liver homogenate, supernatant, and protein fractions.…”

Section: Methodsmentioning

confidence: 99%

Sites of competition in the selective hepatic uptake of rifamycin-SV, flavaspidic acid, bilirubin, and bromsulphthalein

Kenwright¹,

Levi²

1974

Gut

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“…The pH remained constant at 8.6. Bilirubin concentrations were determined in triplicate at the beginning and end of the incubation by the method of Mertens et al (1972), with diazotized p-iodoaniline, by using 0.5 ml of medium. The enzyme activity was expressed in munits/mg of protein, 1 munit being equivalent to 1 nmol of bilirubin degraded/min.…”

Section: Bilirubin Oxidase Assaymentioning

confidence: 99%

Enzymic oxidation of unconjugated bilirubin by rat liver

Cárdenas-Vázquez¹,

Yokosuka²,

Billing³

1986

Biochemical Journal

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The presence of the enzyme bilirubin oxidase, which degrades bilirubin in vitro, was demonstrated in the liver. Subcellular-fractionation experiments indicate that bilirubin oxidase is located in both the inner and outer membranes of the mitochondria. The mean rate of the reaction is 1.57 +/- 0.38 (S.D.) nmol of bilirubin degraded/min per mg of mitochondrial protein (munits/mg of protein). With respect to the overall breakdown of bilirubin, the enzyme has a Km' of 136 microM-bilirubin and a Vmax.' of 9.13 munits/mg of protein. Its activity is influenced by the ionic strength of the media and is inhibited by KCN, thiol reagents, NADH and albumin. The enzyme is aerobic, and between 1 and 1.5 mol of O2 are consumed per mol of bilirubin degraded. The products of the reaction include propentdyopents. The hepatic bilirubin oxidase activity of the jaundiced Gunn-rat liver is not significantly different from that of the Sprague-Dawley rat, and it is not induced by beta-naphthoflavone.

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Determination of bilirubin in liver homogenate with diazotized p-lodoaniline

Cited by 12 publications

References 9 publications

The Role of Dietary Lipid in the Regulation of Unconjugated Hyperbilirubinaemia in Gunn Rats

The Role of Dietary Lipid in the Regulation of Unconjugated Hyperbilirubinaemia in Gunn Rats

Sites of competition in the selective hepatic uptake of rifamycin-SV, flavaspidic acid, bilirubin, and bromsulphthalein

Enzymic oxidation of unconjugated bilirubin by rat liver

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