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An 18O double-labelling study of haemoglobin catabolism in the rat.

ABSTRACT: The degradation of haemoglobin to bilirubin in the rat was investigated by 18O labelling of the molecular oxygen consumed by the animal. The oxygen atoms incorporated into bilirubin were derived from two different oxygen molecules. Implications of this finding for the mechanism of haem catabolism in vivo are discussed; both verdohaem and a dioxygen-bridged compound appear to be excluded at intermediates.

SUBMITTER: Brown SB

PROVIDER: S-EPMC1165773 | biostudies-other | 1975 Sep

REPOSITORIES: biostudies-other

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The mechanism of haem catabolism. Bilirubin formation in living rats by [18O]oxygen labelling.

Project description:1. The pathway of haem breakdown in living rats was studied by using 18O in the oxygen that the animals consumed. By cannulation of the common bile duct and collection of bile, labelled bilirubin was isolated and its mass spectrum determined. One set of results was obtained for a rat to which haemoglobin had been intravenously administered and another set obtained for a rat that was not given exogenous haem. Isomerization of bilirubin IXalpha to the XIIIalpha and IIIalpha isomers did not occur to any significant extent. The 18O-labelling pattern obtained in the bilirubin was consistent with a Two-Molecule Mechanism, whereby the terminal lactam oxygen atoms of bilirubin are derived from different oxygen molecules. The consequences of this mechanism are discussed in terms of the possible intermediates of the catabolic pathway. 2. 18O-labelled bilirubin appeared in the bile in less than 10 min after exposure of the animals to labelled oxygen. This result suggests that all of the chemical transformations involving production of biliverdin, reduction to bilirubin and conjugation of the bilirubin are fast processes. 3. The quantitative recovery of label obtained in the experiments suggests that there is little or no exchange of newly synthesized bilirubin with existing bilirubin pools in the animal.

| S-EPMC1183896 | biostudies-other

The mechanism of haem catabolism. A study of haem breakdown in spleen microsomal fraction and in a model system by 18O labelling and metal substitution.

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Project description: Not available

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Evaluation of pH during cytostomal endocytosis and vacuolar catabolism of haemoglobin in Plasmodium falciparum.

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A study of the mechanism of quercetin oxygenation by 18O labelling. A comparison of the mechanism with that of haem degradation.

Project description:The mechanism of quercetin oxygenation, which is formally analogous to haem degradation, was studied by using 18O labelling. In both the enzymic oxygenation (catalysed by quercetinase) and the non-enzymic reaction (base-catalysed), both oxygen atoms incorporated into product were derived from a single oxygen molecule. Quercetin oxygenation therefore occurs by a classical dioxygenase mechanism and is not an appropriate model for study of the mechanism of haem catabolism.

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Haemoglobin catabolism: the role of ferrihaems in studies of the degradation pathway.

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Quantitative blood flow measurement in rat brain with multiphase arterial spin labelling magnetic resonance imaging.

Project description:Cerebral blood flow is an important parameter in many diseases and functional studies that can be accurately measured in humans using arterial spin labelling (ASL) MRI. However, although rat models are frequently used for preclinical studies of both human disease and brain function, rat CBF measurements show poor consistency between studies. This lack of reproducibility is due, partly, to the smaller size and differing head geometry of rats compared to humans, as well as the differing analysis methodologies employed and higher field strengths used for preclinical MRI. To address these issues, we have implemented, optimised and validated a multiphase pseudo-continuous ASL technique, which overcomes many of the limitations of rat CBF measurement. Three rat strains (Wistar, Sprague Dawley and Berlin Druckrey IX) were used, and CBF values validated against gold-standard autoradiography measurements. Label positioning was found to be optimal at 45°, while post-label delay was optimised to 0.55 s. Whole brain CBF measures were 109 ± 22, 111 ± 18 and 100 ± 15 mL/100 g/min by multiphase pCASL, and 108 ± 12, 116 ± 14 and 122 ± 16 mL/100 g/min by autoradiography in Wistar, SD and BDIX cohorts, respectively. Tumour model analysis shows that the developed methods also apply in disease states. Thus, optimised multiphase pCASL provides robust, reproducible and non-invasive measurement of CBF in rats.

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Tyrosyl radical in haemoglobin and haptoglobin-haemoglobin complex: how does haptoglobin make haemoglobin less toxic?

Project description:One of the difficulties in creating a blood substitute on the basis of human haemoglobin (Hb) is the toxic nature of Hb when it is outside the safe environment of the red blood cells. The plasma protein haptoglobin (Hp) takes care of the Hb physiologically leaked into the plasma - it binds Hb and makes it much less toxic while retaining the Hb's high oxygen transporting capacity. We used Electron Paramagnetic Resonance (EPR) spectroscopy to show that the protein bound radical induced by H 2O 2 in Hb and Hp-Hb complex is formed on the same tyrosine residue(s), but, in the complex, the radical is found in a more hydrophobic environment and decays slower than in unbound Hb, thus mitigating its oxidative capacity. The data obtained in this study might set new directions in engineering blood substitutes for transfusion that would have the oxygen transporting efficiency typical of Hb, but which would be non-toxic.

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