J. Phys. IV France 125 (2005) 717-720
Determination of bilirubin by thermal lens spectrometry and studies of its transport into hepatic cellsA. Margon1, M. Terdoslavich2, A. Cocolo2, G. Decorti3, S. Passamonti2 and M. Franko1
1 Nova Gorica Polytechnic, School of Environmental Sciences, PO Box 301, 5001 Nova Gorica, Slovenia
2 University of Trieste, Department of Biochemistry, Biophysics and Macromolecular Chemistry, Via Giorgeri 1, 34127 Trieste, Italy
3 University of Trieste, Department of Biomedical Sciences, Via Giorgeri 7, 34127 Trieste, Italy
The liver is responsible for clearance of bilirubin, the end product of heme catabolism, from the bloodstream. The main aim of our investigation was to determine the role of the carrier protein bilitranslocase in bilirubin uptake into the liver. Our experiments consisted of exposing cell cultures to bilirubin solutions under different conditions and measuring the uptake of bilirubin into the cells. However, since bilirubin is only slightly soluble in aqueous solution (< 70 nM at pH 7.4), we had to use bilirubin concentrations that are far below the limit of detection of the commonly used techniques (e.g. LOD for HPLC with UV-Vis detection M). TLS showed up to be a suitable technique for investigation of bilirubin uptake with an LOD of 2 nM. Under basal conditions, bilirubin uptake did not occur. However, increase of cytosolic NADH due to catabolism of specific substrates (e.g. lactate or ethanol) seemed to trigger bilirubin uptake. Furthermore, bilirubin uptake was completely inhibited by addition of specific anti-bilitranslocase antibodies. We can thus infer that, under these conditions, bilitranslocase is the main bilirubin transporter.
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