Large agarose beads for extracorporeal detoxification systems. Preparation and enzymatic properties of agarose-bound UDP-glucuronyltransferase.
UDP-glucuronlytransferase, E.C. 2.4.1.17, has been solubilised from the microsomal fraction of liver homogenate from phenobarbital pretreated rabbits by lipase or detergent treatments. A 110-fold purification of the enzyme with respect to the crude homogenate was achieved by precipitation and column separations. The cholate-detergent solubilised enzyme was far more stable than that prepared by the lipase method. The partially purified UDP-glucuronyltransferase has been covalently bound to cyanogen bromide-activated agarose in the form of large haemocompatible beads to the extent of 0.22 mg protein per mg agarose dryweight, equivalent to about 25 mg of swollen gel. The acceptors for glucuronidation employed were the non-physiological phenolic compounds p-nitrophenol and 1-naphthol, and an exogenous and endogenous substance of physiological importance, namely paracetamol and phenol respectively. The immobilised enzyme exhibited at least 80% of the original activity of the solubilised enzyme, and the catalytic function was preserved for a much longer period of time in the carrier-bound form. The system described in this publication could well be applied in an extracorporeal liver assist device for the replacement of glucuronidation function.
D000818:Animals; D001187:Artificial Organs; D004800:Enzymes, Immobilized / Q000378:metabolism*; D005260:Female; D005965:Glucuronates / Q000378:metabolism; D014453:Glucuronosyltransferase / Q000378:metabolism*; D006464:Hemoperfusion; D006501:Hepatic Encephalopathy / Q000378:metabolism; D008099:Liver; D008107:Liver Diseases / Q000378:metabolism; D008862:Microsomes, Liver / Q000201:enzymology; D011134:Polysaccharides; D011506:Proteins / Q000032:analysis; D011817:Rabbits; D012685:Sepharose; D014535:Uridine Diphosphate Glucuronic Acid / Q000378:metabolism
D004800:Enzymes, Immobilized; D005965:Glucuronates; D011134:Polysaccharides; D011506:Proteins; D014535:Uridine Diphosphate Glucuronic Acid; D012685:Sepharose; D014453:Glucuronosyltransferase