Peter F. Dodds

Incorporation of xenobiotic carboxylic acids into lipids.

Over thirty-six different xenobiotic carboxylic acids have been reported to form xenobiotic lipids. The majority form triacylglycerol analogs or cholesterol esters with fewer reports of polar lipids being formed. As yet there is insufficient information to deduce a relationship between the structure of the xenobiotic acid and its activity as a substrate for lipid biosynthesis, although the ability to form a CoA ester appears to be important. The action of monoacylglycerol acyltransferase, diacylglycerol acyltransferase, lecithin cholesterol acyltransferase and a carboxylesterase in synthesizing xenobiotic lipids has been demonstrated. One xenobiotic lipid has been shown to be the cause of granulomatous changes and there are some indications that others may prove to be of toxicological or pharmacological significance. Detailed investigations into several aspects of xenobiotic lipid biochemistry are still required.

The incorporation of 3-phenoxybenzoic acid and other xenobiotic acids into xenobiotic lipids by enzymes of the monoacylglycerol pathway in microsomes from adult and neonatal tissues

The incorporation of 3-phenoxybenzoic acid (3PBA) into xenobiotic lipids by enzymes of the monoacylglycerol (MG) pathway was measured using microsomes prepared from rat liver as an enzyme source. The mean activities of the three enzymes involved were: acyl-CoA synthetase, 1.1 nmol/min/mg protein; MG acyltransferase, 75 pmol/min/mg protein; and diacylglycerol acyltransferase, 11.4 pmol/min/mg protein. MG and DG acyltransferase also showed activity with benzoyl-CoA or 1-naphthylacetyl-CoA as acyl donor but none with clofibryl-CoA or 2,4-dichlorophenoxyacetyl-CoA. MG acyltransferase activity, using 3PBA-CoA, was higher in microsomes from rat intestinal mucosa and pig liver, and lower in rat adipose tissue, rat liver and mouse liver. This ranking of activities corresponds to published activities using natural substrates. There was a large increase in MG acyltransferase, using either 3PBA-CoA or palmitoyl-CoA as substrate, in microsomes from the livers of rats 16-18 days old. Lysophosphatidic acid (lyso-PA) and lysophosphatidylethanolamine (lyso-PE), but not other phospholipids or detergents, stimulated MG acyltransferase activity more than two-fold. Lyso-PA (5 microM) increased the Vmax but had little effect on the Km for 2-hexadecylglycerol, whereas 100 microM lyso-PE decreased the Km and had a smaller effect on the Vmax. These results illustrate that the incorporation of xenobiotic acids into diacyl- and triacylglycerol by enzymes of the MG pathway may be a more general phenomenon than was previously suspected and that it may be subject to a variety of developmental and physiological controls.

Xenobiotic triacylglycerol formation in isolated hepatocytes

The formation of neutral lipophilic metabolites from five xenobiotic carboxylic acids was studied in isolated rat hepatocytes. Oleic acid was used as a positive control. Rates of formation of lipids lay in the order: oleic acid greater than phytanic acid greater than ibuprofen greater than 3-phenoxybenzoic acid greater than indomethacin and 3-phenylbutanoic acid (rates were undetectable with the last two substrates). The process was saturable with the maximum rates at about 0.5 mM substrate concentration. Supplementation of the hepatocyte system with glycerol enhanced the yields of lipid products. The hepatocytes also effectively modelled the in vivo metabolism of ibuprofen, 3-phenoxybenzoic acid and 3-phenylbutanoic acid with oxidations and classical conjugation reactions predominating over xenobiotic lipid formation.

Activation of protein kinase C by an aromatic xenobiotic diacylglycerol analogue

Protein kinase C; Xenobiotic lipid; 3‐Phenoxybenzoic acid; Diacylglycerol; Phorbol ester

A Search for Resistance Gene-Specific Receptor Proteins in Lettuce Plasma Membrane

We describe a novel approach towards the detection of receptor proteins in the plasma membrane of lettuce which are responsible for the recognition of avirulent isolates of the downy mildew fungus Bremia lactucae. Heat-shock (HS) causes the gene-specific suppression (for up to 3 days) of the hypersensitive reaction controlled by the resistance gene Dm5/8. This effect is consistent with the loss of a Dm5/8-specific receptor from the plasma membrane (PM) and its slow recovery. In order to detect PM proteins conforming to the criteria defined by HS experiments, PM-enriched fractions were isolated by 2-phase partitioning from heat-shocked and unheated cotyledons and their polypeptide composition successfully analysed by 2D IEF/SDS-PAGE. Two polypeptides of 52.5 kD (putative receptors) were significantly reduced in concentration after HS in cultivars containing Dm5/8 but were unaffected by HS in cultivars which lack this gene. Future analysis of the putative receptors is discussed.