ferruginol and salvin

Rosemary extracts containing the phenolic diterpenes carnosic acid and its derivative carnosol are approved food additives used in an increasingly wide range of products to enhance shelf-life, thanks to their high anti-oxidant activity. We describe here the elucidation of the complete biosynthetic pathway of carnosic acid and its reconstitution in yeast cells. Cytochrome P450 oxygenases (CYP76AH22-24) from Rosmarinus officinalis and Salvia fruticosa already characterized as ferruginol synthases are also able to produce 11-hydroxyferruginol. Modelling-based mutagenesis of three amino acids in the related ferruginol synthase (CYP76AH1) from S. miltiorrhiza is sufficient to convert it to a 11-hydroxyferruginol synthase (HFS). The three sequential C20 oxidations for the conversion of 11-hydroxyferruginol to carnosic acid are catalysed by the related CYP76AK6-8. The availability of the genes for the biosynthesis of carnosic acid opens opportunities for the metabolic engineering of phenolic diterpenes, a class of compounds with potent anti-oxidant, anti-inflammatory and anti-tumour activities.

Topics: Abietanes; Anti-Inflammatory Agents; Antineoplastic Agents; Antioxidants; Escherichia coli; Genetic Vectors; Humans; Magnetic Resonance Spectroscopy; Mutagenesis; Mutagenesis, Site-Directed; Mutation; Oxidants; Oxygen; Phylogeny; Rosmarinus; Saccharomyces cerevisiae; Salvia; Spectrometry, Mass, Electrospray Ionization

The antioxidant activity of the abietane-type diterpene ferruginol was evaluated by comparison with that of carnosic acid, ( ± )-α-tocopherol and dibutylhydroxytoluene using 2,2-diphenyl-1-picrylhydrazyl, β-carotene bleaching and linoleic acid assays. Ferruginol had the lowest antioxidant activity of this group using the 2,2-diphenyl-1-picrylhydrazyl and β-carotene methods in polar solvent buffer. However, ferruginol exhibited stronger activity than carnosic acid and α-tocopherol for linoleic acid oxidation under non-solvent conditions. Five peaks corresponding to ferruginol derivatives were detected through GC-MS analysis of the reaction between ferruginol and methyl linoleate. The three reaction products were identified as dehydroferruginol, 7β-hydroxyferruginol and sugiol, and the other two peaks were assumed to be 7α-hydroxyferruginol and the quinone methide derivative of ferruginol. The time course of the reaction suggests that the quinone methide was produced early in the reaction and reacted further to produce dehydroferruginol, 7-hydroxyferruginol and sugiol. Thus, we inferred that quinone methide formation was a key step in the antioxidant reaction of ferruginol.

Topics: Abietanes; Antioxidants; Diterpenes; Molecular Structure; Plant Extracts

Carnosic acid (CA) is the main phenolic diterpene of rosemary (Rosmarinus officinalis L., Lamiaceae) and presents gastroprotective effect in vitro and in vivo. To determine structure-activity relationships, seventeen esters and ethers of CA were prepared, comprising aliphatic, aromatic, and heterocyclic compounds. The naturally occurring 12-O-methylcarnosic acid (14) was also included in the study. The compounds were evaluated for their gastroprotective activity in the HCl/EtOH-induced gastric lesions model in mice, and for cytotoxicity in human adenocarcinoma AGS cells, Hep G2 hepatocellular carcinoma cells, and human lung fibroblasts. At 10 mg/kg, some of the CA derivatives (5, 8, 9, 12, 14, and 18) were more effective preventing gastric lesions than the reference compound lansoprazole at the same dose. The dibenzoate 9, diindoleacetate 12, and the derivative 18 showed the best gastroprotective effect combined with the lowest cytotoxicity.

Topics: 2-Pyridinylmethylsulfinylbenzimidazoles; Abietanes; Animals; Anti-Ulcer Agents; Antioxidants; Cell Line; Disease Models, Animal; Dose-Response Relationship, Drug; Humans; Inhibitory Concentration 50; Lansoprazole; Male; Methylation; Mice; Plant Components, Aerial; Plant Extracts; Random Allocation; Rosmarinus; Stomach Ulcer; Structure-Activity Relationship