Witold Danikiewicz

Contribution of the mevalonate and methylerythritol phosphate pathways to the biosynthesis of dolichols in plants.

Plant isoprenoids are derived from two biosynthetic pathways, the cytoplasmic mevalonate (MVA) and the plastidial methylerythritol phosphate (MEP) pathway. In this study their respective contributions toward formation of dolichols in Coluria geoides hairy root culture were estimated using in vivo labeling with 13C-labeled glucose as a general precursor. NMR and mass spectrometry showed that both the MVA and MEP pathways were the sources of isopentenyl diphosphate incorporated into polyisoprenoid chains. The involvement of the MEP pathway was found to be substantial at the initiation stage of dolichol chain synthesis, but it was virtually nil at the terminal steps; statistically, 6–8 isoprene units within the dolichol molecule (i.e. 40–50% of the total) were derived from the MEP pathway. These results were further verified by incorporation of [5-2H]mevalonate or [5,5-2H2]deoxyxylulose into dolichols as well as by the observed decreased accumulation of dolichols upon treatment with mevinolin or fosmidomycin, selective inhibitors of either pathway. The presented data indicate that the synthesis of dolichols in C. geoides roots involves a continuous exchange of intermediates between the MVA and MEP pathways. According to our model, oligoprenyl diphosphate chains of a length not exceeding 13 isoprene units are synthesized in plastids from isopentenyl diphosphate derived from both the MEP and MVA pathways, and then are completed in the cytoplasm with several units derived solely from the MVA pathway. This study also illustrates an innovative application of mass spectrometry for qualitative and quantitative evaluation of the contribution of individual metabolic pathways to the biosynthesis of natural products.

Role of polyisoprenoids in tobacco resistance against biotic stresses

Infection with avirulent pathogens, tobacco mosaic virus (TMV) or Pseudomonas syringae pv. tabaci induced accumulation of polyisoprenoid alcohols, solanesol and a family of polyprenols [from polyprenol composed of 14 isoprene units (Pren-14) to -18, with Pren-16 dominating] in the leaves of resistant tobacco plants Nicotiana tabacum cv. Samsun NN. Upon TMV infection, solanesol content was increased seven- and eight-fold in the inoculated and upper leaves, respectively, while polyprenol content was increased 2.5- and 2-fold in the inoculated and upper leaves, respectively, on the seventh day post-infection. Accumulation of polyisoprenoid alcohols was also stimulated by exogenously applied hydrogen peroxide but not by exogenous salicylic acid (SA). On the contrary, neither inoculation of the leaves of susceptible tobacco plants nor wounding of tobacco leaves caused an increase in polyisoprenoid content. Taken together, these results indicate that polyisoprenoid alcohols might be involved in plant resistance against pathogens. A putative role of accumulated polyisoprenoids in plant response to pathogen attack is discussed. Similarly, the content of plastoquinone (PQ) was increased two-fold in TMV-inoculated and upper leaves of resistant plants. Accumulation of PQ was also stimulated by hydrogen peroxide, bacteria (P. syringae) and SA. The role of PQ in antioxidant defense in cellular membranous compartments is discussed in the context of the enzymatic antioxidant machinery activated in tobacco leaves subjected to viral infection. Elevated activity of several antioxidant enzymes (ascorbate peroxidase, guaiacol peroxidase, glutathione reductase and superoxide dismutase, especially the CuZn superoxide dismutase isoform) and high, but transient elevation of catalase was found in inoculated leaves of resistant tobacco plants but not in susceptible plants.

SELECTIVE DEHYDRATION OF GLUCOSE TO HYDROXYMETHYLFURFURAL AND A ONE-POT SYNTHESIS OF A 4-ACETYLBUTYROLACTONE FROM GLUCOSE AND TRIOXANE IN SOLUTIONS OF ALUMINIUM SALTS

Abstract Saturated water solutions of Al 2 (SO 4 ) 3 and AlCl 3 were applied as solvent/matrices for dehydration of Glc to hydroxymethylfurfural (HMF). Addition of oxygen ligands: methanol, ethanol, THF, furan, dibutyl ether, ethyl orthoformate and trioxane influenced the yield and selectivity, the best being observed with ethanol. When Glc and trioxane were present together in reacting solution, formation of a 4-acetylbutyrolactone was observed.

Divergent pattern of Polyisoprenoid alcohols in the tissues of coluria geoides: A new electrospray lonization ms approach

Polyisoprenoid alcohols of the plant Coluria geoides were isolated and analyzed by HPLC with UV detection to determine the nature of the polyprenol and dolichol mixture in the organs studied. In roots, a family of dolichols (Dol-15 to Dol-23, with Dol-16 dominating, where Dol-n is dolichol composed of n isoprene units) was accompanied by traces of polyprenols of similar chain lengths, whereas in hairy roots grown in vitro, identical patterns with a slightly broader chain-length range were found. Conversely, in leaves and seeds polyprenols were the dominant form, and their pattern was shifted toward longer chains (maximal content of Pren-19, where Pren-n is polyprenol composed of n isoprene units). Interestingly, the pattern of dolichols in seeds and leaves (in which Dol-17 dominated) was similar to that found in roots.Structures of the dolichols and polyprenols isolated were confirmed by the application of a new HPLC/electrospray ionization-MS method, which also offers a much higher sensitivity in detection of these compounds compared to a UV detector. The highest sensitivity was obtained when the [M+Na]+ ions of polyprenols and dolichols were recorded in the selected ion monitoring mode and a small amount of sodium acetate solution was added post-column to enhance the formation of these ions in an electrospray ion source.

Concentrated water solutions of salts as solvents for reaction of carbohydrates. Part 2. Influence of some magnesium salts and some ruthenium species on catalysis of dehydration of glucose

Saturated water solutions of MgCl2 and MgSO4 were applied as solvents/matrices for dehydration of glucose. The product of reaction between RuCl3(aq.) and Ag2SO4 abbreviated as ‘RuSO4’ was used as catalyst. As the most selective systems are: MgSO4/benzene(ethanol)/RuSO4 the mechanistic hypothesis was designed for these cases. Interaction of glucose with MgSO4RuSO4 consists of: Mg2+ complexation of glucose oxygens; hydrogen bonds between the glucose OH groups and the SO2−4 groups; between the glucose OH groups and water, hydroxy ligands in the Ru complex and SO2−4 donation to the glucose carbons.

Mass spectrometry studies on meso‐substituted corroles and their photochemical decomposition products

Corroles, ring-contracted analogs of porphyrins, are an important class of compounds which have attracted the attention of many researchers in the fields of organic, coordination and physical chemistry. In the present work, the stability and the decomposition pathways of a diverse set of meso-substituted corroles have been studied using mass spectrometry (MS), UV-Vis absorption and preparative methods combined with NMR spectroscopy. Four different ionization methods (electrospray ionization, field desorption, atmospheric pressure photoionization and atmospheric pressure chemical ionization) were utilized to investigate light- and oxygen-induced decomposition in various solvents. It was found that the rate of decomposition in MeCN is significantly higher than in CH(2)Cl(2), hexane, MeOH and ethyl acetate. HR-MS combined with CID-MS/MS enabled us to identify the products of initial decomposition. Surprisingly, numerous smaller open-chain compounds were also detected. Large-scale decomposition of a corrole bearing sterically hindered substituents at positions 5 and 15 allowed us to isolate mg quantities of three decomposition products: isocorrole and isomeric biliverdin-type species. These are formed as a result of oxygen attack on the meso-10 position.

Ruthenium-amido complexes: synthesis, structure, and catalytic activity in olefin metathesis.

Olefin metathesis has become a powerful reaction for the formation of carbon–carbon bonds in organic and polymer chemistry. Availability of well-defined ruthenium-based catalysts (e.g., 1–3) tolerant of moisture, oxygen, various functional groups and normal organic or polymer processing conditions has greatly expanded the scope and applications of this process and has made it widely used in synthesis.

Direct nitromethylation of nitronaphthalene and its heteroanalogues. A new method for functionalization of nitroarenes

Abstract Some nitroarenes react with nitromethane anion to form nitromethyl derivatives which are easily converted into corresponding nitroaromatic aldehydes.

Structural elucidation of specific noncovalent association of folic acid with native cyclodextrins using an ion mobility mass spectrometry and theoretical approach.

The combination of ion mobility mass spectrometry studies and theoretical calculations including docking studies permitted a detailed structural description of noncovalent complexes of folic acid (FA) and native cyclodextrins (α-CD, β-CD, and γ-CD). The mode of noncovalent association depended on the cavity size of the cyclodextrin. The structure of FA/α-CD represented the exclusion complex in which the aminobenzoic moiety and the aromatic pteridine ring of folic acid remain outside the cyclodextrin cavity, while the glutamate residue is anchored in the interior of the α-cyclodextrin. A rotaxane-type structure was proposed for the FA/β-CD complex with the aminobenzoic part of FA being trapped in the central cavity of β-CD. The glutamate residue and the aromatic pteridine ring interact with the primary and secondary rim hydroxyl residues, respectively, enhancing complex stability. Two possible structures of FA/γ-CD were suggested, the first one being analogous to the FA/β-CD complex and the second one being more stable-in which the aromatic pteridine ring penetrates into the CD cavity while the glutamate residue with the aminobenzoic part of FA is exposed to the cone exterior of CD at its wider edge. Further insight into the association behavior of the folic acid toward cyclodextrins evaluated by thermodynamic calculations indicates that the process is highly exothermic. The complex stability increased in the order FA/α-CD < FA/β-CD < FA/γ-CD. This order is consistent with the previously determined relative gas-phase stability established based on the dissociation efficiency curves of the FA/CD complexes.

The effects of statins on the mevalonic acid pathway in recombinant yeast strains expressing human HMG-CoA reductase.

BackgroundThe yeast Saccharomyces cerevisiae can be a useful model for studying cellular mechanisms related to sterol synthesis in humans due to the high similarity of the mevalonate pathway between these organisms. This metabolic pathway plays a key role in multiple cellular processes by synthesizing sterol and nonsterol isoprenoids. Statins are well-known inhibitors of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR), the key enzyme of the cholesterol synthesis pathway. However, the effects of statins extend beyond their cholesterol-lowering action, since inhibition of HMGR decreases the synthesis of all products downstream in the mevalonate pathway. Using transgenic yeast expressing human HMGR or either yeast HMGR isoenzyme we studied the effects of simvastatin, atorvastatin, fluvastatin and rosuvastatin on the cell metabolism.ResultsStatins decreased sterol pools, prominently reducing sterol precursors content while only moderately lowering ergosterol level. Expression of genes encoding enzymes involved in sterol biosynthesis was induced, while genes from nonsterol isoprenoid pathways, such as coenzyme Q and dolichol biosynthesis or protein prenylation, were diversely affected by statin treatment. Statins increased the level of human HMGR protein substantially and only slightly affected the levels of Rer2 and Coq3 proteins involved in non-sterol isoprenoid biosynthesis.ConclusionStatins influence the sterol pool, gene expression and protein levels of enzymes from the sterol and nonsterol isoprenoid biosynthesis branches and this effect depends on the type of statin administered. Our model system is a cheap and convenient tool for characterizing individual statins or screening for novel ones, and could also be helpful in individualized selection of the most efficient HMGR inhibitors leading to the best response and minimizing serious side effects.