Reactions of d-glucose,d-xylose,and d-erythrose with 2-methyl-2-propanethiol

The reaction of d-glucose and d-xylose with 2-methyl-2-propanethiol in conc. hydrochloric acid yielded tert-butyl 1-thioglycopyranosides (products of kinetic and thermodynamic control). Di-tert-butyl dithioacetals (12–14) were obtained from the acetylated aldehydo-derivatives of d-glucose, d-xylose, and d-erythrose. On brief treatment with conc. hydrochloric acid, 12 and 13 gave tert-butyl 1-thio-α- and -β-glycopyranosides and 14 gave the corresponding furanosides.     

1,2-O-Trichloroethylidene acetal group protected 3,5-dieno-1,4-furanose derivatives

The preparation of 3,5-(E)-dieno-3,5,6,8-tetradeoxy-(S)-1,2-O-trichloroethylidene-alpha-D-glycero-octo-1,4-furano-7-ulose starting from either 1,2-O-(S)-trichloroethylidene-alpha-D-glucofuranose (beta-chloralose) or 1,2-O-(S)-trichloroethylidene-alpha-D-galactofuranose (galactochloralose) and the preparation of methyl 3,5-(E)-dieno-3,5,6-trideoxy-(S)-1,2-O-trichloroethylidene-alpha-D-glycero-hepta-1,4-furano-uronate starting from beta-chloralose are described. Endocyclic double bond formations were realised by the elimination of 3-acetoxy groups using DMF-sodium bicarbonate. This elimination was not successful when the starting compound was 1,2-O-(R)-trichloroethylidene-alpha-D-glucofuranose (alpha-chloralose), where the trichloromethyl group occupies the endo position.     

d-Xylopentadialdo-1,4-furanose as a novel bifunctional reagent for enzyme immobilization

The novel bifunctional reagent, d-xylopentadialdo-1,4-furanose, was prepared by the specific oxidation of d-glucose and used for the immobilization of model proteins, such as ovalbumin and subtilisin, onto aminoethylcellulose. The results were compared with that obtained during the immobilization of these proteins to the same support using glutaraldehyde as reagent. As glutaraldehyde and d-xylopentadialdo-1,4-furanose are both dialdehydes, the same pH optima for the binding reaction (near neutrality) were found. The difference between the two reagents was only found in the resulting operational stability. The operational stability was higher in the case of subtilisin bound to aminoethylcellulose using d-xylopentadialdo-1,4-furanose. This is believed to be the result of higher hydrophilicity of the novel bifunctional reagent.     

2,3-Dimethoxy-5-methyl-1,4-benzoquinones and 2-methyl-1,4-naphthoquinones: glycation inhibitors with lipid peroxidation activity

Anti-glycation activity of our anti-oxidant quinone library was measured and several 2,3-dimethoxy-5-methyl-1,4-benzoquinones and 2-methyl-1,4-naphthoquinones were identified as novel inhibitors of glycation, of which 2,3-dimethoxy-5-methyl-1,4-benzoquinones 13b is the most potent glycation inhibitor with around 50 microM of the IC(50) value.     

Electron transfer in DNA duplexes containing 2-methyl-1,4-naphthoquinone

2-Methyl-1,4-naphthoquinone (menadione, MQ) was linked to synthetic oligonucleotides and exposed to near-UV light to generate base radical cations in DNA. This model system of electron transfer induced alkali-labile breaks at GG doublets, similar to anthraquinone and metallointercalators systems. In sharp contrast to other systems, the photolysis of MQ–DNA duplexes gave interstrand cross-links and alkali-labile breaks at bases on the complementary strand opposite the MQ moiety. For sequences with an internal MQ, the formation of cross-links with A and C opposite the MQ moiety was 2- to 3-fold greater than that with G and T. The yield of cross-links was more than 10-fold greater than that of breaks opposite MQ, which in turn was more than 2-fold greater than breaks at GG doublets. The yield of damage at GG doublets greatly increased for a sequence with a terminal MQ. The distribution of base damage was measured by enzymatic digestion and HPLC analysis (dAdo > dThd > dGuo > dCyd). The formation of novel products in MQ–DNA duplexes was attributed to the ability of excited MQ to generate the radical cations of all four DNA bases; thus, this photochemical reaction provides an ideal model system to study the effects of ionizing radiation and one-electron oxidants.     

Reaction of 2'-deoxyribonucleosides with cis- and trans-1,4-dioxo-2-butene

cis-1,4-Dioxo-2-butene is a toxic metabolite of furan, while the trans-isomer is a product of deoxyribose oxidation in DNA. It has recently been reported that both cis- and trans-1,4-dioxo-2-butene react with the 2'-deoxynucleosides dC, dG, and dA to form novel diastereomeric oxadiazabicyclo(3.3.0)octaimine adducts. We have now extended these studies with kinetic and spectroscopic analyses of the reactions of cis- and trans-1,4-dioxo-2-butene, as well as the identification of novel adducts of dA. The reaction of dC with trans-1,4-dioxo-2-butene was observed to be nearly quantitative and produced two interchanging diastereomers with a second-order rate constant of 3.66 x 10(-2)M(-1)s(-1), which is nearly 10-fold faster than the reaction with the cis-isomer (3.74 x 10(-3)M(-1)s(-1)). HPLC analyses of reactions of 1,4-dioxo-2-butene with both dA and 9-methyladenine (pH 7.4, 37 degrees C) revealed multiple products including a novel pair of closely eluting fluorescent species of identical mass ([M+H] m/z 420), each of which contains two molecules of 1,4-dioxo-2-butene, and a more abundant but unstable and non-fluorescent species ([M+H] m/z 414). Partial structural characterization of the fluorescent adducts of dA revealed the presence of the oxadiazabicyclo(3.3.0)octaimine ring system common to the dC adducts. These results support the genotoxic potential of furan metabolites and products of deoxyribose oxidation.     

Structure of 2-methyl-3-VIII-dihydrooctaprenyl-1,4-naphthoquinone from Halococcus morrhuae

Abstract The position of saturation of the dihydrogenated menaquinone-8 from Halococcus morrhuae has been investigated by mass spectrometry (MS) and proton nuclear magnetic resonance spectrometry (¹H-NMR). The results of the present study demonstrate that the terminal isoprene (8th from the ring) is saturated, and the quinone corresponds to 2-methyl-3-VIII-dihydrooctaprenyl-1,4-naphthoquinone.     

Interaction of menadione (2-methyl-1,4-naphthoquinone) with glutathione

The interaction of menadione with reduced glutathione (GSH) led to a removal of menadione and formation of menadione-GSH conjugate and glutathione disulfide (GSSG). The changes in thiol level were essentially biphasic with an initial rapid decrease in GSH and appearance of GSSG (less than 1 min) followed by secondary less pronounced changes. The interaction of menadione and GSH caused an oxygen uptake and both superoxide anion radical and hydrogen peroxide were produced during the reaction, the amount dependent on the GSH/menadione ratio. Catalase did not protect against the initial decrease in GSH level but markedly inhibited the secondary changes while superoxide dismutase had little effect. These results suggest that the initial changes in thiol level are the result in part of a redox reaction between menadione and GSH as well as conjugate formation, whilst the secondary changes reflect conjugate formation and the activity of other oxidants such as hydrogen peroxide. The potential biological significance of this reaction was investigated using hepatocytes depleted of reduced pyridine nucleotides and thus not able to perform enzyme-catalyzed reduction of menadione. In these cells menadione induced GSSG formation at a rate similar to that observed in control cells. This suggests that quinone-induced oxidative challenge caused by the chemical interactions of a quinone and glutathione may have biological relevance.     

Major adenine products from 2-methyl-1,4-naphthoquinone-sensitized photoirradiation at 365 nm

In this article we report the isolation and characterization of major products of adenine in dinucleoside monophosphates upon 2-methyl-1,4-naphthoquinone (menadione)-sensitized UVA irradiation. Our results show that the major products form via the coupling between the menadione moiety and the exocyclic amino group of adenine. Similar reactions were not observed for cytosine. To our knowledge, this is the first report about the direct reaction between a DNA base and a photosensitizer under 365-nm ultraviolet light irradiation. Our results are consistent with previous observation showing that N(6) radical formed on adenine upon UVA irradiation.     

Effect of inducers of DT-diaphorase on the toxicity of 2-methyl- and 2-hydroxy-1,4-naphthoquinone to rats

It has previously been shown that rats pre-treated with butylated hydroxyanisole (BHA), a well-known inducer of the enzyme DT-diaphorase, are protected against the toxic effects of 2-methyl-1,4-naphthoquinone but are made more susceptible to the harmful action of 2-hydroxy-1,4-naphthoquinone. In the present experiments, the effects of BHA have been compared with those of other inducers of DT-diaphorase. Rats were dosed with BHA, butylated hydroxytoluene (BHT), ethoxyquin (EQ), dimethyl fumarate (DMF) or disulfiram (DIS) and then challenged with a toxic dose of the naphthoquinones. All the inducers protected against the haemolytic anaemia induced by 2-methyl-1,4-naphthoquinone in rats, with BHA, BHT and EQ being somewhat more effective than DMF and DIS. A similar order of activity was recorded in the relative ability of these substances to increase hepatic activities of DT-diaphorase, consistent with a role for this enzyme in facilitating conjugation and excretion of this naphthoquinone. In contrast, all the compounds increased the haemolytic activity of 2-hydroxy-1,4-naphthoquinone. DMF and DIS were significantly more effective in this regard than BHA, BHT and EQ. DMF and DIS also caused a much greater increase in levels of DT-diaphorase in the intestine, suggesting that 2-hydroxy-1,4-naphthoquinone is activated by this enzyme in the gut. BHA, BHT and EQ had no effect on the nephrotoxicity of 2-hydroxy-1,4-naphthoquinone, but the severity of the renal lesions was decreased in rats pre-treated with DMF and DIS. The results of the present experiments show that modulation of tissue levels of DT-diaphorase may not only alter the severity of naphthoquinone toxicity in vivo, but may also change the relative toxicity of these substances to different target organs.     

Reactivity of thiols towards derivatives of 2- and 6-methyl-1,4-naphthoquinone bioreductive alkylating agents

The stoichiometry and kinetics of the anaerobic reactions between some thiols and derivatives of 2- and 6-methyl-1,4-naphthoquinones in water were measured using stopped flow spectrophotometry. The stoichiometry of the reaction with representative compounds was 1:2 thiol:quinone, a finding consistent with the formation of a hydroquinone as well as a thioether in the reaction. The first-order dependence of rate on thiol concentration, and the pH-dependent rate constants indicated that the thiolate anion was involved in the rate-limiting step, with rate constants at pH 7.6 generally increasing in the order glutathione (GSH) less than cysteamine less than dithiothreitol (DTT) less than cysteine. Despite the lower reactivity of GSH, the half-lives of the uncatalyzed conjugation reaction of these quinones at typical biological concentrations of GSH (e.g. 2 mM) ranged from about 2.0 to 20 s at pH 7.6 and 25 degrees C. The implications of these reactions in the use of naphthoquinones as potential bioreductive alkylating agents and as hypoxic cell radiosensitizers are discussed.     

Sensitized photo-oxidation of thymidine by 2-methyl-1,4-naphthoquinone. Characterization of the stable photoproducts

The near ultraviolet photolysis of an aerated aqueous solution of thymidine containing 2-methyl-1,4-naphthoquinone gives rise to two main classes of photoproducts as a result of the initial formation of a pyrimidine radical cation. These photo-oxidation products have been separated by high performance liquid chromatography and further characterized by various spectroscopic techniques including fast atom bombardment mass spectrometry and high field 1H and 13C nuclear magnetic resonance analysis. This photoreaction constitutes an excellent model to study the chemical properties of the thymidine radical cation which is expected to be one of the primary consequences of the direct effects of ionizing radiation.     

Menadione- (2-methyl-1,4-naphthoquinone-) dependent enzymatic redox cycling and calcium release by mitochondria

The results presented in this paper reveal the existence of three distinct menadione (2-methyl-1,4-naphthoquinone) reductases in mitochondria: NAD(P)H:(quinone-acceptor) oxidoreductase (D,T-diaphorase), NADPH:(quinone-acceptor) oxidoreductase, and NADH:(quinone-acceptor) oxidoreductase. All three enzymes reduce menadione in a two-electron step directly to the hydroquinone form. NADH-ubiquinone oxidoreductase (NADH dehydrogenase) and NAD(P)H azoreductase do not participate significantly in menadione reduction. In mitochondrial extracts, the menadione-induced NAD(P)H oxidation occurs beyond stoichiometric reduction of the quinone and is accompanied by O2 consumption. Benzoquinone is reduced more rapidly than menadione but does not undergo redox cycling. In intact mitochondria, menadione triggers oxidation of intramitochondrial pyridine nucleotides, cyanide-insensitive O2 consumption, and a transient decrease of delta psi. In the presence of intramitochondrial Ca2+, the menadione-induced oxidation of pyridine nucleotides is accompanied by their hydrolysis, and Ca2+ is released from mitochondria. The menadione-induced Ca2+ release leaves mitochondria intact, provided excessive Ca2+ cycling is prevented. In both selenium-deficient and selenium-adequate mitochondria, menadione is equally effective in inducing oxidation of pyridine nucleotides and Ca2+ release. Thus, menadione-induced Ca2+ release is mediated predominantly by enzymatic two-electron reduction of menadione, and not by H2O2 generated by menadione-dependent redox cycling. Our findings argue against D,T-diaphorase being a control device that prevents quinone-dependent oxygen toxicity in mitochondria.     

Biosynthesis of tocopherols: A re-examination of the biosynthesis and metabolism of 2-methyl-6-phytyl-1,4-benzoquinol

A number of previous studies of the involvement of 2-methyl-6-phytyl-1,4-benzoquinol in the biosynthesis of α-tocopherol have failed to take account of the fact that this quinol and its quinone have very similar chromatographic properties to those of 2-methyl-3-phytyl-1,4-benzoquinol and 2-methyl-3-phytyl-1,4-benzoquinone respectively. It has now been shown that the two quinones can be separated from each other either by multidevelopment TLC or by HPLC and that the claims made earlier with regard to the biosynthesis and metabolism of 2-methyl-6-phytyl-1,4-benzoquinol in chloroplasts are correct. In particular, it has been established that this quinol is the only methyl phytylbenzoquinol formed from homogentisate and phytyl pyrophosphate in chloroplast preparations. It has also been shown for the first time that lettuce chloroplasts are able to synthesize ³H-labelled α- and γ-tocopherols from [methylene-³H] homogentisate.     

Novel epoxide formation in the reaction of 2-bromo-3-methyl-1,4-naphthoquinone with 1,3-propanedithiol

A novel epoxide 2 was formed as the major product in the reaction of 2-bromo-3-methyl-1,4-naphthoquinone with 1,3-propanedithiol in the presence of triethylamine in 92% yield. Molecular oxygen is suggested to be the source of the added oxygen in 2, an oxidation product of its precursor 3. A strong base such as triethylamine is required to abstract the methyl hydrogen of 1,4-naphthoquinones, leading to the formation of 3 as well as 2.