Mixture Of Adducts Research Articles

Preparation of diastereomerically pure 9-carboxybicyclo[6.1.0]nonane derivatives

Reactions of cyclooctene (1) with dibromocyanoacetic esters and copper(I) bromide give (8–10) : 1 mixtures of isomers (2, 3), not stereochemically pure compounds as reported by others. The stereochemistry is elucidated by independent synthesis of one diastereomer (3a). Carbenoid addition of alkoxycarbonylmethylene to 1 and 1,5-cyclooctadiene leads also to exo/endo adduct mixtures. Methods are developed to generate diastereomerically pure compounds (exo-7a, exo-8, endo-9, exo-10, endo-10) from these. Endo esters of this series undergo very facile base catalyzed epimerization.

Synthesis of Chiral alpha,delta-Dioxygenated Allylic Stannanes as Reagents for Carbohydrate Synthesis and Homologation.

The delta-oxygenated allylic stannanes 4.4 and 4.5, prepared through addition of Bu(3)SnLi to gamma-OTBS crotonaldehyde 4.3c followed by etherification of the adduct with TBS-Cl or MOM-Cl, undergo transmetalation with InCl(3) and in situ addition to aldehydes leading to mainly anti adducts 5.1 or 5.2, accompanied by varying amounts of syn diastereomers. Selectivities of >95:5 can be realized with the MOM reagent 4.5 and ynals 4.3d and 4.3e or cyclohexanecarboxaldehyde 4.3a. With enals 4.3b and 4.3c, 80:20 mixtures of anti and syn adducts are formed. The S enantiomer 10.1 of stannane 4.5 has also been prepared as a reagent for carbohydrate synthesis. Accordingly, addition to alpha-ODPS acetaldehyde 10.2 in the presence of InCl(3) leads to the adduct 10.3 as an inseparable 90:10 mixture of anti and syn diastereomers. Dihydroxylation of the OTBS derivative 10.4 affords the potential altrose precursor 10.5 in 81% yield.

Diastereoselective reactions of enolates

Abstract

First Michael Addition Reaction of α-Substituted N-Diphenylmethyleneglycinate with Ethyl Propiolate. Synthesis of α-Substituted (E)-3,4-Dehydroglutamic Acids

(E)-α-substituted 3,4-dehydro glutamic acids 4, were prepared from ethyl N-diphenylmethyleneglycinate 5 by alkylation with a suitable alkyl halide followed by a Michael type addition reaction with ethyl propiolate, giving rise to a mixture of EZ adducts 78. Sequential hydrolysis of the substituted glycine synthons 78 afforded the titled compounds 4 as single diastereomers.

Reaction between Metabolically Activated Acetaminophen and Phosphorothioate Oligonucleotides

Assessment of toxic or mutagenic risks associated with phosphorothioate oligonucleotides (PTO) is important. In vitro and in vivo data have shown that PTOs are nontoxic and nonmutagenic. However, these studies do not address interactions between PTOs and other compounds. The sulfur on PTOs may provide a novel reactive center on a DNA molecule for drug interactions. This study chose acetaminophen (ACAP) as a model drug because ACAP is oxidized to the reactive intermediate N-acetyl-p-benzoquinone imine (NAPQI), which reacts with sulfur-containing compounds. Reaction of dCTP(S) with NAPQI or activated ACAP formed a product distinct from the reactants. Analysis of the product by fast atom bombardment mass spectroscopy gave a molecular weight consistent with NAPQI bound to a sulfur. Higher-molecular-weight products were seen on a polyacrylamide gel electrophoresis after incubation of fluorescein-labeled PTO with NAPQI. These products were not seen after incubation of a phosphodiester oligonucleotide with NAPQI. 31P NMR analysis confirmed the existence of a heterogenous mixture of adducts between a PTO and NAPQI. Nonsequence-specific PTOs of various lengths were tested for their ability to reduce ACAP toxicity. Cell viability showed that larger PTOs provided greater protection. We evaluated the ability of NAPQI to cause mutations in the LacZ gene of pBluescript plasmid which contained phosphorothioate linkages at designed locations within the gene. In addition, the ability of ACAP to cause mutations in the HGPRT locus in cells grown in dATP(S)-containing medium was measured. No mutations were seen in either assay. Based upon these results, activated ACAP is reactive with PTOs in vitro, although this interaction is nontoxic and nonmutagenic.

Synthesis of vinyl sulfide analogs of 2,3-oxidosqualene and their inhibition of 2,3-oxidosqualene lanosterol-cyclases

Syntheses of all trans (6 E)-5-, (10 E)-9-, (14 E)-16- and (18 E)-20-thia-2,3-oxidosqualenes as inhibitors of 2,3-oxidosqualene-lanosterol cyclase (OSC) are reported. To mimic the natural geometry of 2,3-oxidosqualene (2,3-OS), we required E-vinyl sulfides which were prepared by condensation of sulfur-substituted Wittig-Horner reagents (α-thioterpenoidyl diphenylphosphine oxides) with appropriate aldehydes. Mixtures of syn and anti α-hydroxydiphenylphosphine adducts were separated by chromatography and the syn isomers were transformed to the E-vinyl sulfides. Both (6 E)-5- and (18 E)-20-thia-2,3-OS inhibited OSC from Candida albicans (IC 50 = 47 and 0.2 μM, respectively) and rat liver (IC 50 = 7.7 and 0.32 μM, respectively). Their activities were compared with those of previously synthesized (6 E)-8- and (14 E)-13-thia-2,3-OSs (IC 50 = 0.68 and 45 μM, C. albicans, IC 50 = 34 and 61 μM, rat liver, respectively). The best inhibitor among these compounds for the OSC of C. albicans and rat liver is the (18 E)-20-thia-2,3-OS. This result suggests that modification of C-20 region of the 2,3-OS skeleton is an attractive strategy for development of OSC inhibitors.

Hetero‐Diels‐Alder Additions of α,β‐Unsaturated‐Acyl Cyanides. Part 3. Syntheses of 3‐bromo‐2‐ethoxy‐3,4‐dihydro‐2H‐pyran‐6‐carbonitriles, and about their transformation to 2‐ethoxy‐2H‐pyrans

AbstractCycloadditions of the α,β‐unsaturated‐acyl cyanides 1–3 with (Z)‐or (E)‐1‐bromo‐2‐ethoxyethene (4) may be performed at moderate temperatures and provide in good yields the 3‐bromo‐2‐ethoxy‐3,4‐dihydro‐2H‐pyran‐6‐carbonitriles 5–7, respectively (Scheme 1). Diastereoisomeric pairs of products result at room temperature merely from the ‘endo’‐ and ‘exo’‐transition states; more complex mixtures appear above 60° as a consequence of (Z)/(E)‐isomerization of 4. The relative stability of the anomers of 5 and 6 is explored by treatment with BF3·Et2O. Acid alcoholysis (MeOH or EtOH) of 5 leads to acetals 9a, b of 4‐bromo‐5‐oxopentanoate. Alkyl (2Z,4E)‐5‐ethoxypenta‐2,4‐dienoates 12, 17, and 20, are formed in alcoholic alkoxide solutions from 5, 6, and 7, respectively, which is compatible with the intermediacy of 2‐alkoxy‐2H‐pyrans and their valence tautomers, α,β‐unsaturatedacyl cyanides. Methoxide addition to the CN group competes with dehydrobromination in case of 5; it leads to 3‐bromo‐3,4‐dihydro‐2H‐pyran‐6‐carboximidate 13 (ca. 50% at −20°) which can be hydrolyzed to the methyl carboxylate 14. DBU (1,8‐diazabicyclo[5,4,0]undec‐7‐ene) in benzene converts 5 to 6‐ethoxy‐2‐oxohexa‐3,5‐dienenitrile (11), the ring‐opening product of an obviously unstable 2‐ethoxy‐2H‐pyran; the same reagent dehydrobrominates 6 to 2‐ethoxy‐4‐methyl‐2H‐pyran‐6‐carbonitrile (15). HBr Elimination from 7 takes place with great ease in presence of pyridine, or even during chromatography on alumina, and leads to the stable ethyl 6‐cyano‐2‐ethoxy‐2H‐pyran‐4‐carboxylate (18); this dimerizes at room temperature to give a 1:3 mixture of tricyclic adducts ‘endo’‐21 and ‘exo’‐21. The structure of the latter is established by an X‐ray crystallographic analysis.

Tetrakis (μ-difluoroacetato) dimolybdenum(II): solution chemistry and mass spectrum

The complex Mo 2(O 2CCHF 2) 4 has been prepared and characterized, and its solution chemistry has been studied. In acetone, it exists as the axial adduct; in pyridine, it exists as a mixture of axial and monoequatorial adducts which undergo exchange. It forms axial, mono- and diequatorial adducts with tributylphosphine in acetone; these adducts also exchange. This complex undergoes less carboxylate ring-opening with pyridine or tributylphosphine than does Mo 2(O 2CCF 2) 4 under identical conditions. In the mass spectrometer, Mo 2(O 2CCHF 2) 4 fragments via early losses of CHFCO 2 and F, processes analogous to those undergone by Mo 2(O 2CCF 3) 4.

Phosphorus hydrazides as building blocks for potential photoaffinity labels. Synthesis and co-ordination chemistry of perfluoroaryl azide conjugates of phenylphosphonothioic dihydrazide

The phosphorus hydrazide PhP(S)(NMeNH2)21 reacted with the azido functionalized aldehyde 4-N3C6F4CHO to give a mixture of Schiff-base adducts with one or two perfluoroaryl azido substituents on 1. However, the reaction with R′CHO (R = 4-O2NC6H4, 3-O2NC6H4 or 2,4-Me2C6H3) resulted in a considerable selectivity of incorporating one aromatic substituent in the terminal hydrazide unit of 1via Schiff-base coupling to produce PhP(S)(NMeNH2)(NMeNCHR′)(R′= 4-O2NC6H43, 3-O2NC6H44 or 2,4-Me2C6H35). These mono-Schiff-base adducts 3–5 undergo further Schiff-base coupling with azidotetrafluorobenzaldehyde to produce the azido-functionalized phosphorus hydrazides PhP(S)-(NMeNCHC6F4N3)(NMeNCHR′)(R′= 4-O2NC6H47, 3-O2NC6H48 or 2,4-Me2C6H39). The ligating properties of the representative phosphorus hydrazides 2 and 9 with palladium (II) were investigated. The crystal structure of the complex [PdCl2{PhP(S)(NMeNH2)(NMeNCHC6F4N3-4)}] reveals that the palladium(II) is bound in a chelating cis arrangement to the phosphorus chalcogenide and the hydrazine nitrogen via a five-membered metallacyclic framework: monoclinic, space group P21/n, a= 7.826(10), b= 18.051 (20), c= 18.330(3)A, β= 98.196(7)°, Z= 4, R= 0.038 and R′= 0.053.

Reaction of C60 with cyclopent-2-enone acetals. A convenient access to chiral C60 derivatives

C60 reacts with cyclopent-2-enone ethylene acetal to give C60-fused norbornan-2-one acetal; modification of the starting acetal with a chiral auxiliary leads to a mixture of diastereoisomeric adducts from which a pair of enantiomerically pure C60-fused norbornan-2-ones are readily obtained.

The development of controllable metal-chelate curing agents with improved storage stability

The novel latent epoxy curing agent and commercial epoxy system designated MY750/Cu(PGE-EMI)4Cl2 shows remarkable properties under both extended storage and cure conditions. For example, after a period of 2600 hours at ambient temperature the viscosity of an MY750/Cu(PGE-EMI)4Cl2 mixture is still low enough (ca.3x106 cps) to be measured at ambient temperature, while the corresponding MY750/(PGE-EMI) adduct mixture is too viscous to be measured (ca. 8x106 cps) after only 430 hours. The slower increase in viscosity of the complex mixture indicates that it is much more stable than its parent imidazole when dispersed in an epoxy prepolymer over an extended period of time. FTIR and 1H n.m.r. data suggest that after heating an MY750/Cu(PGE-EMI)4Cl2 mixture for 5 minutes at high temperatures (120–140°C) and a room temperature quench, almost no further cure occurs thereafter.

Separation of 32P-Postlabeled DNA Adducts of Polycyclic Aromatic Hydrocarbons and Nitrated Polycyclic Aromatic Hydrocarbons by HPLC

The 32P-postlabeling assay, thin-layer chromatography, and reverse-phase high-pressure liquid chromatography (HPLC) were used to separate DNA adducts formed from 10 polycyclic aromatic hydrocarbons (PAHs) and 6 nitrated polycyclic aromatic hydrocarbons (NO2-PAHs). The PAHs included benzo[j]fluoranthene, benzo[k]fluoranthene, indeno[1,2,3-cd]pyrene, benzo[a]pyrene, chrysene, 6-methylchrysene, 5-methylchrysene, and benz[a]anthracene. The NO2-PAHs included 1-nitropyrene, 2-nitrofluoranthene, 3-nitrofluoranthene, 1,6-dinitropyrene, 1,3-dinitropyrene, and 1,8-dinitropyrene. Separation of seven of the major PAH-DNA adducts was achieved by an initial PAH HPLC gradient system. The major NO2-PAH-DNA adducts were not all separated from each other using the initial PAH HPLC gradient but were clearly separated from the PAH-DNA adducts. A second NO2-PAH HPLC gradient system was developed to separate NO2-PAH-DNA adducts following one-dimensional TLC and HPLC analysis. HPLC profiles of NO2-PAH-DNA adducts were compared using both adduct enhancement versions of the 32P-postlabeling assay to evaluate the use of this technique on HPLC to screen for the presence of NO2-PAH-DNA adducts. To demonstrate the application of these separation methods to a complex mixture of DNA adducts, the chromatographic mobilities of the 32P-postlabeled DNA adduct standards (PAHs and NO2-PAHs) were compared with those produced by a complex mixture of polycyclic organic matter (POM) extracted from diesel emission particles. The diesel-derived adducts did not elute with the identical retention time of any of the PAH or NO2-PAH standards used in this study. HPLC analyses of the NO2-PAH-derived adducts (butanol extracted) revealed the presence of multiple DNA adducts.(ABSTRACT TRUNCATED AT 250 WORDS)

Formation and behavior of 1-arylpropene-amine exciplexes

Abstract The photochemical reactions of several 1-arylpropenes with diethylamine and trimethylamine in hexane and acetonitrile solution have been investigated. The amines quench the fluorescence of the singlet 1-arylpropenes with rate constants which are dependent upon the aryl substituent. The formation of fluorescent exciplex intermediates is observed for some of the 1-arylpropenes with triethylamine. Preparative irradiation results in the formation of 1-arylpropene-amine adducts as well as 1-arylpropene isomerization, reduction, and dimerization. In the case of 1-phenyl-propene a single adduct is formed with either diethylamine or triethylamine. Mixtures of regioisomeric adducts are formed in the reactions of some of the 1-arylpropenes. The effects of aryl substituents upon exciplex formation and photochemical behavior are discussed.

Mutation spectra in salmonella of complex mixtures: Comparison of urban air to benzo[a]pyrene

We used an ion-exchange procedure coupled to the Salmonella assay to fractionate the dichloromethane-extractable particulate organics from an urban air sample collected in Boise, Idaho. A resulting base/neutral fraction contained 81% of the mutagenic activity but only 36% of the mass of the unfractionated sample. Chemical analysis showed that polycyclic aromatic hydrocarbons (PAHs) accounted for much of the mutagenic activity of the air sample. Colony probe hybridization, PCR, and DNA sequence analysis were then used to determine the mutations induced by the complex mixtures and a model PAH, benzo[a]pyrene (BAP) in approximately 900 revertants of the frameshift hisD3052 allele and approximately 400 revertants of the base-substitution hisG46 allele. The majority (93-94%) of the mutations induced at the frameshift allele in strain TA98 by the whole or base/neutral fraction of the urban air sample was a hotspot 2-base deletion of a CG or GC within the sequence CGCGCGCG. The remaining mutations were complex frameshifts that consisted of -2 or +1 frameshifts associated with a flanking base substitution. BAP induced a somewhat similar pattern of mutations, with 70% being the hotspot mutation, 23% being complex frameshifts, and the remaining being deletions. The inferred base-substitution specificity associated with the complex frameshifts at the hisD3052 allele (primarily G.C-->T.A transversions) was consistent with the observation that this same transversion was the primary mutation induced by the whole urban air sample and BAP at the base-substitution allele in strain TA100. At the frameshift allele, adducts that promote correct incorporation/slippage could account for hotspot mutations, whereas those that promote misincorporation/slippage could account for complex frameshifts. At the base-substitution allele, a mixture of adducts or of adducts with multiple conformations could account for the observed proportion of transitions and transversions. Combined with the bioassay-directed chemical analysis, these results from the first mutation spectra of a complex mixture suggest that such spectra reflect the dominance of particular classes of chemical mutagens within the mixture.

Generation and cycloadditions of 4,5-dihydrooxazole- and oxazolidine-4-carbonitrile N-oxides

A route to 2-phenyl-4,5-dihydrooxazole-4-carbonitrile N-oxide 7 has been developed starting from (S)-serine methyl ester and involving dehydration of the 4-nitromethyl derivative 13 using m-tolylene diisocyanate and triethylamine. Cycloaddition of the nitrile oxide to styrene afforded a 55:45 mixture of diastereoisomeric dihydroisoxazoles 21 and 22; furazan N-oxides 19 and 20 were also formed in a competing dimerisation. The analogous (R)-oxazolidine-4-carbonitrile oxide 8, also prepared from (S)-serine methyl ester, reacted similarly with styrene, oct-1-ene and diethyl fumarate to afford ca. 1:1 mixtures of adducts. The structures of the dihydrooxazole–dihydroisoxazole adduct 21 and furazan N-oxide 20 were determined by X-ray crystallography.

Class-specific immunoadsorption purification for polycyclic aromatic hydrocarbon-DNA adducts.

An immunoenrichment procedure has been developed for applications in the detection and identification of a broad range of polycyclic aromatic hydrocarbon (PAH)-DNA adducts at very low abundance. The procedures are based on a monoclonal antibody raised to r-7,trans-8,trans-9-trihydroxy-cis-10-(N2-deoxyguanosyl-5'-phospha te)- 7,8,9,10-tetrahydrobenzo[a]pyrene (anti-BPDE-N2-dG) which has been tested for cross-reactivity towards DNA and proteins (bovine serum albumin, chicken gamma globulin and human globin) covalently modified with a range of PAH diol-epoxides. The antibody recognised DNA adducted with the diol-epoxides of benzo[a]pyrene, benz[a]anthracene, chrysene, dibenz[a,h]anthracene or picene. The antibody also cross-reacted with the 7,8,9,10-tetraol derived from benzo[a]pyrene and the 1,2,3,4-tetraols of benz[a]anthracene and chrysene. The degree of cross-reactivity was greatest for PAH adducts with structural features similar to anti-BPDE-N2-dG proximate to the base attachment. The antibody also recognised a range of PAHs adducted to human globin; these included adducts of benzo[a]pyrene, benz[a]anthracene and chrysene diol-epoxides. This wide range of recognition provides good evidence for the class-specific recognition of PAH adducts by the antibody. When immobilized on Sepharose 4B and used in the immunoadsorption purification of adducted nucleotides, the antibody selectively enriched adducts of benzo[a]pyrene, benz[a]anthracene and chrysene from normal nucleotides. Quantitative measurements with [14C]benzo[a]pyrene-DNA adducts showed that the immobilized antibody was able to enrich benzo[a]pyrene adducts from a DNA hydrolysate containing adducts at a level of 1 adduct/10(10) normal nucleotides. In addition, this immunoadsorption technique was effective in enriching a mixture of DNA adducts formed in the skin of CF1 mice treated cutaneously with a mixture of [3H]benzo[a]pyrene and [3H]chrysene. Class-specific immunoenrichment procedures for DNA adducts are important in assisting the identification of genotoxic components in complex mixtures. The performance characteristics of this immobilized antibody suggest that it may be suitable for application in the detection, identification and monitoring of human exposures to low levels of PAHs.

Investigation of isothiocyanate addition to alkylidene derivatives of malononitrile and cyanoacetic acid esters

AbstractAddition of arylisothiocyanates to active methylene compounds leads to a variety of compounds depending on the structure of the starting material and conditions used to conduct the addition. Addition of arylisothiocyanate to 1c leads to a pyrido[2,3‐d]pyrimidine resulting from addition of a second mole of cyanate to the initial adduct. Addition of arylisothiocyanate to 1b led to a mixture of pyridine and thiopyran adducts, while addition to 1a led to open chain structures.

Detection of polycyclic aromatic hydrocarbon-DNA adducts in human lung.

Synchronous fluorescence spectroscopy has been combined with immunoaffinity chromatography (IAC) and HPLC to detect polycyclic aromatic hydrocarbon (PAH)-DNA adducts and measure r-7,t-8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE)-DNA adducts in human tissues and cells. A monoclonal antibody (8E11) that recognizes a range of PAH-DNA adducts, but not chemically unrelated adducts, was used to prepare IAC columns. Samples of DNA (25 from human lung and 8 positive and negative controls) were hydrolyzed enzymically and subjected to IAC. Adducts captured by the antibodies and eluted in NaOH (50 mM) were analyzed for fluorescent properties. The spectral fluorescence excitation-emission matrices suggested the presence of mixtures of PAH-DNA adducts in some of the eluates. The eluates were subsequently hydrolyzed with acid (HCl, 0.1 N, 3 hr) and reanalyzed by synchronous fluorescence spectroscopy using a wavelength differential of 34 nm. In 6 of the 25 human lung DNA samples, materials with HPLC retention times identical to benzo[a]pyrene-7,10/8,9-tetrahydrotetrol were found to have fluorescence characteristics indistinguishable from pyrene. Comparisons with appropriate standards indicated that BPDE-DNA adduct levels were between 1 and 40 adducts in 10(8) unmodified nucleotides. No correlation was observed between lung DNA-adduct levels and measures of recent smoking (serum cotinine), but tissue samples taken from different portions of the same lungs showed variation in the DNA adduct levels detected. This finding complicates interpretation of the data and has important implications for the design of future experiments.

A Highly Sensitive and Specific Method for Quantitation of O-Alkylated DNA Adducts and Its Application to the Analysis of Human Tissue DNA

Formation and accumulation of O6-alkylguanine and O4-alkylthymine in human tissues is possibly the most relevant marker for cancer risk. Because humans are chronically exposed to diverse kinds of chemicals and eventual DNA structural modifications are supposed to be a complex mixture of adducts at very low levels, it is essential to use an assay with extremely high sensitivity and specificity. We have established a quantitation method, called PREPI, for O6-methylguanine, O4-methylthymine, and O4-ethylthymine by the combination of prefractionation by HPLC, 32P-postlabeling, and immunoprecipitation. The detection limit was about 1 fmole for all three adducts, enabling us to analyze about 1 x 10(-8) levels as a molar ratio to normal counterpart using 100 micrograms of DNA. In a pilot experiment, we analyzed 11 peripheral blood samples from healthy volunteers. O6-Methylguanine was detected in all the cases with a mean value of 2.0 +/- 1.3 x 10(-8) (range, 0.78-4.6 x 10(-8)). Neither O4-methylthymine nor O4-ethylthymine was above the detection limit of 0.8 x 10(-8) as a ratio to thymine.

A highly sensitive and specific method for quantitation of O-alkylated DNA adducts and its application to the analysis of human tissue DNA.

Formation and accumulation of O6-alkylguanine and O4-alkylthymine in human tissues is possibly the most relevant marker for cancer risk. Because humans are chronically exposed to diverse kinds of chemicals and eventual DNA structural modifications are supposed to be a complex mixture of adducts at very low levels, it is essential to use an assay with extremely high sensitivity and specificity. We have established a quantitation method, called PREPI, for O6-methylguanine, O4-methylthymine, and O4-ethylthymine by the combination of prefractionation by HPLC, 32P-postlabeling, and immunoprecipitation. The detection limit was about 1 fmole for all three adducts, enabling us to analyze about 1 x 10(-8) levels as a molar ratio to normal counterpart using 100 micrograms of DNA. In a pilot experiment, we analyzed 11 peripheral blood samples from healthy volunteers. O6-Methylguanine was detected in all the cases with a mean value of 2.0 +/- 1.3 x 10(-8) (range, 0.78-4.6 x 10(-8)). Neither O4-methylthymine nor O4-ethylthymine was above the detection limit of 0.8 x 10(-8) as a ratio to thymine.