The chemistry and biogenesis of the S-containing metabolites of vinyl chloride in rats.

In order to determine whether vinyl chloride yields chloroethylene oxide in vivo, the biogenesis of the various urinary S-containing metabolites in rats has been investigated.N-Acetyl-S-(2-hydroxyethyl)cysteine is a major vinyl chloride metabolite in rats, but according to the method of protective esterification that is used, so either N-acetyl-S-(2-chloroethyl)cysteine or N-acetyl-S-(2-hydroxyethyl)cysteine may be isolated from the body fluids. N-Acetyl-S-vinylcysteine is a second related metabolite. These S-containing vinyl chloride metabolites are not mutagenic in S. typhimurium. Neutral methanol methylates N-acetyl-S-(2-hydroxyethyl)cysteine. N-Acetyl-S-(2-methoxyethyl)cysteine plus N-acetyl-S-vinylcysteine degrade to give the volatile S-(2-methoxyethyl)(prop-1 or 2-enyl)sulphide.Administration of several vinyl chloride metabolites and closely related compounds to rats shows that chloroacetaldehyde and S-(carboxymethyl)cysteine, but not chloroacetic acid, lie on a pathway or pathways connecting vinyl chloride with thiodiglycollic acid. The fact (a) that chloroacetaldehyde affords both thiodiglycollic acid and N-acetyl-S-(2-hydroxyethyl)cysteine in the animal and (b) that S-(carboxymethyl)cysteine has been identified amongst the hydrolytic products from an hepatic extract prepared from vinyl chloride-treated animals is consistent with the formation of chloroacetaldehyde, and with the reaction of chloroethylene oxide or chloroacetaldehyde with glutathione in the presence of a glutathione S-epoxide transferase to give the identified S-containing metabolites.     

Metabolic and mutagenicity studies on DDT and 15 derivatives. Detection of 1,1-bis(p-chlorophenyl)-2,2-dichloroethane and 1,1-bis(p-chlorophenyl)-2,2,2-trichloroethyl acetate (kelthane acetate) as mutagens in Salmonella typhimurium and of 1,1-bis(p-chlorophenyl) ethylene oxide, a likely metabolite, as an alkylating agent.

Using a novel in vitro technique, whereby microsomal enzymes were embedded in an agar layer to prolong their viability, 1,1-bis(p-chlorophenyl) ethylene(DDNU), a mammalian metabolite of 1,1-bis(p-chlorophenyl)-2,2,2-trichloroethane (DDT), was converted by microsomal mono-oxygenases of mouse liver into 1,1-bis(p-chlorophenyl)-1,2-ethanediol (DDNU-diol). The putative epoxide intermediate, 1,1-bis(p-chlorophenyl)ethylene oxide (DDNU-oxide), a new compound, was synthesized; it showed weak alkylating activity with 4-(4-nitrobenzyl)pyridine but was not mutagenic in Salmonella typhimurium strains TA100 and TA98. DDT and 13 of its metabolites or putative synthetic derivatives, including 1,1-bis(p-chlorophenyl)-2,2-dichloroethylene (DDE), 1 1,1-bis(p-chlorophenyl)-2-chloroethylene (DDMU), 1,1-bis(p-chlorophenyl)-2-chloroethane (DDMS)-DDNU, 2,2-bis(p-chlorophenyl)ethanol (DDOH), bis(p-chlorophenyl)acetic acid (DDA) and 1,1-bis(p-chlorophenyl)-2,2,2-trichloroethanol (Kethane), caused no mutagenic effects in S. typhimurium strains TA100 or TA98, either in the presence or absence of a mouse-liver microsomal fraction. 1,1-Bis(p-chlorophenyl)-2,2,2-trichloroethyl acetate (Kelthane acetate) was a direct-acting mutagen in strain TA100, whereas 1,1-bis(p-chlorophenyl)-2,2-dichloroethane (DDD) was mutagenic in TA98, only in the presence of a mouse-liver microsomal system. The results are discussed in relation to possible pathways whereby DDT is activated to mutagenic and/or carcinogenic metabolites.     

Synthesis and anti-rhinovirus activity of novel 3-[2-(pyridinyl)vinyl]substituted -2H-chromenes and -4H-chromen-4-ones

Human rhinoviruses (HRVs) are the most common cause of viral respiratory infections and their complications. So far, no anti-viral agent has been approved for prevention or treatment of HRV infections. Pursuing our researches on small molecules with anti-rhinovirus activity, in this paper we describe the synthesis and in vitro anti-HRV 1B and 14 properties of new [2-(2H-chromen-3-yl)vinyl]pyridines and 3-[2-(pyridinyl)vinyl]-4H-chromen-4-ones. Generally, the synthesized compounds interfered with the replication of both serotypes at the micromolar or submicromolar concentrations. Preliminary results on their mechanism of action, performed on selected (E)-2-[2-(2H-chromen-3-yl)vinyl]pyridine, indicate an interference with the early step(s) of HRV 1B and 14 replication, probably at the uncoating level.     

Alkylating properties and genetic activity of 4-vinylcyclohexene metabolites and structurally related expoxides

The mutagenicity of the epoxides 4-vinyl-1,2-epoxycyclohexane, 4-epoxy-ethyl-1,2-epoxycyclohexane, 4-epoxyethyl-1,2-dihydroxycyclohexane, 1,2-epoxycyclohexane and styrene oxide was assayed on the TA100 strain of S. typhimurium and V79 Chinese hamster cells. In the latter cell system, both point mutation (6-thioguanine resistance) and chromosomal damage (anaphase bridges and micronuclei) were scored. Genetic effects were related to the alkylating properties of the epoxides. For this purpose, alkylation of 4-(p-nitrobenzyl)pyridine (NPB) and sodium-p-nitrothiophenolate (NTP) was measured and values for the substrate constant (s) were calculated. 4-Epoxyethyl-1,2-epoxycyclohexane, 1,2-epoxycyclohexane and styrene oxide, characterized by the highest reactivity toward NBP and by an s value in the vicinity of 1, were mutagenic in all test systems. 4-Vinyl-1,2-epoxycyclohexane and 4-epoxyethyl-1,2-dihydroxycyclohexane, characterized by lower NBP reactivity and higher s values (1.30–1.38), did not induce reversion in S. typhimurium or 6-thioguanine-resistant mutants in V79 cells, but were as effective as the 3 other compounds in the induction of chromosomal damage.     

Contrasting growth properties of Nocardioides JS614 on threedifferent vinyl halides

Applied Microbiology and Biotechnology - Ethene (ETH)-grown inocula of Nocardioides JS614 grow on vinyl chloride (VC), vinyl fluoride (VF), or vinyl bromide (VB) as the sole carbon and energy...     

手性Schiff－Base在手性PTC条件下的双不对称诱导烷基化

本文报道了以苯甲醛甘氨酸酯〔乙酯,（—）—簿荷醇酯〕西佛碱作为反应底物,烯丙基溴,溴苄,对硝基溴作为烷基化试剂,在以（—）—Ｎ—基氯化辛可宁丁,（＋）－Ｎ基氯化辛可宁作为催化剂的固液相转移条件下的双不对称诱导烷基化反应,进而水解得到α—光学活性氨基酸。光学产率为２．５７—２２．４％,实验中观察到了双不对称诱导效应。     

The biological fate in rats of vinyl chloride in relation to its oncogenicity.

The main eliminative route for [14C]vinyl chloride after oral, i.v. or i.p. administration to rats is pulmonary; both unchanged vinyl chloride and vinyl chloride-related CO2 are excreted by that route and the other [14C] metabolites via the kidneys. After intragastric administration, pulmonary output of unchanged vinyl chloride is proportional to the logarithm of reciprocal dose. Excretion patterns after i.v. and i.p. injections are predictable from the characteristics of excretion following oral administration. Pulmonary excretion of unchanged vinyl chloride after oral dosing is complete within 3-4 h, but pulmonary elimination of CO2 and renal excretion of metabolites occupies 3 days. In comparison, 99% of a small i.v. dose is excreted unchanged within 1 h of injection; 80% within 2 min. The rate of elimination of a single oral doses of [14C]vinyl chloride is uninfluenced by up to 60 days' chronic dosing with the unlabelled substance. The distribution volume of vinyl chloride as displayed by whole-animal autoradiography agrees with deductions from excretion data. Small localization of 14C in the para-auricular region of appropriate sections occurs in sectioned tubules, belonging possibly to the Zymbal glands. Biotransformation of vinyl chloride into S-(2-chloroethyl) cysteine and N-acetyl-S-(2-chloroethyl) cysteine occurs through addition of cysteine, and biotransformation into: (i) chloroacetic acid, thiodiglycollic acid and glutamic acid, and (ii) into formaldehyde (methionine, serine), CO2 and urea is explicable in terms of an associative reaction with molecular O2 involving a singlet oxygen bonded transition state in dynamic equilibrium with a cyclic peroxide ground state. There is no evidence for chloroethylene oxide formation.Thiodiglycollic acid is the major metabolite of chloroacetic acid in rats; more than 60% of the dose. The interaction of vinyl chloride and of its primary metabolites with the intermediates of mammalian metabolism is discussed in relation to the oncogenicity of that substance.     

Nucleophilic selectivity and reaction kinetics of chloroethylene oxide assessed by the 4-(p-nitrobenzyl)pyridine assay and proton nuclear magnetic resonance spectroscopy

The nucleophilic selectivity (Swain-Scott's constant s) of chloroethylene oxide (CEO), an ultimate carcinogenic metabolite of vinyl chloride, was determined to be 0.71 using the 4-(p-nitrobenzyl)pyridine (NBP) assay (Spears method). The molar extinction coefficient of the adduct formed between NBP and CEO was measured; and the second-order rate constants for the reactions of CEO with NBP and with thiosulfate were estimated at three temperatures. The disappearance of CEO and the formation of chloroacetaldehyde (CAA) and glycolaldehyde (GCA) were followed in D2O or a mixture of D2O/hexadeuterated acetone (acetone-d6), using Fourier transform proton nuclear magnetic resonance spectroscopy (1H-FTNMR). Evidence was obtained that CEO reacts with chloride ions to yield CAA at a rate constant of about 17 M-1 h-1 in D2O/acetone-d6 (1 : 1, v/v) at 280 K. Under the same conditions, the first-order rate constant kr for the thermal rearrangement of CEO into CAA was estimated to be approximately 0.41 h-1. These data suggest that the isomerization of CEO may be a minor reaction in physiological saline. These chemical properties of CEO are discussed in relation to the mechanism of vinyl chloride-induced carcinogenesis.     

Adenosine and its receptor agonists regulate nitric oxide production and RAW 264.7 macrophages via both receptor binding and its downstream metabolites-inosine

Adenosine and its receptor agonists enhanced the production of nitric oxide (NO) in lipopolysaccharide (LPS)-treated RAW 264.7 cells. The enhancement of LPS-induced NO production by adenosine, as represented by the amount of its oxidation products, nitrite and nitrate, was inhibited by adenosine uptake inhibitors, such as dipyridamole, S(4-nitrobenzyl)-6-thioinosine (NBTI) and S(4-nitrobenzyl)-6-thioguanosine (NBTG). These indicate that the uptake of adenosine by macrophages is a prerequisite for the enhancement effects observed. A downstream metabolite of adenosine, inosine, also potentiated the LPS-induced NO production in a dose-dependent manner while its enhancement effect was also inhibited by dipyridamole. However, the degree of enhancement by inosine on NO production and nitric oxide synthase (NOS) activity in LPS-treated RAW 264.7 was weaker than the effect of adenosine. Furthermore, adenosine agonists also enhanced the NO production in a dose-dependent manner, but were not specific for A1, A2 nor A3 adenosine receptor. Adenosine uptake inhibitors had no effects on the enhancement activity of the adenosine receptor agonists. Thus, extracellular receptor/s may also play an important role in the observed enhancement responses. The results of this study indicate that the enhancement effects of adenosine on NO production in macrophages could be mediated by the extracellular adenosine receptors as well as the downstream metabolites of adenosine.     

The modification of tryptophan in bovine thrombin.

2-Hydroxy-5-nitrobenzyl bromide, at a 100-fold molar excess, was observed to react withthrombin at pH 4.0 to give a modified enzyme which possessed 20% of the fibrinogen clotting activity and 80% of the esterase activity compared to a control preparation. Spectrophotometric analysis of the modified protein indicated that this effect on catalytic activity was associated with the incorporation of 1 mol of reagent per mol of thrombin. Amino acid analysis showed no loss of amino acids other than tryptophan. The reaction of N-bromosuccinimide with thrombin at 2-fold molar excess resulted in the modification of one tryptophan per mol of enzyme with the loss of 80% of the fibrinogen clotting activity with, as above, a considerably smaller loss of esterase activity. Oxidation of thrombin with N-bromosuccinimide decreased the extent of subsequent tryptophan modification with 2-hydroxy-5-nitrobenzyl bromide. Thrombin modified with 2-hydroxy-5-nitrobenzyl bromide showed a 3-4 fold increase in Km and a decrease in V for the ester substrate. The reaction of thrombin with 2-acetoxy-5-nitrobenzyl bromide, a substrate analogue, also resulted in the inactivation of the enzyme. The data are interpreted to show the presence of a tryptophan residue at or near the enzyme's substrate binding site.     

The chemistry of the reaction of 2-hydroxy-5-nitrobenzyl bromide with his-32 of alpha-lactalbumin.

His-32 of bovine or human alpha-lactalbumin reacts with the tryptophan reagent 2-hydroxy-5-nitrobenzyl bromide at pH 7. The reaction depends on the native conformation of the alpha-lactalbumin molecule and it is restricted to position 1 of the imidazole nucleus. The synthesis and characterization of 1-(2-hydroxy-5-nitrobenzyl)-histidine, 3-(2-hydroxy-5-nitrobenzyl)-histidine and 1,3-bis(2-hydroxy-5-nitrobenzyl)-histidine are described.     

Electrospinning of poly(vinyl alcohol)-water-soluble quaternized chitosan derivative blend

Defect free mats containing a cationic polysaccharide, chitosan derivative such as N-[(2-hydroxy-3-trimethylammonium)propyl] chitosan chloride (HTCC), have been prepared using electrospinning of an aqueous solution of poly(vinyl alcohol) (PVA)-HTCC blends. HTCC, a water-soluble derivative of chitosan, was synthesized via the reaction between glycidyl-trimethylammonium chloride and chitosan. Solutions of PVA-HTCC Blends were electrospun. The morphology, diameter and structure of the produced electrospun nanofibres were examined by scanning electron microscopy (SEM). The average fibre diameter was in the range of 200-600 nm. SEM images showed that the morphology and diameter of the nanofibres were mainly affected by weight ratio of the blend and applied voltage. The results revealed that increasing HTCC content in the blends decreases the average fibre diameter. These observations were discussed on the basis of shear viscosities and conductivities of the spinning solutions. Microbiological assessment showed that the PVA-HTCC mats have a good antibacterial activity against Gram-positive bacteria, Staphylococcus aureus, and Gram-negative bacteria, Escherichia coli.     

Effects of protein-modifying reagents on an isoenzyme of potato apyrase

Treatment of an isoenzyme of potato apyrase of high adenosine triphosphatase/adenosine diphosphatase (ATPase/ADPase) ratio with iodine, N-acetylimidazole or tetranitromethane inactivates the ATPase activity of this enzyme faster than its ADPase activity. There was protection by substrates with the two last-named substances. This and the appearance of nitrotyrosine suggests the participation of tyrosyl residues in both enzymic activities of potato apyrase. The participation of thiol groups is excluded by the insensitivity of apyrase to p-chloromercuribenzoate. Also, 2-hydroxy-5-nitrobenzyl bromide or carboxymethylation produce the same rate of inactivation of ATPase and ADPase activities. Substrates protect both activities from inactivation. Hydrogen peroxide and photo-oxidation inactivate ATPase activity faster than ADPase activity. There is no protection by substrates. Analysis of pH effects on V_max. and K_m suggest different pK values for the amino acid residues at the ATP and ADP sites.     

Oxime-phosphazenes containing dioxybiphenyl groups

2,2-Dichloro-4,4,6,6-bis[spiro(2′,2′′-dioxy-1′,1′′-biphenylyl]cyclotriphosphazene (2) was obtained from the reaction of hexachlorocyclotriphosphazene (1) with biphenyl-2,2′-diol. 2,2-Bis(4-formylphenoxy)-4,4,6,6-bis[spiro(2′,2′′-dioxy-1′,1′′-biphenylyl]cyclotriphosphazene (3) was synthesized from the reaction of 2 with 4-hydroxybenzaldehyde. The novel oxime-cyclophosphazene containing dioxybiphenyl groups (4) was synthesized from the reaction of 3 with hydroxylaminehydrochloride in pyridine. The reactions of this oxime-cyclophosphazene with propanoyl chloride, allyl bromide, acetyl chloride, methyl iodide, benzoyl chloride, 4-methoxybenzoyl chloride, benzenesulfonyl chloride, chloroacetyl chloride, ethyl bromide, benzyl chloride and 2-chlorobenzoyl chloride were studied. Disubstituted compounds were obtained from the reactions of 4 with propanoyl chloride, allyl bromide, acetyl chloride, methyl iodide, benzoyl chloride, 4-methoxybenzoyl chloride, chloroacetyl chloride, ethyl bromide, and 2-chlorobenzoyl chloride, however, the oxime groups on 4 rearranged to nitrile (11) in the reaction of 4 with benzenesulfonyl chloride. A monosubstituted compound was obtained from the reaction of 4 with benzyl chloride. All products were generally obtained in high yields. The structures of the compounds were defined by elemental analysis, IR, ¹H, ¹³C and ³¹P NMR spectroscopy.     

Enantioselective vinylation of aldehydes with the vinyl Grignard reagent catalyzed by magnesium complex of chiral BINOLs

Enantioselective vinylation of aldehydes via direct catalytic asymmetric Grignard reaction of aldehdyes and the vinyl Grinard reagent is a long‐standing challenge. This work demonstrated that the magnesium (S)‐3,3′‐dimethyl BINOLate enantioselectively catalyze the direct vinylation of aldehydes with the deactivated vinylmagnesium bromide by bis(2‐[N,N′‐dimethylamino]ethyl) ether (BDMAEE) in the addition of n‐butylmagnesium chloride. The highest ee of 63% was achieved up to date.     

Quantitative comparison of carcinogenicity, mutagenicity and electrophilicity of 10 direct-acting alkylating agents and of the initial O6:7-alkylguanine ratio in DNA with carcinogenic potency in rodents

The quantitative relationship between carcinogenicity in rodents and mutagenicity in Salmonella typhimurium was examined, by using 10 monofunctional alkylating agents, including N-nitrosamides, alkyl methanesulfonates, epoxides, β-propiolactone and 1,3-propane sultone. The compounds were assayed for mutagenicity in two S. typhimurium strains (TA1535 and TA100) and in plate and liquid assays. The mutagenic activity of the agents was compared with their alkylating activity towards 4-(4′-nitrobenzyl)pyridine and with their half-lives (solvolysis constants) in an aqueous medium. No correlations between these variables were found, nor was mutagenic activity correlated with estimates of carcinogenicity in rodents.

There was a positive relationship between carcinogenicity and the initial ratios of 7-: O⁶-alkylguanine formed or expected after their reaction with double-stranded DNA in vitro. The results suggest that alkylation of guanine at position O⁶ (or at other O atoms of DNA bases) may be a critical DNA-base modification that determines the overall carcinogenicity of these alkylating agents in rodents.     

On the reactivities of the tryptophan residues of human alpha-lactalbumin to 2-hydroxy-5-nitrobenzyl bromide.

The reaction of human alpha-lactalbumin with the tryptophan reagent 2-hydroxy-5-nitrobenzyl bromide has been studied. This protein has 3 tryptophan residues (Trp-60, Trp-104 and Trp-118) all of which are accessible to the reagent at pH 2.7 or 7. Trp-60 of human alpha-lactalbumin is much more reactive than Trp-60 of bovine alpha-lactalbumin (Barman, T. E. (1972) Biochim. Biophys. Acta 257, 297-313). As with bovine alpha-lactalbumin, at pH 2.7, 2-hydroxy-5-nitrobenzyl bromide is specific for tryptophan but at pH 7 His-32 also reacts. When treated with the tryptophan reagent, both alpha-lactalbumins lose their specifier protein activities in the lactose synthase (UDPgalactose:D-glucose 4-beta-galactosyltransferase, EC 2.4.1.22) reaction.     

Structure and action of heteronemertine polypeptide toxins: Inactivation of Cerebratulus lacteus toxin B-IV concomitant with tryptophan alkylation

Reaction of Cerebratulus lacteus toxin B-IV with 2-hydroxy-5-nitrobenzyl bromide at pH 4.5 results in modification of toxin tryptophan residues and loss of biological activity. With relatively small reagent excesses, one tryptophan per molecule is modified without major effect on toxicity. Further reaction results in modification of a second residue of tryptophan and loss of at least 95% of the toxic activity. Modification of one or both tryptophan residues is without significant effect on the secondary structure of the protein. The specificity of each phase of the reaction has been assessed by fingerprint analysis of peptides derived from toxin modified to differing extents with 2-hydroxy-5-nitrobenzyl bromide. It is thus possible to show that tryptophan-5 reacts first and tryptophan-30 only under more rigorous conditions. It thus appears that tryptophan-30 is essential for full neurotoxic activity.     

Syntheses stereoselectives de 1-thioglycosides

2,3,4,6-Tetra-O-acetyl-β-d-mannopyranosyl chloride (2) was obtained in 70% yield by the action of lithium chloride on 2,3,4,6-tetra-O-acetyl-α-d-mannopyranosyl bromide (1) in hexamethylphosphoric triamide. p-Nitrobenzenethiol reacted with 1 and 2 as well as with 2,3,4,6-tetra-O-acetyl-α-d-glucopyranosyl bromide (9) or its β-d-chloro analog (10), giving exclusively and in good yield the corresponding p-nitrophenyl 1-thioglycosides of inverted anomeric configuration. The 1,2-cis-d-manno and -glucop-nitrophenylglycosides were likewise prepared. α-d-Glucopyranosyl 1-thio-α-d-glucopyranoside was similarly obtained by the action of the sodium salt of 1-thio-α-d-glucopyranose on the β-chloride 10 in hexamethylphosphoric triamide, or by treatment of 10 with sodium sulfide, with subsequent deacetylation. Analogous procedures allowed the preparation of β-d-mannopyranosyl 1-thio-β-d-mann opyranoside, the corresponding α,β anomer and α-d-glucopyranosyl 1-thio-α-d-mannopyranoside, starting from bromide 1, 1-thio-α-d-mannopyranose (8),and chloride 10, respectively. When acetone was used as solvent, the reaction between 1 and 8 led instead to the α,α anomer. The thio disaccharides that are interglycosidic 4-thio analogs of methyl 4-O-(β-d-galactopyranosyl)-α-d-galactopyranoside, methyl α-cellobioside, and methyl α-maltoside, respectively, were obtained by way of the peracetates of methyl 4-thio-α-d-galactopyranoside and -glucopyranoside by reaction of the corresponding thiolates with tetra-O-acetyl-α-d-galactopyranosyl bromide, bromide 9, or chloride 10, respectively, in hexamethylphosphoric triamide. These 1-thioglycosides, and (1→1)- and (1→4)-thiodisaccharides, were characterized by ¹H- and ^{1 3}C-n.m.r. spectroscopy. Correlations were established between the polarity of the sulfur atom and certain proton and carbon chemical-shifts in the 1-thioglycosides in comparison with the O-glycosyl analogs; these correlations permitted in particular the unambigous attribution of anomeric configuration.     

Mutagenicity of vinyl chloride,chloroethyleneoxide, chloroacetaldehyde and chloroethanol

Exposure of S. typhimurium strains TA 1530, TA 1535 and G-46 to vinyl chloride increased the number of his⁺ rev./plate 16, 12 or 5 times over the spontaneous mutation rate. The mutagenic response for TA 1530 strain was enhanced 7, 4 or 5-fold when fortified S-9 liver fractions from humans, rats or mice were added. In TA 1530 strain, chloroacetic acid showed only toxic effects, while chloroacetaldehyde, chloroethanol and chloroethyleneoxide caused a mutagenic response. The latter compound was shown to be a strong alkylating agent.