Effect of Transpiration-reducing Chemicals on Growth, Flowering, and Stomatal Opening of Tomato Plants

Phenyl mercuric acetate, 8-hydroxyquinoline, N-dimethylamino succinamic acid, or 2-chloroethyl trimethyl ammonium chloride were sprayed on 37-day-old tomato (Lycopersicon esculentum Mill. cv. Sioux) plants seven times at weekly intervals. Plants of nearly normal appearance resulted with all treatments except 2-chloroethyl trimethyl ammonium chloride. There was no change in leaf number, but 2-chloroethyl trimethyl ammonium chloride increased the number of flowers. 2-Chloroethyl trimethyl ammonium chloride and phenyl mercuric acetate caused earlier flowering. Yield was not affected significantly. Stomatal opening was reduced 80% immediately after spraying with phenyl mercuric acetate or 2-chloroethyl trimethyl ammonium chloride, but 6 days after spraying, the reduction in stomatal opening was only 30 to 40%. Wilting was delayed 8 days by phenyl mercuric acetate and 4 days by 2-chloroethyl trimethyl ammonium chloride and N-dimethyl amino succinamic acid treatments, when water was withheld 59 days after the final spray application.     

Penetration of chemicals into the Malus leaf cuticle

The adaxial leaf cuticle of Malus pumila was examined by electron microscopy to determine possible avenues for transcuticular movement of foliarly applied chemicals. Cutin-embedded polysaccharide microfibrils originated at the outer epidermal cell wall and occasionally extended to the cuticle surface. Lamellae, ca. 4 nm wide, usually were oriented parallel to the cuticle surface. When oriented perpendicular to the surface, they extended nearly to the subjacent wall layer from the surface. Aqueous solutions of uranyl acetate, silver nitrate and phenyl mercuric acetate applied to the cuticle surface of leaf segments floated on solutions of phosphate salts or thiocarbohydrazide (TCH) reacted within the cuticle to form insoluble electron-opaque deposits indicative of their avenues of transcuticular movement. Uranyl phosphate deposits were observed only in the polysaccharide microfibrils of chloroform: methanolextracted leaves. Silver-TCH deposits were observed in the microfibrils of both extracted and nonextracted leaf cuticles. Phenyl mercuric acetate-TCH deposits were randomly dispersed throughout the extracted cuticle and not associated with the polysaccharide microfibrils.Abbreviations TCH thiocarbohydrazide - PMA phenyl mercuric acetate     

Synthesis and characterization of methyl 6-O-alpha- and -beta-D-galactopyranosyl-beta-D-galactopyranoside

Sequential tritylation, acetylation and detritylation of methyl beta-D-galactopyranoside gave crystalline methyl 2,3,4-tri-O-acetyl-beta-D-galactopyranoside (4) and methyl 2,3,6-tri-O-acetyl-beta-D-galactopyranoside, the latter being the minor product resulting from acetyl migration. Reaction of 4 with 2,3,4,6-tetra-O-acetyl-alpha-D-galactosyl bromide in benzene, in the presence of mercuric cyanide and mercuric bromide, gave the alpha- and beta-D-(1----6)-linked disaccharides (7 and 9, respectively) in high yield, and their structure was confirmed by 1H- and 13C-n.m.r. 1d. and 2d. spectroscopy. O-Deacetylation of 7 gave the hitherto unknown, crystalline methyl 6-O-alpha-D-galactopyranosyl-beta-D-galactopyranoside. O-Deacetylation of 9 gave the corresponding, beta-D-linked disaccharide methyl glycoside, the physical constants of which are discussed with respect to controversial data in the literature.     

High-pressure liquid chromatographic analysis for identification of in vitro and in vivo metabolites of 4-phenethyl-5-[4-(1-(2-hydroxyethyl)-3,5-dimethyl-4-pyrazolylazo)phenyl]-2,4-dihydro-3H-1,2,4-triazole-3-thione in rats

All azo colorants whose metabolism can liberate a carcinogenic arylamine, are suspected of having carcinogenic potential. Therefore, a new azo compound 4-phenethyl-5-[4-(1-(2-hydroxyethyl)-3,5-dimethyl-4-pyrazolylazo)phenyl]-2,4-dihydro-3H-1,2,4-triazole-3-thione (substrate) was prepared to investigate its in vitro and in vivo biotransformation in rats by HPLC. Chromatographic separation of substrate and its metabolites was performed using a Chromasil C(18) column. The mobile phase consisted of acetonitrile and water in a linear gradient system. From the biotransformation of this compound, the reduction metabolite 4-(2-phenethyl)-5-(4-aminophenyl)-2,4-dihydro-3H-1,2,4-triazole-3-thione was identified by comparing it to reference standard by HPLC-DAD. In the in vivo study, identification of the unknown peak which was the N-acetylation metabolite was confirmed by LC-MS spectrometry. Besides this, the azo compound was reduced to its corresponding amine in intestinal and cytosolic parts. In addition, oxidation of the methyl group and the phenyl ring, and reduction of azo group to hydrazo were identified in the cytosolic part using LC-MS.     

Interaction of aspartate aminotransferase with mercurochrome. Relationship of an exposed thiol group of the enzyme to the active centre.

Mercurochrome strongly inhibits aspartate transaminase and 2,3-dicarboxyethylated aspartate transaminase. The native enzyme exhibits a biphasic time-course of inactivation by mercurochrome with second-order rate constants 1.62 x 10(4) M-1 - min-1 and 2.15 x 10(3) M-1 - min-1, whereas the modified enzyme is inactivated more slowly (second-order rate constant 6.1 x 10(2) M-1 - min-1) under the same conditions. The inhibitor inactivates native and modified enzyme in the absence as well as in the presence of substrates. Mercurochrome-transaminase interaction is accompanied by a red shift in the absorption maximum of the fluorochrome of about 10 nm. Difference spectra of the mercurochrome-enzyme system versus mercurochrome, compared with analogous spectra of mercurochrome-ethanol, revealed that the spectral shifts recorded during mercurochrome-transaminase interaction are similar to those that occur when mercurochrome is dissolved in non-polar solvents. Studies of mercurochrome complexes with native or modified transaminase, isolated by chromatography on Sephadex G-25, revealed that native transaminase is able to conjugate with four mercurochrome molecules per molecule, but the modified enzyme is able to conjugate with only two mercurochrome molecules per molecule.     

Pseudomonas putida Strains Which Constitutively Overexpress Mercury Resistance for Biodetoxification of Organomercurial Pollutants

Improved biocatalysts for mercury (Hg) remediation were generated by random mutagenesis of Pseudomonas putida with a minitransposon containing merTPAB, the structural genes specifying organomercury resistance. Subsequent selection for derivatives exhibiting elevated resistance levels to phenylmercury allowed the isolation of strains that constitutively express merTPAB at high levels, conferring the ability to cleave Hg from an organic moiety and reduce the freed Hg(II) to the less toxic elemental form, Hg⁰, at greater rates. Constitutive overexpression of merTPAB had no apparent effect on culture growth rates, even when Hg(II) was initially present at otherwise toxic concentrations. These properties were also combined with benzene and toluene catabolism, allowing detoxification of the metal component of phenyl mercuric acetate, as well as degradation of its aromatic moiety.     

Biomonitoring of low levels of mercurial derivatives in water and soil by Allium micronucleus assay

The Allium micronucleus (MNC) assay was developed to monitor low levels of mercury in aquatic and terrestrial environments. Four mercurial derivatives namely mercuric chloride (MC), methyl mercuric chloride (MMC), phenyl mercuric acetate (PMA) and a methoxy ethyl mercuric chloride based fungicide, Emisan-6, were tested to assess the sensitivity and versatility of the Allium MNC assay. Allium bulbs were set directly on water and soil contaminated with known levels of mercurial derivatives (0.0001-10.00 ppm). On the 5th day the endpoints measured were root length, mitoses with spindle abnormality and cells with MNC in root meristems. The effective concentrations of the test chemicals that cause 50% of root length as compared to control (EC50) were determined from dose-response curves so obtained. The lowest effective concentration tested (LECT) and highest ineffective concentration tested (HICT) for each of the mercurial derivatives for the induction of spindle malfunction and MNC were determined. It was found that EC50, LECT and HICT values for mercurial derivatives in soil were higher than those in water. The frequencies of cells with MNC and mitoses with spindle abnormality were highly correlated indicating that MNC is a good parameter of spindle malfunction. The present approach increased the sensitivity of the Allium assay by 10-fold, the detection limit being 0.001-0.1 ppm and 0.1-1.0 ppm in aquatic and terrestrial environments respectively, depending on the species of mercury.     

Genetic and physical map of the 2,4-dichlorophenoxyacetic acid-degradative plasmid pJP4.

Plasmid pJP4 is an 80-kilobase, IncP1, broad-host-range conjugative plasmid of Alcaligenes eutrophus encoding resistance to mercuric chloride and phenyl mercury acetate and degradation of 2,4-dichlorophenoxyacetic acid, 2-methyl-4-chlorophenoxyacetic acid, and 3-chlorobenzoate. By the use of cloning, transposon mutagenesis, and restriction endonuclease analysis, a biophysical and genetic map of pJP4 was generated.     

A method for the identification of human peripheral blood T lymphocytes by sequential immunogold and esterase double staining

A double staining method involving the sequential use of monoclonal OKT hybridoma antibodies applied in the colloidal immunogold method and followed by a simultaneously capturing azo dye method for the detection of acid alpha-naphthyl acetate esterase (ANAE) is described. Mononuclear leukocytes isolated from human peripheral blood using a Ficoll-Hypaque density gradient were stained. M-pattern ANAE-positive monocytes (diffuse staining) were excluded from the lymphocyte counts. 80 +/- 5% of all lymphocytes were T-pattern ANAE positive (dot-like staining) and 77 +/- 3% were OKT3 positive. 86 +/- 6% of all ANAE-positive T-pattern lymphocytes were also OKT3 positive, and 89 +/- 6% of all OKT3-positive lymphocytes were also ANAE positive. This indicates that ANAE is a good marker for total human T lymphocytes. 53 +/- 10% of human peripheral blood lymphocytes were OKT4 positive and 87 +/- 8% of all OKT4-positive lymphocytes were also ANAE positive. 30 +/- 6% of all lymphocytes were OKT8 positive, and only 26 +/- 18% of all OKT8-positive lymphocytes were ANAE negative. This indicates that ANAE cannot be used to distinguish T-helper and T-suppressor lymphocytes as identified by monoclonal antibodies.     

黑磷纳米片光催化甲基橙降解机理

偶氮废水大量排放到环境中会对生态系统和人类健康造成严重威胁,发展偶氮染料废水的高效处理技术具有实际意义。光催化法由于工艺简单、处理彻底等优点具有应用前景。本研究利用液相剥离法制备黑磷纳米片(LBP),以甲基橙(MO)为例,考察LBP对偶氮染料的光催化能力;利用淬灭及荧光探针试验判断体系中参与反应的主要瞬态物种;结合液相色谱-质谱的产物鉴定结果,阐明光催化降解机理。结果表明: MO在酸性(pH=3.0)和碱性(pH=11.0)条件下的降解速率(k_obs)高于中性条件下(pH=7.0)。LBP光照下产生羟基自由基(·OH)进攻偶氮键使双键断裂生成中间产物,后者被·OH继续氧化,生成主要降解产物N,N-二甲基-4-(2-对苯甲基肼)苯胺、2-二甲胺基-5-((4-(二甲胺基)苯基)二氮基)苯酚和N,N-二甲基-4-硝基苯胺。     

Purification and properties of an enzyme catalyzing the splitting of carbon-mercury linkages from mercury-resistant Pseudomonas K-62 strain. I. Splitting enzyme 1.

An enzyme (S-1) which catalyzes the splitting of carbon-mercury linkages of organomercury compounds was purified about 24-fold from the cell-free extract of mercury-resistant Pseudomonas K-62 strain by treatment with streptomycin, precipitation with ammonium sulfate, and successive chromatography on Sephadex G-150, DEAE-Sephadex, and DEAE-cellulose. A purified preparation of the enzyme showed a single band on polyacrylamide gel electrophoresis, and was colorless. The molecular weight of the enzyme was estimated to be 19,000, and Km was 5.3 X 10(-5) M for p-chloromercuribenzoic acid (PCMB). The temperature and pH optimum for the reaction were 50degrees and 7.0, respectively. The enzyme was capable of catalyzing the decomposition of methylmercuric chloride (MMC), ethylmercuric chloride (EMC), phenylmercuric acetate (PMA), and PCMB in the presence of a sulfhydryl compound to form a mercuric ion plus methane, ethane, benzene, or benzoic acid, respectively. The mercuric ion thus formed was reduced to metallic mercury by metallic mercury-releasing enzyme (MMR-enzyme).     

Studies on the 1,1-diphenyl-2-picrylhydrazyl radical scavenging mechanism for a 2-pyrone compound

The radical scavenging mechanisms for the 2-pyrone compound, 4-hydroxy-3,6-dimethyl-2H-pyrane-2-one (1), and the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical (4) in several solvent systems were evaluated by the quantitative change in compounds detected at 270 nm and subsequent HPLC analyses. The HPLC profile for each condition suggested that the reaction proceeded by a different mechanism in each solvent system. In organic solvents (CHCl3, iso-propanol, and EtOH), 1-[4-(3,4-dihydro-3,6-dimethyl-2,4-dioxo-2H-pyran-3-yl) phenyl]-1-phenyl-2-picrylhydrazine (2) was produced as an adduct of the DPPH radical and 1. On the other hand, the reaction in a buffer solution (an acetate buffer at pH 5.5) gave several degradation products with 1[4-(2,3-dihydro-2,5-dimethyl-3-oxo-fur-2-yl) phenyl]-1-phenyl-2-picrylhydrazine (5), this being structurally elucidated by spectroscopic analyses. The decrease of the DPPH radical in each reaction system suggests that compound 1 could scavenge about 1.5-1.8 equivalents of the radical in organic solvents and about 3.5-3.9 in the buffer solution.     

Degradation of Organomercurials in Bacillus cereus

Degradation of phenylmercuric acetate (PMA) and p-chloromercuribenzoic acid (PCMB) by cells of a mercury resistant strain of Bacillus cereus was demonstrated. Degradation of PMA was also demonstrated with the crude extract of the B. cereus cells grown in nutrient broth containing 1 μm PMA or 10 μm HgCl₂. The crude extract seems to split the carbon-mercury bond of PMA and form benzene and the mercuric ion as degradation products. The splitting enzyme was separated from mercuric reductase which catalyzes the reduction of the mercuric ion by gel filtration on Sephadex G 100.     

On reactions of steroidal 23-oxo and 23,24-epoxysapogenins with Lewis acids

The reaction of 23-oxotigogenin acetate with TMSOTf in THF afforded the corresponding bisnorcholanic lactone in 60% yield. The analogous reactions carried out in dichloromethane or benzene gave the rearranged products—the isomeric spirostanic ketone (10-15%) and bisfuran (40-42%). Similar products were also obtained upon treatment of 23,24-epoxysapogenins with BF₃. The epoxides treated with TiCl₄ afforded mostly chlorohydrins and no rearranged products were detected.     

Benzofuran- and furan-2-yl-(phenyl)-3-pyridylmethanols: synthesis and inhibition of P450 aromatase

A series of benzofuran-2-yl-(phenyl)-3-pyridylmethanol derivatives were prepared using an efficient 1-step procedure in good yields. In addition furan-2-yl-(phenyl)-3-pyridylmethanol derivatives were also prepared to determine the effect of the benzene ring in benzofuran with respect to inhibitory activity. The pyridylmethanol derivatives were all evaluated in vitro for inhibitory activity against aromatase (P450(AROM), CYP19), using human placental microsomes. The benzofuran-2-yl-(phenyl)-3-pyridylmethanol derivatives showed good to moderate activity (IC50 = 1.3-25.1 microM), which was either better than or comparable with aminoglutethimide (IC50 = 18.5 microM) but lower than arimidex (IC50 = 0.6 microM), with the 4-methoxyphenyl substituted derivative displaying optimum activity. Molecular modelling of the benzofuran-2-yl-(4-fluorophenyl)-3-pyridylmethanol derivative suggested activity to reside with the (S)-enantiomer. The furan-2-yl-(phenyl)-3-pyridylmethanol derivatives were devoid of activity indicating the essential role of the benzene ring of the benzofuran component for enzyme binding.     

Distribution of free sulfhydryl and disulfide groups among platelet membrane proteins.

Reactive sulfhydryl and disulfide groups were identified in platelet membrane proteins resolved by sodium dodecyl sulfate-polycrylamide gel electrophoresis. Platelet membranes treated with N-ethyl(1-14C)maleimide, phenyl(203Hg)mercuric acetate and p-chloro(203Hg)mercuribenzoate showed similar patterns of distribution of sulfhydryl groups among the sodium dodecyl sulfate-solubilized membrane proteins. Four major and two minor polypeptides ranging in molecular weight from greater than 200 000 to 20 000 were found to have reactive SH groups. Reduction of membrane proteins by sulfite coupled with subsequent mercaptide formation of the resultant monothiols led to the identification of four polypeptides with disulfide bonds. Reaction of platelet membranes with 14C-labeled 5,5'-dithio-bis(2-nitrobenzoic acid) resulted changes in the distribution profile of the solubilized membrane proteins suggestive of a polymerization process dependent upon, 5,5'-dithio-bis(2-nitrobenzoic acid)-induced intermolecular disulfide interchange.     

Synthesis and antimalarial activity of some new 1,2-dioxolane derivatives

The synthesis of 1,2-dioxolane derivatives in two different acetophenone series, as simplified models of natural coumarins is described. 2-Acetyl-3-acetoxy-4-(3-hydroperoxy-3-methylbut-1-enyl)phenyl acetate and 2-acetyl-5-acetoxy4-(3-hydroperoxy-3-methylbut-1-enyl) phenyl acetate synthons are used as precursors to these structures. In vitro antimalarial activity of the 1,2-dioxolane derivatives has been investigated.     

On the Productivity and Properties of l-Asparaginase from Escherichia coli A–l–3

Oxidation of grayanotoxin (GTX) II with lead (IV) acetate in methanol gave a new derivative, the 1(R)-spiro-3,6(S),14,16-tetra-hydroxy-5-keto derivative. Treatment of GTX-II tetraacetate in acetic acid by using Pb(IV) acetate as an oxidizing agent gave a novel 1,5-seco-GTX derivative, Δ¹⁽¹⁰⁾-1,5-seco-GTX-pentaacetate, together with the 1,5-seco-GTX-1(R) derivative. Oxidation of GTX-II-tetraacetate with Tl(III) acetate in acetic acid or benzene gave the 1,5-seco-GTX-1(S) derivative.     

Fluorescence energy transfer measurements of spatial relationships between sulfhydryl groups of thiolase I from porcine heart

Mitochondrial thiolase I from pig heart has been found to have at least two and possibly three reactive sulfhydryl residues at or near the active site [Izbicka-Dimitrijevi?, E., & Gilbert, H. F. (1982) Biochemistry 21, 6112-6118; Izbicka-Dimitrijevi?, E., & Gilbert, H. F. (1984) Biochemistry 23, 4318-4324]. In the native enzyme, fluorescein mercuric acetate reacts with two of the sulfhydryl groups and inactivates the enzyme with a rate constant of 1.6 X 10(-4) M-1 s-1 in 0.1 M Tris-acetate, pH 7.0. The presence of saturating (250 microM) concentrations of acetoacetyl coenzyme A protects against both modification and inactivation. The acetyl enzyme, a normal intermediate in the reaction catalyzed by thiolase, is not inactivated by fluorescein mercuric acetate although one sulfhydryl group out of five per mole of thiolase subunit is still available for reaction with the reagent. Fluorescein mercuric acetate and S-mercurio-N-dansyl-L-cysteine (Dns-Cys-SHg+) have been used to differentially label two of the sulfhydryl groups in thiolase. The distance between Dns-Cys-SHg+ (donor) and fluorescein mercuric acetate (acceptor) determined by the fluorescence energy transfer is less than 14 A. The fluorescent analogue of coenzyme A, 1,N6-etheno coenzyme A, is recognized by thiolase as a substrate (Km = 21 microM); however, substrate inhibition and equilibrium dialysis show that the affinity of the free enzyme for CoA is quite low (Ki = 100 microM). The quantum yield of the fluorescence of the three thiolase tryptophan residues is low (0.024), corresponding to about 12% of the fluorescence expected from equivalent concentrations of tryptophan.     

Synthesis and Antimalarial Activity of Some New 1,2-Dioxolane Derivatives

The synthesis of 1,2-dioxolane derivatives in two different acetophenone series, as simplified models of natural coumarins is described. 2-Acetyl-3-acetoxy-4-(3-hydroperoxy-3-methylbut-1-enyl)phenyl acetate and 2-acetyl-5-acetoxy-4-(3-hydroperoxy-3-methylbut-1-enyl) phenyl acetate synthons are used as precursors to these structures. In vitro antimalarial activity of the 1,2-dioxolane derivatives has been investigated.