Inhibition of horseradish peroxidase byN-ethylamide ofo-sulfobenzoylacetic acid

The carboxylic groups of horseradish peroxidase were modified by 1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide metho-p-toluenesulfonate by the Koshland method. The catalytic properties of the native and modified peroxidase were studied in the presence ofN-ethylamide ofo-sulfobenzoylacetic acid (EASBA) at pH 5.0–7.5. In the oxidation ofo-dianisidine, EASBA is a competitive inhibitor of the carbidiimide-modified peroxidase, and it increases bothK _m andV _m in the case of the native enzyme. These data show that at least one of the carboxylic groups modified with carbodiimide is located at the area of the peroxidase active site.     

Substrate specificity of peroxidase isoenzymes for hydrogen donors

The investigation of the substrate specificity of the anionic peroxidase isoenzymes, isolated from the zone of differentiation of the primary roots ofZea mays, for some representatives of phenolic compounds and aromatic amines, as hydrogen donors, is reported. The investigation was carried out electrophoretically with peroxidase isoenzymes partially purified by a combination of gel filtration by Sephadex G-25 and Sephadex G-100. A difference in the substrate specificity of the individual isoenzymes is observed. It was established that the anionic peroxidase isoenzymes showed a similarity in total number and relative activity on staining with bivalent phenols and difference on staining with trivalent phenols, as hydrogen donors. A greater number of isoenzymes was stained with benzidine ando-dianisidine and a lesser number witho- andp-phenylendiamine. The substrate specificity of the peroxidase isoenzymes was compared for guaiacol and benzidine. The substrate specificity of peroxidase soenzymes was discussed as regards their diverse role in the plant metabolism.     

NADH diaphorase polymorphism in European fallow deer

NADH diaphorase polymorphism is reported for European fallow deer (D. d. dama) from Coto de Doñana National Park (Spain). Allele frequencies ofp=0.682 (anodic variant) andq=0.318 (cathodic variant) in 110 specimens suggest population genetic usefulness of this biallelic system in an otherwise fairly monomorphic cervid species.     

Regulation of enzymic oxidation of indole-3-acetic acid by phenols: Structure-activity relationships

Mono- and diphenols were tested for their effects on the decarboxylation of [1-¹⁴C]IAA catalysed by purified horseradish peroxidase (EC 1.11.1.7) in the presence or absence of 2,4-dichlorophenol (DCP). The number of hydroxyl groups and their position relative to each other and the nature and position of other substituents on the aromatic ring were found to affect the activity. Although the effects were complex, the following generalizations may be made. (1) Monophenols produce activation when no other cofactor is present. p-Substituted monophenols are more active than o- or m-compounds. In the presence of DCP, the activity varies from slight activation to strong inhibition. (2) m-Diphenols also produce activation in the absence of other cofactors while o- and p-diphenols, with the exception of 3,4-dihydroxyacetophenone and 3,4-dihydroxypropiophenone, produce strong inhibition in the presence or absence of DCP. The o-diphenolsare degraded in the IAA-oxidizing enzyme system and thus produce only a temporary inhibition. (3) m-Diphenols and 3,4-dihydroxyacetophenone produce a sustained inhibition in the presence of DCP. (4) Substitution at position 2 significantly alters the activity of m-diphenols. (5) O-Methylation alters the activity of most o-diphenols. In the absence of DCP, o-methoxyphenols and certain other phenols such as 3,4-dihydroxyacetophenone and 2,6-dihydroxyacetophenone either promote or inhibit IAA oxidation depending on concentration.     

Cytokinin activity of some derivatives of n- n- butyl- n′ - aryl thiourea and their effect on the nitrogen metabolism in barley seedlings

The cytokinin activity of a group of derivatives of N-n-butyl-N′-arylthiourea and their effect on the uptake of amino acids and the accumulation of total nitrogen in seven-day barley seedlings have been investigated. It was established that N-n-butyl-N″-2,4-dinitrophenyl-thiosemicarbazide, N-n-butyl-N′-o-,m- andp-chlorophonylthioureas, N-n-butyl-N′-3,4-dichlorophenylthiourea and N-n-butyl-N′-2-methyl-4-ehlorophenylthiourea possess a high activity with respect to the retardation of the chlorophyll breakdown. Substances with a high cytokinin activity cause an increase in the amount of total nitrogen and amino acids, mainly of glutamic acid, aspartic acid and serine in the roots and loaves of seven-day barley seedlings.     

Peroxidase isozymes of first internodes of sorghum: tissue and intracellular localization and multiple peaks of activity isolated by gel filtration chromatography

Electrophoretic analyses using Sepraphore III strips indicate the presence of a minimum of five bands of peroxidase activity detectable with o-dianisidine and H₂O₂ in extracts from first internodes of Sorghum vulgare var. Wheatland milo. Three of these isozymes were anodic and two were cathodic forms at pH 8.3. The relative amounts of these forms are compared in zero time and incubated excised internodes, stelar and cortical tissues of internodes, and in other parts of the plant. Localization of these isozymes with respect to walls and cytoplasm was characterized by differential centrifugation after grinding of the internodes and by an in situ extraction of walls by centrifugation after vacuum infiltration. Using the latter in situ method, 32% of the total activity of the fast moving cathodic form was exchanged from the wall after infiltration with 50 mm CaCl₂. Only trace amounts of the other isozymes were localized in the walls of the cortex. The isozymes were eluted as two peaks from columns of Sephadex G-100 and three peaks from Agarose A-15m. Although such groupings may be due to asymmetric molecules and ionic interactions as well as to molecular weight differences, they may indicate associations with complexes or membranes of different cytoplasmic constituents.     

Partial characterization of peroxidase isoenzymes from rust-affected wheat leaves

Four anodic peroxidase isoenzymes from wheat leaves were purified by column chromatography and their kinetic behavior with common substrates were examined. One isoenzyme is more active in wheat resistant to stem rust fungi and differed from the others in carbohydrate content and also by a specific activity 2–4-fold higher with non-physiological electron donors. As a substrate, eugenol exhibited kinetic behavior different from p-phenylenediamine, guaiacol or o-dianisidine with all isoenzymes. All four isoenzymes showed similar pH and temperature optima and kinetic behavior and apparent K_m values for both H₂O₂ and non-physiological electron donors.     

Cryoenzymology of β-galactosidase: Effects of cryosolvents

Experimental support for the use of fluid aqueous organic solvent systems and subzero temperatures in mechanistic studies of β-galactosidase is presented. The enzyme was stable and retained catalytic activity and structural integrity in 50% aqueous dimethyl sulfoxide and 60% aqueous methanol at 0°C; at lower temperatures higher concentrations of cosolvent may be successfully used. The effects of dimethyl sulfoxide on the catalytic and structural properties of the enzyme were investigated in detail. For the β-galactoside-catalyzed h ydrolysis ofo-nitrophenyl-β-D-galactoside the value ofk _cat decreased in a linear manner with increasing cosolvent concentration, whereasK _m increased exponentially. The decrease ink _cat paralleled the decrease in water concentration, consistent with rate-limiting hydrolysis of a galactosylenzyme intermediate. The increase inK _m is attributed to less favorable partitioning of the substrate to the active site in the cryosolvent compared to aqueous solution. ThepH*-rate profile for this reaction at 0°C in 50% dimethyl sulfoxide was similar to that in aqueous solution, withpK*₁=5.8 andpK*₂=8.0. Linear Arrhenius plots, with energies of activation of 13.9 and 16.0 kcal mol^?1, respectively, were obtained for the β-galactosidase-catalyzed hydrolysis ofo-nitrophenyl- andp-nitrophenyl-β-D-galactosides in 50% dimethyl sulfoxide at temperatures to ?57°C. Examination of the intrinsic fluorescence and ultraviolet spectra of the enzyme as a function of increasing cosolvent concentration showed no evidence for structural perturbation up to and including 50% dimethyl sulfoxide at 0°C. We conclude that these cryosolvent systems are suitable for mechanistic investigations of β-galactosidase, in particular for trapping intermediates at subzero temperatures.     

Hormonal control of isoperoxidases in lentil embryonic axis

Detachment of the cotyledons from the lentil (Lens culinaris Med.) embryonic axis causes in the latter an increase in total peroxidase activity which is shown to be due to enhancement of specific cathodic isoperoxidases. Kinetin treatment of attached or detached axes promotes activity of essentially the same cathodic isoperoxidases. In addition kinetin enhances the activity of two anodic peroxidases and represses specifically that of a cathodic one. Abscisic acid inhibits the production of all isoenzymes in the presence or absence of kinetin. Cytokinin and abscisic acid actions are discussed in relation to the nature of a wounding hormone and the role of natural inhibitors in cotyledons during germination. Indoleacetic acid stimulates the activity of certain isoenzymes which are also stimulated by kinetin, whereas in others the effects of the two hormones are different. Specific inverse effects of indoleacetic acid and kinetin are demonstrated on the two most cathodic isoperoxidases. Indoleacetic acid-kinetin interactions on the cathodic isoperoxidases have been found in the literature and are discussed as a possible mechanism for explaining interactions of the two regulators on growth and other physiological processes.     

The anaerobic biodegradation ofo-, m- andp-cresol by sulfate-reducing bacterial enrichment cultures obtained from a shallow anoxic aquifer

Summary Sulfate-reducing bacterial enrichments were obtained from a shallow anoxic aquifer for their ability to metabolize eithero-, m-, orp-cresol. GC/MS and simultaneous adaptation experiments suggested that the anaerobic decomposition ofp-cresol proceeds by the initial oxidation of the aryl methyl group to formp-hydroxybenzoic acid. This intermediate was then converted to benzoic acid. Benzoic acid and a hydroxybenzaldehyde were also found in spent culture fluids from ano-cresol-degrading enrichment culture. This result, in addition to others, suggested thato-cresol may also be anaerobically degraded by the oxidation of the methyl substituent. An alternate pathway for anaerobicm-cresol decomposition might exist. Enrichment cultures obtained with eitherp- oro-cresol degraded both of these substrates but notm-cresol. In contrast, am-cresol enrichment culture did not metabolize theortho orpara isomers. Anaerobic biodegradation in all enrichment cultures was inhibited by molybdate and oxygen, and was dependent on the presence of sulfate as a terminal electron acceptor. The stoichiometry of sulfate-reduction and substrate depletion by the various enrichment cultures indicated that the parent cresol isomers were completely mineralized. This result was confirmed by the conversion of¹⁴C-labeledp-cresol to¹⁴CO₂. These results help clarify the fate of alkylated aromatic chemicals in anoxic aquifers.     

Cometabolic transformation ofo-xylene in a biofilm system under nitrate reducing conditions

The purpose of this work was to investigate the anaerobic transformation ofo-xylene in a laboratory biofilm system with nitrate as an electron acceptor.o-Xylene was degraded cometabolically with toluene as primary carbon source. A mass balance showed thato-xylene was not mineralized but transformed.o-Methyl-benzalcohol ando-methyl-benzaldehyde were identified as intermediates ofo-xylene transformation which resulted in the formation ofo-methyl-benzoic acid as an end product. A cross inhibition phenomenon was observed between toluene ando-xylene. The presence of toluene was necessary for stimulation ofo-xylene transformation, but above a toluene concentration of 1–3 mg/L theo-xylene removal rate dramatically decreased. In returno-xylene inhibited the toluene degradation at concentrations above 2–3 mg/L.     

Alternative pathways foro-xylene orm-xylene andp-xylene degradation in aPseudomonas stutzeri strain

Pseudomonas stutzeri OX1 is able to grow ono-xylene but is unable to grow onm-xylene andp-xylene, which are partially metabolized through theo-xylene degradative pathway leading to the formation of dimethylphenols toxic to OX1.P. stutzeri spontaneous mutants able to grow onm-xylene andp-xylene have been isolated. These mutants soon lose the ability to grow ono-xylene. Data from HPLC analyses and from induction studies suggest that in these mutantsm-xylene andp-xylene could be metabolized through the oxidation of a methyl substituent.P. stutzeri chromosomal DNA is shown to share homology with pWW0 catabolic genes. In the mutant strains the region homologous to pWW0 upper pathway genes has undergone a genomic rearrangement.Abbreviations BADH benzylalcohol dehydrogenase - cat catechol - C23O catechol 2,3-dioxygenase - 2,3-,3,4-,2,4-,2,6-,3,5-2,5-DMP 2,3-,3,4-,2,4-,2,6-,3,5-,2,5-dimethylphenol - 2-MBOH 2-methylbenzyl alcohol - 3-MBOH 3-methylbenzyl alcohol - 4-MBOH 4-methylbenzyl alcohol - m-,p-tol m-,p-toluate - o-,m-,p-xyl o-,m-,p-xylene     

Cell Wall Free Space of Cucumis Hypocotyls Contains NAD and a Blue Light-Regulated Peroxidase Activity

Solutions were obtained from the cell wall free space of red light-grown cucumber (Cucumis sativus L.) hypocotyl sections by a low-speed centrifugation technique. The centrifugate contained NAD and peroxidase but no detectable cytoplasmic contamination, as indicated by the absence of the activity of glucose-6-phosphate dehydrogenase from the cell wall solution. Peroxidase activity centrifuged from the cell wall of red light-grown cucumber hypocotyl section could be resolved into at least three cathodic isoforms and two anodic isoforms by isoelectric focusing. Treatment of red light-grown cucumber seedlings with a 10-minute pulse of high-intensity blue light increased the level of cell wall peroxidase by about 60% and caused a qualitative change in the anodic isoforms of this enzyme. The increase in peroxidase activity was detectable within 25 minutes after the start of the blue light pulse, was maximal at 35 minutes, and declined to control levels by 45 minutes of irradiation. The inhibitory effect of blue light on hypocotyl elongation was more rapid than the effect of blue light on total wall peroxidase activity, leading to the conclusion that growth and peroxidase activity are not causally related.     

Mesophilic and thermophilic BTEX substrate interactions for a toluene-acclimatized biofilter

Benzene, toluene, ethylbenzene and xylene (BTEX) substrate interactions for a mesophilic (25°C) and thermophilic (50°C) toluene-acclimatized composted pine bark biofilter were investigated. Toluene, benzene, ethylbenzene, o-xylene, m-xylene and p-xylene removal efficiencies, both individually and in paired mixtures with toluene (1:1 ratio), were determined at a total loading rate of 18.1 g m^–3 h^–1 and retention time ranges of 0.5–3.0 min and 0.6–3.8 min for mesophilic and thermophilic biofilters, respectively. Overall, toluene degradation rates under mesophilic conditions were superior to degradation rates of individual BEX compounds. With the exception of p-xylene, higher removal efficiencies were achieved for individual BEX compounds compared to toluene under thermophilic conditions. Overall BEX compound degradation under mesophilic conditions was ranked as ethylbenzene >benzene >o-xylene >m-xylene >p-xylene. Under thermophilic conditions overall BEX compound degradation was ranked as benzene >o-xylene >ethylbenzene >m-xylene >p-xylene. With the exception of o-xylene, the presence of toluene in paired mixtures with BEX compounds resulted in enhanced removal efficiencies of BEX compounds, under both mesophilic and thermophilic conditions. A substrate interaction index was calculated to compare removal efficiencies at a retention time of 0.8 min (50 s). A reduction in toluene removal efficiencies (negative interaction) in the presence of individual BEX compounds was observed under mesophilic conditions, while enhanced toluene removal efficiency was achieved in the presence of other BEX compounds, with the exception of p-xylene under thermophilic conditions.     

A new species ofScymnodalatias from the southern oceans,and comments on other squaliform sharks

Scymnodalatias albicauda sp. nov. is described from two specimens taken at high latitudes (45°S and 49°S). It is distinguished fromS. sherwoodi, only known species of the genus, by having white markings on the caudal fin, the second dorsal posterior tip almost reaching the upper caudal fin, shorter snout and head, smaller eye and larger fins. Relationships ofScymnodalatias to the generaScymnodon, Centroscymnus, andZameus are discussed, based chiefly on dermal denticle structure.Scymnodalatias andZameus uniquely share transverse ridges on their dermal denticles, and on this character they are treated as sister-groups. Comments on the above genera,Z. squamulosus and some species ofScymnodon are made to clarify their systematic status. As a result, it is proposed thatScymnodon includesichiharai, macracanthus, plunketi, andringens, thatCentroscymnus includescoelolepis, crepidater, crypt acanthus, andowstonii, and thatZameus includessquamulosus.     

Effects of nutritional stress on brain tyramine concentration and dopamine turnover

This is an investigation of the effect of nutritional stress at various ages on the levels of thep- andm-isomers of tyramine in the caudate nucleus of the rat. For comparison, the effects of nutritional stress on the concentration and turnover of dopamine were also studied. Nutritional stress induced in pre-weaning (3 weeks of age) or post-weaning (up to 9 weeks of age) rats resulted in a decrease in the concentration ofp-tyramine and an increase in the concentration ofm-tyramine in the caudate nucleus. Dopamine concentration or turnover in the caudate nucleus was not affected by pre-weaning undernutrition; in the olfactory tubercles, however, a significant decrease was observed in dopamine turnover, calculated from the decrease in homovanillic acid levels after monoamine oxidase inhibition. The results suggest the changes observed are dependent on the availability of the amino acid precursorsp- andm-tyrosine and their competition towards aromatic-l-amino acid decarboxylase.     

Solgerites Reeside, 1932 (Cretaceous Ammonoidea) a synonym ofForresteria Reeside, 1932, with a revision ofSolgerites brancoi (Solger, 1904) (Cretaceous Ammonoidea) from Cameroon

A revision of the surviving “Barroisiceras” specimens from Cameroon described by SOLGER (1904) suggests thatBarroisiceras brancoi, type species ofSolgerites REESIDE, 1932, including varietiesmitis andarmatus of SOLGER, andBarroisiceras hoberfellneri alstadenensis SOLGER non SCHLüTER are variants of a single species and thatSolgerites is a synonym ofForresteria (Forresteria) REESIDE, 1932, of whichEboroceras BASSE, 1947 also is a synonym.     

Redox-mediated oxidation and removal of aromatic amines from polluted water by partially purified bitter gourd (Momordica charantia) peroxidase

Here, the role of bitter gourd peroxidase has been investigated for the treatment of water contaminated with aromatic amines. Most of the aromatic amines were recalcitrant to the action of bitter gourd peroxidase. However, these aromatic amines were oxidized by bitter gourd peroxidase in the presence of a redox mediator, o-dianisidine HCl. The maximum oxidation of aniline was found to be in the buffer of pH 5.0 at 40 °C in the presence of 0.5 mM H₂O₂ and 0.15 mM o-dianisidine HCl. Aromatic amines oxidized and removed from wastewater were 65% aniline, 50% m-toluidine, 86% m-chloroaniline, 54% p-aminobenzoic acid, 61% diphenylamine and 95% N,N-dimethylaniline. Benzidine and p-nitroaniline were recalcitrant to the action of this enzyme even in the presence of o-dianisidine HCl. Complex mixtures of aromatic amines were treated by bitter gourd peroxidase. These mixtures were removed to varying extent, mixtures A, B and C were oxidized to 59%, 56% and 62%, respectively. Mixtures D, E and F were marginally oxidized to 30%, 14% and 16%, respectively.     

Degradation ofortho-chlorophenol,para-chlorophenol, and 2,4-dichlorophenoxyacetic acid by the bacterial community of anaerobic sludge

The bacterial community of anaerobic sludge could degradeo-chlorophenol,p-chlorophenol, and 2,4-dichlorophenoxyacetic acid at concentrations as high as 100 mg/l. The time needed for the degradation of a given chlorinated phenol derivative increased 1.5- to 2-fold upon a twofold increase in its concentration (from 50 to 100 mg/l). The duration of the adaptation period depended on the compound studied and on its concentration. The degradation of 2,4-dichlorophenoxyacetic acid proceeded via 2,4-dichlorophenol andp-chlorophenol as intermediates; the degradation ofo-chlorophenol occurred with the formation of phenol. The dynamics ofp-chlorophenol degradation and chloride ion accumulation were studied.     

Correlation between Chlorophyll and Chlorogenic Acid Content in Tobacco Leaves

Developmental stages of tobacco (Nicotiana tabacum L. cv. Burley 21) flower and capsule were correlated with tissue contents of polyphenols and activities of phenylalanine ammonialyase, polyphenoloxidase, and peroxidase. Chlorogenic acid, scopolin, and scopoletin were present in most tissues, whereas rutin and two dihydroxyphenolic glycosides concentrated primarily in the corolla and placenta, respectively. Ovules contained only chlorogenic acid. As development progressed, polyphenols accounted for nearly 15% of the dry weight in the green capsule of field-grown plants. Fertilization triggered a rapid increase of chlorogenic acid in the ovary. When l-phenylalanine-U-¹⁴C was fed to the detached green capsules and capsule parts, an incorporation of radioactivity into chlorogenic acid and dihydroxyphenolic glycosides occurred which suggested in situ synthesis of these compounds. This was subtantiated by a positive correlation between phenylalanine ammonia-lyase activity and polyphenol accumulation. High polyphenoloxidase activity was associated mainly with the ovary, whereas peroxidase activity was maximal during senescence of all tissues. Polyacrylamide gel slab electrophoresis revealed five cathodic bands and one diffuse zone with poly-phenoloxidase activity in flower extracts. Two anodic poly-phenoloxidase isozymes appeared only in the fertilized ovary. Among 17 peroxidase isozymes, six cathodic forms were present throughout floral development, and the anodic ones increased in number and activity at the later stages of capsule growth.