Zinc increases vulnerability of rat thymic lymphocytes to arachidonic acid under in vitro conditions

Previous studies on the cytotoxicity of arachidonic acid (ARA) elucidated the involvement of oxidative stress and Ca²⁺. In the present study, the Zn²⁺-related cytotoxicity of ARA was studied by a flow cytometric technique with appropriate fluorescent probes in rat thymocytes. Addition of 10 μM ZnCl₂ enhanced the increase in cell lethality induced by 10 μM ARA. The removal of Zn²⁺ by Zn²⁺ chelators attenuated the ARA-induced increase in cell lethality. Thus, Zn²⁺ is suggested to be involved in ARA cytotoxicity. ARA at 3–10 μM elevated intracellular Zn²⁺ level. The Zn²⁺ chelators attenuated the ARA-induced increase in intracellular Zn²⁺ level while ARA significantly increased intracellular Zn²⁺ level in the presence of 3 μM ZnCl₂, suggesting the involvement of external Zn²⁺. Zn²⁺ reportedly exerts cytotoxic action under oxidative stress induced by hydrogen peroxide, via an excessive increase in intracellular Zn²⁺ levels. Since ARA induces oxidative stress, the simultaneous administration of zinc and ARA may be harmful.     

Membrane-permeant chelators can attenuate Zn2+-induced cortical neuronal death

Chelating extracellular Zn²⁺ with the membrane-impermeant Zn²⁺ chelator, CaEDTA, can inhibit toxic Zn²⁺ influx and subsequent neuronal death. However, this drug does not cross the blood-brain barrier. In the present study, we explored the ability of two membrane-permeant Zn²⁺ chelators to inhibit Zn²⁺-induced death of cultured cortical neurons. Addition of either the high affinity (K_D=10^−15.6) Zn²⁺ chelator, N, N, N’, N’, tetrakis (2-pyridylmethyl) etylenediaminepentaethylene (TPEN), or the low affinity (K_D=10⁻⁶) Zn²⁺ chelator, 1-hydroxypyridine-2-thione (pyrithione), to the culture medium following exposure to extracellular Zn²⁺ reduced subsequent neuronal death, even if chelator administration was delayed by up to 1 h. Indeed, some delay was essential for neuroprotection with pyrithione, as co-administration of pyrithione together with extracellular Zn²⁺ increased levels of [Zn²⁺]_i and cell death compared to the levels induced by Zn²⁺ alone. TPEN, but not pyrithione, was intrinsically toxic at high concentrations, likely due to excessive chelation of [Zn²⁺]_i, as this intrinsic toxicity was reduced by prior addition of extracellular Zn²⁺. These data point to a potential therapeutic role for membrane-permeant Zn²⁺ chelators, perhaps especially possessing low Zn²⁺ affinity, in attenuating neuronal death after certain acute insults.     

Zn2+, derived from cell preparation,partly attenuates Ca2+-dependent cell death induced by A23187, calcium ionophore,in rat thymocytes

A23187, a calcium ionophore, is used to induce Ca²⁺-dependent cell death by increasing intracellular Ca²⁺ concentration ([Ca²⁺]_i) under in vitro condition. Since this ionophore also increases membrane permeability of metal divalent cations such as Zn²⁺ and Fe²⁺ rather than Ca²⁺, trace metal cations in cell suspension may affect Ca²⁺-dependent cell death induced by A23187. Therefore, the effects of chelators for divalent metal cations, EDTA and TPEN, on the A23187-induced cytotoxicity were cytometrically examined in rat thymocytes. The cytotoxicity of A23187 was attenuated by 1 mM EDTA while it was augmented by 50 μM EDTA and 10 μM TPEN. These changes were statistically significant. The A23187-induced increase in Fluo-3 fluorescence intensity, a parameter for [Ca²⁺]_i, was significantly reduced by 1 mM EDTA while it was not the case for 50 μM EDTA and 10 μM TPEN. The intensity of FluoZin-3 fluorescence, a parameter for [Zn²⁺]_i, increased by A23187 was respectively reduced by 50 μM EDTA and 10 μM TPEN. It is suggested that the attenuation of A23187-induced cytotoxicity by 1 mM EDTA is due to the chelation of extracellular Ca²⁺ and Zn²⁺ while the augmentation by 50 μM ETDA or 10 μM TPEN is due to the chelation of extracellular Zn²⁺. The Tyrode’s solution without thymocytes contained 32.4 nM of zinc while it was 216.9 nM in the cell suspension. In conclusion, trace Zn²⁺, derived from cell preparation, partly attenuates the Ca²⁺-dependent cell death induced by A23187.     

Correlation of the physicochemical properties of metal ions with their activation and inhibition of human erythrocytic δ-aminolevulinic acid dehydratase (ALAD) in vitro

Using normal adult human whole blood hemolysates this study determined, in a dose-response fashion, the in vitro effects of Na⁺, Mg²⁺, Al³⁺, Mn²⁺, Cu²⁺, Ag⁺, Zn²⁺, Cd²⁺, Hg²⁺, Ga³⁺, In³⁺, Sn²⁺, Sn⁴⁺, and Pb²⁺ on normal erythrocytic ALAD. The effects of these 14 metal ions on erythrocytic ALAD 50% inhibited by Pb²⁺ were also determined as was the ability of a maximum stimulatory concentration of Zn²⁺ to prevent or reverse the effects of these metal ions on erythrocytic ALAD. The effects of these metal ions were then classified in terms of their oxidation state, characteristic coordination number, coordination geometry, and hard and soft Lewis acid characteristics in order to determine the physical and chemical properties associated with the ability of a metal ion to activate or inhibit erythrocytic ALAD and whether these properties are unique to a single metal ion. Preincubation studies established Zn²⁺ to be the only metal ion to both activate erythrocytic ALAD and to prevent or reverse the Pb²⁺-induced inhibition of erythrocytic ALAD in vitro even after prolonged contact with the enzyme. Therefore future investigations of the use of nontoxic salts of Zn²⁺ as a prophylactic agent or therapeutic adjunct in the prevention or treatment of lead poisoning with its possibly toxic accumulation of ALA are proposed.     

Chelator-induced phytoextraction of zinc and copper by rice seedlings

Solution culture was carried to investigate capacity of synthetic aminopolycarboxylic acids (ethylenediamine tetraacetate, N-hydroxyethylenediaminetriacetic acid, and diethylenetriamine-pentaacetate) for enhancing botanical removal and transport of heavy metals (Cu and Zn) by plants. Biodegradable organic acids (citric acid, malic acid, and oxalic acid) were also selected as alternatives to compare them with synthesized chelating agents for effectiveness. Young rice seedlings (Oryza sativa L. cv. XZX 45) were grown in nutrient solutions treated with single or combined metal solutions in presence or absence of chelating compounds. Calculation by chemical equilibrium program VISUAL MINTEQ showed that different chelating compounds had various complex potential with Cu²⁺ and Zn²⁺ ions, in which synthetic chelators exhibited higher complexed capability than biodegradable organic acids. All applied synthetic aminopolycarboxylic acids significantly decreased removal of metal from nutrient solution (p < 0.01), while more or less effects of organic acids supplied on biosorptive potential were observed with all treatments (p > 0.05), compared with the treatment without metal ligands. Synthetic aminopolycarboxylic acids significantly decreased metal concentrations in plant materials in all treatments (p < 0.01). However, biodegradable organic acids decreased metal concentrations in roots (p < 0.01), but enhanced them in shoots (p < 0.01). Results obtained indicated that synthetic aminopolycarboxylic acids decreased uptake of metals by rice seedlings, but translocation of metals complexed within plant materials was evident. Although exogenous biodegradable organic acids showed negligible effect on botanical removal of metals, metals complexed with organic acids was more mobile than those complexed with other chelating agents. These information collected here had important implication for the use of biodegradable metal chelators in transport of essential micronutrients in plant nutrition.     

Aflatoxin B1 Degradation by a Pseudomonas Strain

Aflatoxin B₁ (AFB₁), one of the most potent naturally occurring mutagens and carcinogens, causes significant threats to the food industry and animal production. In this study, 25 bacteria isolates were collected from grain kernels and soils displaying AFB₁ reduction activity. Based on its degradation effectiveness, isolate N17-1 was selected for further characterization and identified as Pseudomonas aeruginosa. P. aeruginosa N17-1 could degrade AFB₁, AFB₂ and AFM₁ by 82.8%, 46.8% and 31.9% after incubation in Nutrient Broth (NB) medium at 37 °C for 72 h, respectively. The culture supernatant of isolate N17-1 degraded AFB₁ effectively, whereas the viable cells and intra cell extracts were far less effective. Factors influencing AFB₁ degradation by the culture supernatant were investigated. Maximum degradation was observed at 55 °C. Ions Mn²⁺ and Cu²⁺ were activators for AFB₁ degradation, however, ions Mg²⁺, Li⁺, Zn²⁺, Se²⁺, Fe³⁺ were strong inhibitors. Treatments with proteinase K and proteinase K plus SDS significantly reduced the degradation activity of the culture supernatant. No degradation products were observed based on preliminary LC-QTOF/MS analysis, indicating AFB₁ was metabolized to degradation products with chemical properties different from that of AFB₁. The results indicated that the degradation of AFB₁ by P. aeruginosa N17-1 was enzymatic and could have a great potential in industrial applications. This is the first report indicating that the isolate of P. aeruginosa possesses the ability to degrade aflatoxin.     

Climatic, edaphic and altitudinal factors affecting yield and toxicity of Lathyrus sativus grown at five locations in Ethiopia

A 2 years (2005-2006) data analysis based on agronomic, qualitative, climatic and edaphic factors was carried out using 10 grass pea (Lathyrus sativus L.) genotypes grown at five eco-divergent locations (Alem Tena, Debre Zeit, Denbi, Akaki, Chefe Donsa) in Ethiopia. Crop yield showed considerable variability among locations, years and genotypes. Path coefficient analysis indicated that rainfall and days to maturity have a large positive influence on yield. High level of micronutrients Mn²⁺ and S²⁻ negatively affected yield. Path analysis revealed that Zn²⁺/P, days to maturity, yield and K⁺ were dominant variables affecting the response variable β-ODAP (β-N-oxalyl-L-α,β-diaminopropionic acid), the neuro-excitatory amino acid in grass pea seeds considered as the cause of neurolathyrism. Linear correlation analysis between β-ODAP and the 35 factors considered showed that β-ODAP level was positively correlated (r > 0.70) with K⁺ and sunshine hours (ssh) and negatively correlated (r < 0.70) with soil pH, days to maturity and yield. The strongest correlation of ssh with β-ODAP level was found during the phase of crop maturity. Our results suggest that β-ODAP biosynthesis and its response to environmental stress are maximized during the post-anthesis stage.     

Micromolar Zn2+ potentiates the cytotoxic action of submicromolar econazole in rat thymocytes: Possible disturbance of intracellular Ca2+ and Zn2+ homeostasis

Econazole, one of imidazole antifungals, has been reported to exhibit an inhibitory action on Mycobacterium tuberculosis and its multidrug-resistant strains under in vitro and ex vivo conditions. There is a chemotherapeutic potential of econazole against tuberculosis. We have revealed that Zn²⁺ at micromolar concentrations potentiates the cytotoxicity of imidazole antifungals by increasing membrane Zn²⁺ permeability. It is reminiscent of a possibility that econazole exhibits harmful action on human in the presence of Zn²⁺ at a physiological range when the agent is systemically administered. Because it is necessary to characterize the cytotoxic action of econazole in the presence of Zn²⁺, we have cytometrically examined the effects of econazole, ZnCl₂, and their combination on rat thymocytes. ZnCl₂ at concentrations ranging from 1 μM to 30 μM significantly increased the lethality induced by 10 μM econazole in a concentration-dependent manner. Econazole at a sublethal concentration of 1 μM significantly augmented the intensity of side scatter in the presence of micromolar ZnCl₂, suggesting the change in an intracellular circumstance by the combination of econazole and ZnCl₂. Econazole at 0.3 μM or more in the presence of ZnCl₂ increased the intensity of Fluo-3 fluorescence, an indicator for intracellular Ca²⁺. Furthermore, the intensity of FluoZin-3 fluorescence, an indicator for intracellular Zn²⁺, was also augmented by econazole at 0.1 μM or more in the presence of ZnCl₂. Results suggest that the combination of submicromolar econazole with micromolar ZnCl₂ may increase the intracellular concentration of Ca²⁺ and Zn²⁺, leading to disturbance of intracellular Ca²⁺ and Zn²⁺ homeostasis that triggers cytotoxic action.     

Studies with tryptophan metabolites in vitro. effect of zinc,manganese, copper and cobalt ions on kynurenine hydrolase and kynurenine aminotransferase in normal mouse liver

The possible mechanisms by which increasing concentrations of Zn²⁺, Mn²⁺, Cu²⁺, or Co²⁺ may affect the vitamin B₆-dependent enzymes kynurenine hydrolase and kynurenine aminotransferase. were studied in normal mouse liver homogenates. It was found that Zn²⁺ inhibited kynurenine hydrolase. whereas Mn²⁺ activated this enzyme, but both Zn²⁺ and Mn²⁺ activated kynurenine aminotransferase. Co²⁺ and Cu²⁺ inhibited both enzymes. This inhibition is attributed to the blocking and inactivation of the -SH groups of these enzymes and may be due to the adequate sequence of the -SH groups in both enzymes for Cu²⁺ or Co²⁺ action. This is in contrast to the inadequate sequence of these groups in kynurenine aminotransferase for Zn²⁺ action. The decreasing order by which these metal ions inhibit (a) kynurenine hydrolase is Cu²⁺ > Co²⁺ > Zn²⁺, and (b) kynurenine aminotransferase is Cu²⁺ > Co²⁺. The decreasing order of the per cent activation of the aminotransferase enzyme is Mn²⁺ > Zn²⁺. These decreasing orders fall into a more reasonable order approximating the order of complex stability of these metal ions.     

Effect of CH3HgCl and several transition metals on the dopamine neuronal carrier; peculiar behaviour of Zn2+

CH₃Hg⁺ and metal ions inhibited the specific binding of (1-[2-(diphenylmethoxy)ethyl]-4-(3-phenyl-2-[1-³H]propenyl) piperazine) ([³H]GBR 12783) to the dopamine neuronal carrier present in membranes from rat striatum with a general rank order of potency CH₃Hg⁺ > Cu²⁺ > Cd²⁺ > Zn²⁺ > Ni²⁺ = Mn²⁺ = Co²⁺, suggesting that -SH groups are chiefly involved in this inhibition. Five millimolar dithiothreitol reversed the rather stable block of the specific binding produced by Cd²⁺ or Zn²⁺. An increase in the concentration of Na⁺, or addition of either K⁺ or Ca²⁺ reduced the inhibitory effects of metal cations, except Cu²⁺. Zn²⁺ (3 μM) reduced the inhibitory potency of Cd²⁺ on the binding but was ineffective against CH₃Hg⁺ and Cu²⁺. Zn²⁺ at 0.3 to 10 μM significantly enhanced the specific binding of [³H]GBR 12783 and [³H]cocaine by 42 to 146%. Zn²⁺ (3 μM) increased the affinity of all pure uptake inhibitors tested and of the majority of the substrates for the [³H]GBR 12783 binding site. Dissociation experiments revealed that Zn²⁺ both inhibited and enhanced the [³H]GBR 12783 binding by recognizing amino acids located close to or in the radioligand binding site. Micromolar concentrations of Zn²⁺ noncompetitively blocked the [³H]dopamine uptake but they did not modify the block of the transport provoked by pure uptake inhibitors. These findings suggest that Na⁺, K⁺, Ca²⁺ and metal ions could recognize some -SH groups located in the [³H]GBR 12783 binding site; low concentrations of Zn²⁺ could allow a protection of these -SH groups.     

1,10-Phenanthroline stabilizes mRNA of the carcinogen-metabolizing enzyme,cytochrome P450 1a1

1,10-phenanthroline (phen), flufenamic acid, and indomethacin are inhibitors of aldo-keto reductases 1C1 (AKR1C1), but only phen decreased the benzo[a]pyrene (BaP)-induced cytochrome P450 1a1 (Cyp1a1) protein level. Therefore the decrease in the BaP-induced Cyp1a1 protein level was not due to inhibition of Akr1c1, but to phen itself. Phen decreased the BaP-induced Cyp1a1 promoter activity and protein expression, and in contrast, it increased Cyp1a1 mRNA, resulting from an increase in mRNA stability. Phen is also known as a transition metal ion-chelator. Along with the phen study, we also found that Zn²⁺, Fe²⁺ and Cu²⁺ increased Cyp1a1 mRNA and protein stability. Our results show that phen stabilized the mRNA of Cyp1a1, although it decreased cell viability. In addition, Zn²⁺ and Fe²⁺ highly neutralized phen's suppression of Cyp1a1 protein expression, but they only slightly neutralized phen's promotion of mRNA stability and suppression of cell viability, and had no effect on phen's suppression of promoter activity. Phen's effect on Cyp1a1 expression was reversible, which indicates that phen is non-covalently linked to its target. This report elucidates a new role for phen of stabilizing Cyp1a1 mRNA, and provides information for further studies on mRNA stabilization.     

Increase in intracellular Zn concentration by thimerosal in rat thymocytes: Intracellular Zn release induced by oxidative stress

Thimerosal (TMR), an ethylmercury-containing preservative in pharmaceutical products, was recently reported to increase intracellular Zn²⁺ concentration. Therefore, some health concerns about the toxicity of TMR remain because of physiological and pathological roles of Zn²⁺. To reveal the property of TMR-induced increase in intracellular Zn²⁺ concentration, the effect of TMR on FluoZin-3 fluorescence, an indicator of intracellular Zn²⁺, of rat thymocytes was examined. TMR at concentrations ranging from 0.3 μM to 10 μM increased the intensity of FluoZin-3 fluorescence in a concentration-dependent manner under external Ca²⁺- and Zn²⁺-free condition. The threshold concentration was 0.3–1 μM. The increase in the intensity was significant when TMR concentration was 1 μM or more. N,N,N′,N′-Tetrakis(2-pyridylmethyl)ethylenediamine (TPEN), a chelator for intracellular Zn²⁺, completely attenuated the TMR-induced augmentation of FluoZin-3 fluorescence. Hydrogen peroxide (H₂O₂) and N-ethylmaleimide, reducing cellular thiol content, significantly increased FluoZin-3 fluorescence intensity and decreased 5-chloromethylfluorescein (5-CMF) fluorescence intensity, an indicator for cellular thiol. The correlation coefficient between TMR-induced augmentation of FluoZin-3 fluorescence and attenuation of 5-CMF fluorescence was −0.882. TMR also attenuated the 5-CMF fluorescence in the presence of TPEN. Simultaneous application of H₂O₂ and TMR synergistically augmented the FluoZin-3 fluorescence. It is suggested that TMR increases intracellular Zn²⁺ concentration via decreasing cellular thiol content.     

Cd-induced apoptosis was mediated by the release of Ca from intracellular Ca storage

Previous studies found that cadmium (Cd) could induce apoptosis via interfering with the intracellular calcium (Ca) ions homeostasis. But the detailed mechanisms remain poorly understood. In the present study two cell lines (normal human liver cell HL-7702, and tumor cell Raji cell) were exposed to Cd along or co-incubated with ethylene glycol-bis (2-aminoethylether)-N,N,N′,N′-tetraacetic acid (EGTA) and 1,2-bis (2-aminophenoxy)ethane-N,N,N’,N’-tetraacetic acid-acetoxymethyl ester (BAPTA-AM), respectively. After the incubation, the apoptosis and intracellular Ca²⁺ ([Ca²⁺]_i) were measured. Excessive apoptosis was observed both in HL-7702 and Raji cells treated with Cd. Significant elevation of [Ca²⁺]_i was also detected in the cells with higher levels of apoptosis. EGTA (the extracellular Ca²⁺ chelator) decreased Cd-elicited [Ca²⁺]_i (22% in HL-7702 and 41% in Raji cells; p < 0.05) significantly except for apoptosis. However, BAMTA-AM (the [Ca²⁺]_i chelator) attenuated the Cd-elevated [Ca²⁺]_i (78% in HL-7702 and 59% in Raji cells; p < 0.05) and inhibited Cd-induced apoptosis significantly (p < 0.05). These results suggest that (1) Ca²⁺ was primarily generated intracellularly and only a small portion was generated extracellularly; (2) Cd-induced apoptosis was mediated by the release of Ca²⁺ from intracellular Ca storage but not an influx of extracellular Ca²⁺.     

Interaction of metal ions with tRNA: Effects of cadmium,zinc, and magnesium ions on chromatographic behavior of tyrosine and leucine tRNA species

The mechanism by which certain divalent cations may cause toxic effects was studied by examining the in vitro response of certain transfer RNAs to the cations. Two Drosophila tRNAs, those for tyrosine and leucine, were examined by reversed-phase column 5 (RPC-5) chromatography in the presence of each of three metal ions, Zn²⁺, Mg²⁺, and Cd²⁺. As Zn²⁺ concentration increased, both tRNAs eluted at progressively higher ionic strengths and the isoacceptor patterns collapsed and then reappeared. In contrast, when either the Mg²⁺ or the Cd²⁺ concentration increased, both tRNAs eluted at lower ionic strengths and the resolution of each of the isoacceptors was reduced. The resolution of tRNA^Tyr from tRNA^Leu was best with Cd²⁺ and poorest with Mg²⁺. If the observed changes were caused by conformational changes, they should be reversed upon removal of the metal ion; this was found to be so. Recovery of the tRNA after chromatography was reduced with Zn²⁺, presumably through nonspecific hydrolysis. One late-eluting leucyl-tRNA species was labile to incubation in the presence or absence of metal ion, followed by heating to 95°C. This same species was labile to Zn²⁺ exposure in the absence of heating. We suggest that reversed-phase column 5 chromatography is useful for evaluation of the effect of divalent metal ions on tRNA conformation and that the effects of various toxic metal ions could be examined in this manner.     

Allosteric modulation of 5-HT(1A) receptors by zinc: Binding studies

5-HT_1A receptors were studied via [³H]WAY-100635 and [³H]8-OH-DPAT binding to rat brain cortical membranes. We characterized the effect of zinc (Zn²⁺) on the binding properties of the 5-HT_1A receptor. The allosteric ternary complex model was applied to determine the dissociation constant (K_A) of Zn²⁺ and their cooperativity factors (α) affecting the dissociation constants (K_D, K_i) of [³H]WAY-100635, [³H]8-OH-DPAT, and serotonin (5-HT), the endogenous neurotransmitter. Zn²⁺ (5 μM-1 mM) inhibited the binding of agonist/antagonist to 5-HT1A receptors, mostly by decreasing both the ligands' affinity and the maximal number of sites. In [³⁵S]GTPγS binding assays Zn²⁺ behaved as insourmountable antagonist of 5-HT1A receptors, in agreement with radioligand binding assays. The residues involved in the formation of the inhibitory binding site on the 5-HT1A receptor were assessed by using N-ethyl-maleimide (NEM) or diethylpyrocarbonate (DEPC) which modify preferentially cysteine and histidine residues, respectively. Exposure to both agents did not block the negative allosteric effects of Zn²⁺ on agonist and antagonist binding. Our findings represent the first quantitative analysis of allosteric binding interactions for 5-HT_1A receptors. The physiological significance of Zn²⁺ modulation of 5-HT_1A receptors is unclear, but the colocalization of 5-HT_1A receptors and Zn²⁺ in the nervous system (e.g. in the hippocampus and cerebral cortex) suggests that Zn²⁺ released at nerve terminals may modulate signals generated by the 5-HT_1A receptors in vivo. Finally, these findings suggest that synaptic Zn²⁺ may be a factor influencing the effectiveness of therapies that rely on 5-HT_1A receptor activity.     

Studies on the reaction catalyzed by transport (Na, K) adenosine triphosphatase. I. Effects of divalent metals

The effects of divalent metals (Cu²⁺, Zn²⁺, Fe²⁺ and Pb²⁺) on a microsomal preparation of NaK-ATPase (ATP phosphohydrolase, EC 3.6.1.3) from beef cerebral cortex were studied. These metals are all potent inhibitors of the enzyme with I₅₀ values of 1 μM for Cu²⁺ and Zn²⁺, 3 μM for Fe²⁺ and 20 μM for Pb²⁺. Kinetic studies examining the effect of low concentrations of divalent metals on K_m and V for MgATP are reported. The results indicate that Fe²⁺ and Pb²⁺ are competitive inhibitors of NaK-ATPase with K_i values of 1.60 μM and 1.90 μM respectively. Cu²⁺ and Zn²⁺ are noncompetitive inhibitors of NaK-ATPase with K_i values of 1.18 μM and 3.48 μM respectively.     

Potentiation of iron-induced lipid peroxidation by a series of bipyridyls in relation to their ability to reduce iron

We determine the relative abilities of three bipyridyls (Paraquat P ⁺⁺, Benzylviologen B⁺⁺ and Diquat D⁺⁺) to stimulate iron-induced lipid peroxidation in relation to their ability to redox cycle and to reduce iron. The ability to redox cycle increases with the redox potential, in the order: P⁺⁺ < B⁺⁺ < D⁺⁺. The initial rates of Fe³⁺reduction increased from P⁺⁺ to D⁺⁺ and B⁺⁺. NADPH-dependent microsomal lipid peroxidation was measured in different conditions (1) increasing incubation time, 100 μM bipyridyl, 10 μM FeCl₃; (2) increasing concentrations of bipyridyls, 10 μM FeCl₃, 10 min incubation time; (3) increasing concentrations of FeCl₃, 100μM bipyridyl, 10 min incubation time. Bipyridyls potentiated the production of malondialdehyde (MDA), particularly with the shortest incubation times and the lowest concentrations of FeCl₃, in the order of activity P⁺⁺ < D⁺⁺ < B⁺⁺. The strong correlation between the initial rate of lipid peroxidation and Fe³⁺ reduction indicates that the rate of Fe³⁺ reduction determines the intensity of the potentiation effect of bipyridyls on lipid peroxidation—on the condition that the amount of peroxidizable substrate is not the limiting factor.     

Induction of frond abscission by metals and other toxic compounds in Lemna minor

Fronds of the duckweed Lemna minor L. clone St form colonies of different sizes on the basis of stipes connecting mother and daughter fronds for some time after the development of daughter fronds. All the metals (AsO₄³⁻, AsO₂⁻, Cd²⁺, CrO₄²⁻, Co²⁺, Cu²⁺, Ni²⁺, Hg²⁺, Tl⁺ and Zn²⁺) and one non-metal (SeO₄²⁻, SeO₃²⁻) tested here induced frond abscission, thus decreasing the colony size on the basis of a novel mechanism of abscission described recently. Concentration-response curves were created based on percentages of frond abscission after 7 and 24 h of toxic compound application, and response concentrations were calculated accordingly. The following conclusions could be drawn: (1) in most cases the response demonstrates less sensitivity than the bio test based on the ISO protocol 20079. (2) Even applying 1 mM of the metals, AsO₄³⁻, CrO₄²⁻, Co²⁺ and Zn²⁺ did not reach the half-maximal effects. (3) The concentration-response curves are bell-shaped with AsO²⁻, Cd²⁺, Hg²⁺, SeO₃²⁻ and Tl⁺, which demonstrates that abscission is induced by lower but not by higher concentrations. (4) Frond abscission shows fast and sensitive effects (24 h) for Ag⁺, Cu²⁺, AsO²⁻, SeO₄²⁻, SeO₃²⁻ and Tl⁺. The mechanisms and responses described here quantitatively for the first time complement and explain observations within the frame of the ISO protocol. Therefore, frond abscission should be regularly reported in the standard test protocols as abscission always indicates massive physiological effects.     

Effect of a new potent H2-receptor antagonist 3[[[2-[(diaminomethylene)amino]-4-thiazolyl]methyl]thio]-N2-sulfamoylpropionamidine (YM-11170) on gastric mucosal histamine-sensitive adenylate cyclase from guinea pig

The effect of 3[[[2-[(diaminomethylene)amino]-4-thiazolyl]methyl]thio]-N²-sulfamoylpro-pionamidine (YM-11170), a new thiazole H₂-receptor antagonist bearing propionamidine at the terminus of a side chain, on histamine-sensitive adenylate cyclase [ATP pyrophosphate-lyase (cyclizing); EC 4.6.1.1] of gastric mucosa from the guinea pig was studied and compared with that of cimetidine. YM-11170 displaced the concentration-stimulation curve of histamine-sensitive adenylate cyclase to the right with a pA₂ of 7.65 (K_i, = 2.25 × 10^?8M). Stimulation of gastric adenylate cyclase by 0.1 mM histamine was competitively inhibited by YM-11170 and cimetidine in a dose-dependent manner, with ic₅₀ values of 5.9 × 10^?7M and 1.4 × 10^?5M respectively. Hippocampal histamine-sensitive adenylate cyclase in the presence of 0.1 mM histamine was also competitively inhibited by YM-11170 with an ic₅₀ of 1.1 × 10^?7 M. YM-11170 did not affect Gpp(NH)p-, NaF-, PGE₂-stimulated or basal activity of the gastric adenylate cyclase. These data, together with other results, indicate that YM-11170 is a highly selective and potent H₂-receptor antagonist which competes with histamine at the receptor site on the histamine-sensitive adenylate cyclase.