Mechanism of mutagenicity by 5-hydroperoxymethyl-2'-deoxyuridine, an intermediate product of ionizing radiation, in bacteria. HPMdU bacterial mutagenicity and oxidation of DNA bases.

The specific objective was to find what processes are responsible for the mutagenicity of 5-hydroperoxymethyl-2'-deoxyuridine (HPMdU), which is a product of ionizing radiation, and what role transition metal ions play in those processes. We found that HPMdU is a more potent mutagen than its decomposition products 5-hydroxymethyl-2'-deoxyuridine (HMdU) and 5-formyl-2'-deoxyuridine (FdU) in the Salmonella typhimurium strains tested, with the TA100 strain being the most sensitive. HMdU exerted intermediate mutagenicity and FdU was the weakest of the three compounds. At 50 nmoles/plate, HPMdU increased the number of revertants by 4-fold, whereas 1000 nmoles HMdU was required to enhance the number of revertants by 5-fold. Pretreatment of TA100 with o-phenanthroline, a membrane-permeable Fe and Cu chelator, caused an increase in mutagenicity of the low HPMdU doses but inhibited that of the 50 nmoles HPMdU/plate, while desferal, a membrane-impermeable Fe chelator, had virtually no effect. Azide (a catalase inhibitor) enhanced HPMdU mutagenicity, whereas 3-amino-1,2,4-triazole (a catalase and peroxidase inhibitor) and ammonium formate (a hydroxyl radical scavenger) were protective. Preincubation of TA100 cells with 20 and 40 nM HPMdU caused dose-dependent formation of the oxidized DNA base derivatives HMdU, thymidine glycol and 8-hydroxyl-2'-deoxyguanosine (8-OHdG), known hydroxyl radical-mediated oxidation products. Cumulatively, these results suggest that the genetic effects of HPMdU are due to its hydroperoxide moiety, which upon reacting with Fe generates hydroxyl radicals that in turn oxidize neighboring bases in cellular DNA. This also may be a mechanism by which ionizing radiation exerts its long-term effects.     

Decomposition of nucleoside hydroperoxide by metals and metalloproteins

5-Hydroperoxymethyl-2′-deoxyuridine (HPMdU) is formed in DNA by ionizing radiation. Although relatively stable, HPMdU eventually decomposes to two products 5-hydroxymethyl-2′-deoxyuridine (HMdU) and 5-formyl-2′-deoxyuridine (FdU). We show that a number of transition metal ions and metalloproteins accelerate this process. Of the metal ions tested, Sn(II) and Fe(II) were the most active, with the former producing exclusively HMdU, and the latter, a mixture of both. Cu(I), Cu(II), Co(II), and Ni(II) induced a predominant generation of FdU, with copper ions being more effective than Co and Ni. FdU was also preferentially formed in the presence of the iron-containing proteins transferrin and ferritin, whereas HMdU was the major product in the presence of apotransferrin as well as in the presence of ceruloplasmin, a copper-containing protein.     

Superoxide and hydrogen peroxide suppression by metal ions and their EDTA complexes

Redox-active metal ions such as Fe(II)\(III) and Cu(I)\(II) have been proposed to activate reactive oxygen and nitrogen species (RONS) and thus, perpetuate oxidative damage. Here, we show that concentrations of metal ions and EDTA complexes with superoxide-destroying activities equivalent to 1 U SOD are Fe(III) 5.1 microM, Mn(II) 0.77 microM, Cu(II)-EDTA 3.55 microM, Fe(III)-EDTA 2.34 microM, and Mn(II)-EDTA 1.38 microM. The most active being the aquated Cu(II) species which exhibited superoxide-destroying activity equivalent to 2U of SOD at 0.29 microM. Hydrogen peroxide-destroying activities were as follows Fe(III)-EDTA ca. 70 U/mg and aquated Fe(III) 141 U/mg. In contrast, DTPA prevented superoxide-destroying activity and significantly depleted hydrogen peroxide-destroying activity. In conclusion, non-protein bound transition metal ions may have significant anti-oxidant effects in biological systems. Caution should be employed in bioassays when chelating metal ions. Our results demonstrate that DTPA is preferential to EDTA for inactivating redox-active metal ions in bioassays.     

Interactions of quercetin with iron and copper ions: complexation and autoxidation

Quercetin (3,3',4',5,7-pentahydroxyflavone), one of the most abundant dietary flavonoids, has been investigated for its ability to bind Fe(II), Fe(III), Cu(I) and Cu(II) in acidic to neutral solutions. In particular, analysis by UV-visible spectroscopy allows to determine the rate constants for the formation of the 1:1 complexes. In absence of added metal ion, quercetin undergoes a slow autoxidation in neutral solution with production of low hydrogen peroxide (H(2)O(2)) concentrations. Autoxidation is accelerated by addition of the metal ions according to: Cu(I) > Cu(II)>Fe(II) Fe(III). In fact, the iron-quercetin complexes seem less prone to autoxidation than free quercetin in agreement with the observation that EDTA addition, while totally preventing iron-quercetin binding, slightly accelerates quercetin autoxidation. By contrast, the copper-quercetin complexes appear as reactive intermediates in the copper-initiated autoxidation of quercetin. In presence of the iron ions, only low concentrations of H(2)O(2) can be detected. By contrast, in the presence of the copper ions, H(2)O(2) is rapidly accumulated. Whereas Fe(II) is rapidly autoxidized to Fe(III) in the presence or absence of quercetin, Cu(I) bound to quercetin or its oxidation products does not undergo significant autoxidation. In addition, Cu(II) is rapidly reduced by quercetin. By HPLC-MS analysis, the main autoxidation products of quercetin are shown to be the solvent adducts on the p-quinonemethide intermediate formed upon two-electron oxidation of quercetin. Finally, in strongly acidic conditions (pH 1-2), neither autoxidation nor metal complexation is observed but Fe(III) appears to be reactive enough to quickly oxidize quercetin (without dioxygen consumption). Up to ca. 7 Fe(III) ions can be reduced per quercetin molecule, which points to an extensive oxidative degradation.     

Quantitative high-performance liquid chromatography analysis of DNA oxidized in vitro and in vivo

Oxidative modification of genetic material has been implicated as a factor in carcinogenesis, particularly during promotion and progression, and therefore there is a need for sensitive detection of oxidized DNA bases. We developed a method that can be applied to DNA isolated from any source and used to simultaneously quantify oxidized nucleosides without a need to prelabel the DNA or use destructive hydrolytic procedures. This method is based on: (a) enzymatic DNA digestion; (b) HPLC separation of the resultant nucleosides; (c) acetylation of the oxidized nucleosides with [3H]Ac2O (acetic anhydride); (d) removal of the radioactive debris; and (e) quantitative analysis of tritiated nucleoside acetates by HPLC. Enzymatic DNA digestion was optimized using DNase I in the presence of Mg2+ (pH 7), followed by nuclease P1 in the presence of Zn2+ (pH 5.1) and alkaline phosphatase (pH 7.5). Analysis of DNA oxidized with H2O2 in the presence of Fe2+/EDTA for 30 min showed that the levels of 8-OHdG (8-hydroxy-2'-deoxyguanosine) were increased 2.7-fold, HMdU (5-hydroxymethyl-2'-deoxyuridine) 3.15-fold, and FdU (5-formyl-2'-deoxyuridine) 2.5-fold. Although the (-)-isomer of cis-dTG (cis-thymidine glycol) was enhanced 2.3 times, the (+)-isomer remained virtually unchanged. Analysis of DNA isolated from epidermal cells of mice treated in vivo with the tumor promoter TPA (12-O-tetradecanoylphorbol 13-acetate) showed 4.8-, 2.7-, and 8.7-fold increases in the levels of total cis-dTG, 8-OHdG, and HMdU, respectively, and of some unknown DNA oxidation products. These results prove applicability of the 3H-postlabeling method to the analysis of DNA (and potentially RNA) isolated from many sources, including animals and humans.     

Metal-mediated oxidative damage to cellular and isolated DNA by gallic acid, a metabolite of antioxidant propyl gallate

Propyl gallate (PG), widely used as an antioxidant in foods, is carcinogenic to mice and rats. PG increased the amount of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), a characteristic oxidative DNA lesion, in human leukemia cell line HL-60, but not in HP100, which is hydrogen peroxide (H2O2)-resistant cell line derived from HL-60. Although PG induced no or little damage to 32P-5'-end-labeled DNA fragments obtained from genes that are relevant to human cancer, DNA damage was observed with treatment of esterase. HPLC analysis of the products generated from PG incubated with esterase revealed that PG converted into gallic acid (GA). GA induced DNA damage in a dose-dependent manner in the presence of Fe(III)EDTA or Cu(II). In the presence of Fe(III) complex such as Fe(III)EDTA or Fe(III)ADP, GA caused DNA damage at every nucleotide. Fe(III) complex-mediated DNA damage by GA was inhibited by free hydroxy radical (*OH) scavengers, catalase and an iron chelating agent. These results suggested that the Fe(III) complex-mediated DNA damage caused by GA is mainly due to *OH generated via the Fenton reaction. In the presence of Cu(II), DNA damage induced by GA occurred at thymine and cytosine. Although *OH scavengers did not prevent the DNA damage, methional inhibited the DNA damage. Cu(II)-mediated DNA damage was inhibited by catalase and a Cu(I) chelator. These results indicated that reactive oxygen species formed by the interaction of Cu(I) and H2O2 participates in the DNA damage. GA increased 8-oxodG content in calf thymus DNA in the presence of Cu(II), Fe(III)EDTA or Fe(III)ADP. This study suggested that metal-mediated DNA damage caused by GA plays an important role in the carcinogenicity of PG.     

Chemical cleavage of plasmid DNA by Cu(II), Ni(II) and Co(III) desferal complexes.

It is demonstrated that the Cu(II), Co(III) and Ni(II) complexes of a siderophore chelating drug desferal cleave DNA, in contrast to the corresponding Fe(II) complex which does not bring about DNA scission. Hydroxy radical scavengers inhibit the cleavage reaction.     

Tannic acid inhibits in vitro iron-dependent free radical formation

The antioxidant activity of tannic acid (TA), a plant polyphenol claimed to possess antimutagenic and anticarcinogenic activities, was studied by monitoring (i) 2-deoxyribose degradation (a technique for OH detection), (ii) ascorbate oxidation, (iii) ascorbate radical formation (determined by EPR analysis) and (iv) oxygen uptake induced by the system, which comprised Fe(III) complexes (EDTA, nitrilotriacetic acid (NTA) or citrate as co-chelators), ascorbate and oxygen. TA removes Fe(III) from the co-chelators (in the case of EDTA, this removal is slower than with NTA or citrate), forming an iron-TA complex less capable of oxidizing ascorbate into ascorbate radical or mediating 2-deoxyribose degradation. The effectiveness of TA against 2-deoxyribose degradation, ascorbate oxidation and ascorbate radical formation was substantially higher in the presence of iron-NTA (or iron-citrate) than with iron-EDTA, which is consistent with the known formation constants of the iron complexes with the co-chelators. Oxygen uptake and 2-deoxyribose degradation induced by Fe(II) autoxidation were also inhibited by TA. These results indicate that TA inhibits OH formation induced by Fe(III)/ascorbate/O(2) mainly by arresting Fe(III)-induced ascorbate oxidation and Fe(II) autoxidation (which generates Fe(II) and H(2)O(2), respectively), thus limiting the production of Fenton reagents and OH formation. We also hypothesize that the Fe(II) complex with TA exhibits an OH trapping activity, which explains the effect of TA on the Fenton reaction.     

A comparison of cobalt(II) and iron(II) hydroxyl and superoxide free radical formation

We have employed the electron spin resonance spin-trapping technique to study the reaction of Co(II) with hydrogen peroxide in a chemical system and in a microsomal system. In both cases, we employed the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) and were able to detect the formation of DMPO/.OH and DMPO/.OOH. DMPO/.OOH was the predominant radical adduct formed in the chemical system, while the two adducts were of similar concentrations in the microsomal system. The formation of both of these adducts in either reaction system was inhibited by the addition of superoxide dismutase or catalase, and by chelating the cobalt with either ethylenediaminetetraacetic acid (EDTA) or diethylenetriaminepentaacetic acid (DTPA). The incorporation of the hydroxyl radical scavengers ethanol, formate, benzoate, or mannitol inhibited the formation of DMPO/.OH in both systems. We also repeated the study using Fe(II) in place of Co(II). In contrast to the Co(II) results, Fe(II) reacted with hydrogen peroxide to yield only DMPO/.OH, and this adduct formation was relatively insensitive to the presence of added superoxide dismutase. In addition, Fe(II)-mediated DMPO/.OH formation increased when the iron was chelated to either EDTA or DTPA rather than being inhibited as for Co(II). Thus, we propose that Co(II) does not react with hydrogen peroxide by the classical Fenton reaction at physiological pH values.     

Mechanism of metal-mediated DNA damage and apoptosis induced by 6-hydroxydopamine in neuroblastoma SH-SY5Y cells

6-Hydroxydopamine (6-OHDA) is a neurotoxin to produce an animal model of Parkinson's disease. 6-OHDA increased the formation of 8-oxo-7, 8-dihydro-2'-deoxyguanosine (8-oxodG), a biomarker of oxidatively damaged DNA, and induced apoptosis in human neuroblastoma SH-SY5Y cells. Iron or copper chelators inhibited 6-OHDA-induced 8-oxodG formation and apoptosis. Thus, iron and copper are involved in the intracellular oxidatively generated damage to DNA, a stimulus for initiating apoptosis. This study examined DNA damage caused by 6-OHDA plus metal ions using (32)P-5'-end-labelled DNA fragments. 6-OHDA increased levels of oxidatively damaged DNA in the presence of Fe(III)EDTA or Cu(II). Cu(II)-mediated DNA damage was stronger than Fe(III)-mediated DNA damage. The spectrophotometric detection of p-quinone and the scopoletin method showed that Cu(II) more effectively accelerated the 6-OHDA auto-oxidation and H(2)O(2) generation than Fe(III)EDTA. This study suggests that copper, as well as iron, may play an important role in 6-OHDA-induced neuronal cell death.     

DNA methylation by tert-butyl hydroperoxide-iron (II): a role for the transition metal ion in the production of DNA base adducts.

Metabolic degradation of both endogenous and exogenous peroxides is associated with the etiology of several diseases including cancer. Tert-butyl hydroperoxide (TBHP) has been widely employed as a model compound to study the cytotoxicity and promoting effects of organic peroxides. Recently, we reported that incubations of TBHP with iron (II) and calf thymus DNA led to generation of high yields of methyl radicals and to DNA methylation. Interestingly, DNA was methylated to products expected from both free radical and ionic mechanisms such as 8-methylguanine (C8-MeGua) and 7-methylguanine (N7-MeGua), respectively. To elucidate the mechanisms by which methyl radicals can produce different types of DNA adducts, we examined the effects of transition metal ions (iron (II), iron (III) and copper (I)) and metal ion chelators (ethylenediamine-N,N,N",N"-tetraacetate (EDTA) and desferal) on the nature and the yields of the DNA adducts produced during TBHP decomposition. The results led us to propose that a direct methyl radical attack on DNA guanine residues produces C8-MeGua whereas N7-MeGua and 3-methyladenine (N3-MeAde) are likely to be produced by attack of nucleophilic DNA centers on methyl radical generated in situ by the assistance of transition metal ions bound to DNA.     

Polyphenol tannic acid inhibits hydroxyl radical formation from Fenton reaction by complexing ferrous ions

Tannic acid (TA), a plant polyphenol, has been described as having antimutagenic, anticarcinogenic and antioxidant activities. Since it is a potent chelator of iron ions, we decided to examine if the antioxidant activity of TA is related to its ability to chelate iron ions. The degradation of 2-deoxyribose induced by 6 microM Fe(II) plus 100 microM H2O2 was inhibited by TA, with an I50 value of 13 microM. Tannic acid was over three orders of magnitude more efficient in protecting against 2-deoxyribose degradation than classical *OH scavengers. The antioxidant potency of TA was inversely proportional to Fe(II) concentration, demonstrating a competition between H2O2 and AT for reaction with Fe(II). On the other hand, the efficiency of TA was nearly unchanged with increasing concentrations of the *OH detector molecule, 2-deoxyribose. These results indicate that the antioxidant activity of TA is mainly due to iron chelation rather than *OH scavenging. TA also inhibited 2-deoxyribose degradation mediated by Fe(III)-EDTA (iron = 50 microM) plus ascorbate. The protective action of TA was significantly higher with 50 microM EDTA than with 500 microM EDTA, suggesting that TA removes Fe(III) from EDTA and forms a complex with iron that cannot induce *OH formation. We also provided evidence that TA forms a stable complex with Fe(II), since excess ferrozine (14 mM) recovered 95-96% of the Fe(II) from 10 microM TA even after a 30-min exposure to 100-500 microM H2O2. Addition of Fe(III) to samples containing TA caused the formation of Fe(II)n-TA, complexes, as determined by ferrozine assays, indicating that TA is also capable of reducing Fe(III) ions. We propose that when Fe(II) is complexed to TA, it is unable to participate in Fenton reactions and mediate *OH formation. The antimutagenic and anticarcinogenic activity of TA, described elsewhere, may be explained (at least in part) by its capacity to prevent Fenton reactions.     

Simultaneous determination of Fe(III) and Fe(II) in water solutions and tissue homogenates using desferal and 1,10-phenanthroline

At neutral pH values 1,10-phenanthroline forms a colored complex with Fe(II), but it does not form such a complex with Fe(III). On the contrary, only Fe(III) forms with desferal a yellow complex with a g = 4.3 electron paramagnetic resonance (EPR) signal, but Fe(II) is rapidly oxidized by desferal to Fe(III), which gives then a yellow complex. On the basis of these facts, a method for simultaneous determination of both Fe(II) and Fe(III) ions was elaborated using a desferal-phenanthroline mixture. Two ways of detecting Fe(II) and Fe(III) have been suggested: (1) the spectrophotometric method for transparent water solutions, and (2) the EPR-spectrometric method for turbid solutions and tissue homogenates. The latter method was applied for determination of free and weakly bound iron in rat liver. The Fe(II) amount in intact liver was 22.2 ± 7.6 nmol/g of wet tissue; free Fe(III) was not found.     

Copper-dependent cleavage of DNA by bleomycin

DNA strand scission by bleomycin in the presence of Cu and Fe was further characterized. It was found that DNA degradation occurred readily upon admixture of Cu(I) or Cu(II) + dithiothreitol + bleomycin, but only where the order of addition precluded initial formation of Cu(II)--bleomycin or where sufficient time was permitted for reduction of the formed Cu(II)--bleomycin to Cu(I)--bleomycin. DNA strand scission mediated by Cu + dithiothreitol + bleomycin was inhibited by the copper-selective agent bathocuproine when the experiment was carried out under conditions consistent with Cu chelation by bathocuproine on the time scale of the experiment. Remarkably, it was found that the extent of DNA degradation obtained with bleomycin in the presence of Fe and Cu was greater than that obtained with either metal ion alone. A comparison of the sequence selectivity of bleomycin in the presence of Cu and Fe using 32P-end-labeled DNA duplexes as substrates revealed significant differences in sites of DNA cleavage and in the extent of cleavage at sites shared in common. For deglycoblemycin and decarbamoylbleomycin, whose metal ligation is believed to differ from that of bleomycin itself, it was found that the relative extents of DNA cleavage in the presence of Cu were not in the same order as those obtained in the presence of Fe. The bleomycin-mediated oxygenation products derived from cis-stilbene were found to differ in type and amount in the presence of added Cu vs. added Fe. Interestingly, while product formation from cis-stilbene was decreased when excess Fe was added to a reaction mixture containing 1:1 Fe(III) and bleomycin, the extent of product formation was enhanced almost 4-fold in reactions that contained 5:1, as compared to 1:1, Cu and bleomycin. The results of these experiments are entirely consistent with the work of Sugiura [Sugiura, Y. (1979) Biochem. Biophys. Res. Commun. 90, 375-383], who first demonstrated the generation of reactive oxygen species upon admixture of O2 and Cu(I)--bleomycin.     

The reaction between ferrous polyaminocarboxylate complexes and hydrogen peroxide: an investigation of the reaction intermediates by stopped flow spectrophotometry

The reactions of Fe(II)EDTA, Fe(II)DTPA, and Fe(II)HEDTA with hydrogen peroxide near neutral pH have been investigated. All these reactions have been assumed to proceed through an active intermediate, I1, (Formula: see text) where pac is one of the three polyaminocarboxylates mentioned above. I1, whether .OH radical or an iron complex, reacts with ethanol, formate, and other scavengers at rates relative to k2 that, with the exception of t-butanol and benzoate, are similar, but not identical, to those expected for the.OH radical. In contrast, at pH 3, in the absence of ligands the reaction of I1 with Fe2+ was inhibited by ethanol and t-butanol and the reactivity of I1 towards these two scavengers relative to ferrous ion is identical to that exhibited by the hydroxyl radical. When pac = HEDTA, the intermediate of the first reaction reacts with formate ion to form the ferrous HEDTA ligand radical complex, which is characterized by absorption maxima at 295 nm (epsilon = 2,640 M-1 cm-1) and 420 nm (epsilon = 620 M-1 cm-1). For the reaction of Fe(II)HEDTA with H2O2, the following mechanism is proposed: (Formula: see text) where k17 = 4.2 X 10(4) M-1 sec-1 and k19 = 5 +/- 0.2 sec-1.     

Mechanism of metal-mediated DNA damage and apoptosis induced by 6-hydroxydopamine in neuroblastoma SH-SY5Y cells

6-Hydroxydopamine (6-OHDA) is a neurotoxin to produce an animal model of Parkinson's disease. 6-OHDA increased the formation of 8-oxo-7, 8-dihydro-2′-deoxyguanosine (8-oxodG), a biomarker of oxidatively damaged DNA, and induced apoptosis in human neuroblastoma SH-SY5Y cells. Iron or copper chelators inhibited 6-OHDA-induced 8-oxodG formation and apoptosis. Thus, iron and copper are involved in the intracellular oxidatively generated damage to DNA, a stimulus for initiating apoptosis. This study examined DNA damage caused by 6-OHDA plus metal ions using ³²P-5′-end-labelled DNA fragments. 6-OHDA increased levels of oxidatively damaged DNA in the presence of Fe(III)EDTA or Cu(II). Cu(II)-mediated DNA damage was stronger than Fe(III)-mediated DNA damage. The spectrophotometric detection of p-quinone and the scopoletin method showed that Cu(II) more effectively accelerated the 6-OHDA auto-oxidation and H₂O₂ generation than Fe(III)EDTA. This study suggests that copper, as well as iron, may play an important role in 6-OHDA-induced neuronal cell death.     

Pyridoxal isonicotinoyl hydrazone inhibits iron-induced ascorbate oxidation and ascorbyl radical formation

Previous work from our laboratory demonstrated that pyridoxal isonicotinoyl hydrazone (PIH) has in vitro antioxidant activity against iron plus ascorbate-induced 2-deoxyribose degradation due to its ability to chelate iron; the resulting Fe(III)-PIH(2) complex is supposedly unable to catalyze oxyradical formation. A putative step in the antioxidant action of PIH is the inhibition of Fe(III)-mediated ascorbate oxidation, which yields the Fenton reagent Fe(II) [Biochim. Biophys. Acta 1523 (2000) 154]. In this work, we demonstrate that PIH inhibits Fe(III)-EDTA-mediated ascorbate oxidation (measured at 265 nm) and the formation of ascorbyl radical (in electron paramagnetic resonance (EPR) studies). The efficiency of PIH against ascorbate oxidation, ascorbyl radical formation and 2-deoxyribose degradation was dose dependent and directly proportional to the period of preincubation of PIH with Fe(III)-EDTA. The efficiency of PIH in inhibiting ascorbate oxidation and ascorbyl radical formation was also inversely proportional to the Fe(III)-EDTA concentration in the media. When EDTA was replaced by the weaker iron ligand nitrilotriacetic acid (NTA), PIH was much more effective in preventing ascorbate oxidation, ascorbyl radical formation and 2-deoxyribose degradation. Moreover, the replacement of EDTA with citrate, a physiological chelator with a low affinity for iron, also resulted in PIH having a higher efficiency in inhibiting iron-mediated ascorbate oxidation and 2-deoxyribose degradation. These results demonstrate that PIH removes iron from EDTA (or from either NTA or citrate), forming an iron-PIH complex that cannot induce ascorbate oxidation effectively, thus inhibiting iron-mediated oxyradical formation. These results are of pharmacological relevance because PIH has been considered for experimental chelating therapy in iron-overload diseases.     

Comparison of the toxicity of heavy metals to cabbage growth

Summary Cabbage plants were grown for 55 days with a nutrient solution containing 1 and 10 ppm of V, Cr(III), Cr(VI), Mn, Fe, Co, Ni, Cu, Zn, Cd, Hg(I), orHg(II). A comparison of the plant growth and chemical analysis revealed that Cr(VI), Cu, Cd, and Hg(II) in the solution are most toxic to the plant growth (hence detrimental to the cabbage-head formation) and Mn, Fe, and Zn are less toxic than other heavy metals, and that Mn, Zn, Co, Ni, and Cd and translocated into all the plant organs while V, Cr(III), Cr(VI), Fe, Cu, Hg(I), and Hg(II) are accumulated in the roots.     

Determination of photostability and photodegradation products of moxifloxacin in the presence of metal ions in solutions and solid phase. Kinetics and identification of photoproducts

Photostability of moxifloxacin (MOXI) after UVA irradiation in solutions and solid phase, with and without participation of Cu(II), Zn(II), Al(III), and Fe(III) was tested. The studies were carried out by the TLC-densitometric method and LC-MS/MS method. Elaborated and validated chromatography-densitometric method was used for assaying. It was shown that the number and type of photoproducts depend on the environment and type of the metal ion. The studied ions enhanced the degradation of MOXI in solutions, and the influence of Cu(II) and Fe(III) ions was higher than that of Zn(II) and Al(III) ions. In solid phase, in contrast to solutions, all metal ions decreased the photodegradation, however the influence of ions, Al(III) and Zn(II), was weaker than that of Cu(II) and Fe(III) ions. Identification of the degradation products performed with LC-MS/MS and (1)H NMR identified them as: 1-cyclopropyl-6-fluoro-7-amino-8-methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic acid, 1-cyclopropyl-6-fluoro-8-methoxy-4-oxo-7-(2-oxo-octahydro-6H-pyrrolo[3,4-b]pyridine-6-yl)-1,4-dihydroquinoline-3-carboxylic acid, 7-[3-hydroxyamino-4-(2-carboxyethyl)pyrrolidin-1-yl]-1-cyclopropyl-6-fluoro-8-methoxy-4-oxo-1,4-dihydroquinoline-3-carboxylic acid.     

Reduction of Fe(III), Mn(III), and Cu(II) chelates by roots of pea (Pisum sativum L.) or soybean (Glycine max)

Neither the reduction of Fe(III) to Fe(II) by roots nor its induction by Fe-deficiency are unique characteristics of the reductive activities of roots. We show that chelated Mn(III) or chelated Cu(II), as well as chelated Fe(III), may be reduced by Fe-stressed roots of pea (Pisum sativum L.). Deficiency of Fe stimulated the reduction of Fe(III)EDTA about 20-fold, the reduction of Mn(III)CDTA about 11-fold, the reduction of Cu(II)(BPDS)₂ about 5-fold, and the reduction of Fe(III)(CN)₆ by only about 50%. Not only are metals other than Fe reduced as part of the Fe-stress response, but deficiencies of metals other than Fe stimulate the reductive activity of roots. We show that depriving peas or soybeans (Glycine max) of Cu or Zn stimulates the reduction of Fe(III).