Comparison of Usefulness of Four Chelating Agents (EDTA, NTA, ODA and IDA) for the Chromatographic Separation of Micro and Macro Amounts of Rare Earth Elements.

Literature on the use of four chelating agents namely: ethylenediaminetetraacetic acid, nitrilotriacetic acid, diglycolic acid and iminodiacetic acid for the chromatographic separation of micro and macro amounts of rare earth elements was critically reviewed and supplemented with some new unpublished data from our Laboratory. Advantages and disadvantages of ion exchange chromatography both in cation and anion mode as well as ion interaction chromatography techniques, which were used for rare earth elements separation, are discussed. The usefulness of some of the chromatographic systems for micro-macro separations was discussed and demonstrated. The importance of resilience of the separation method to column overloading in some analytical and larger scale separations was emphasized. The methods described in this article might suit well for recovering of individual lanthanides and yttrium from e-waste and other industrial wastes which were fast accumulating in recent years.

Biodegradable chelant-metal complexes enhance cadmium phytoextraction efficiency of Solanum americanum.

Toxic contaminants (metals and metal-containing compounds) are accumulating in the environment at an astonishing rate and jeopardize human health. Remarkable industrial revolution and the spectacular economic growth are the prime causes for the release of such toxic contaminants in the environment. Cadmium (Cd) is ranked the 7th most toxic compound by the Agency for Toxic Substances and Disease Registry (USA), owing to its high carcinogenicity and non-biodegradability even at miniscule concentration. The present study assessed the efficiency of four biodegradable chelants [nitrilotriacetic acid (NTA), ethylenediamine disuccinate (EDDS), ethylene glycol tetraacetic acid (EGTA), and citric acid (CA)] and their dose (5 mM and 10 mM) in enhancing metal accumulation in Solanum americanum Mill. (grown under 24 mg Cd kg-1 soil) through morpho-physiological and metal extraction parameters. Significant variations were observed for most of the studied parameters in response to chelants and their doses. However, ratio of root and shoot length, and plant height stress tolerance index differed non-significantly. The potential of chelants to enhance Cd removal efficiency was in the order - EGTA (7.44%) > EDDS (6.05%) > NTA (4.12%) > CA (2.75%). EGTA and EDDS exhibited dose-dependent behavior for Cd extraction with 10 mM dose being more efficient than 5 mM dose. Structural equation model (SEM) depicted strong positive interaction of metal extraction parameters with chelants (Z-value = 11.61, p = 0.001). This study provides insights into the importance of selecting appropriate dose of biodegradable chelants for Cd extraction, as high chelant concentration might also result in phytotoxicity. In the future, phytoextraction potential of these chelants needs to be examined through field studies under natural environmental conditions.

Fluorescent Single-Walled Carbon Nanotubes for Protein Detection.

Nanosensors have a central role in recent approaches to molecular recognition in applications like imaging, drug delivery systems, and phototherapy. Fluorescent nanoparticles are particularly attractive for such tasks owing to their emission signal that can serve as optical reporter for location or environmental properties. Single-walled carbon nanotubes (SWCNTs) fluoresce in the near-infrared part of the spectrum, where biological samples are relatively transparent, and they do not photobleach or blink. These unique optical properties and their biocompatibility make SWCNTs attractive for a variety of biomedical applications. Here, we review recent advancements in protein recognition using SWCNTs functionalized with either natural recognition moieties or synthetic heteropolymers. We emphasize the benefits of the versatile applicability of the SWCNT sensors in different systems ranging from single-molecule level to in-vivo sensing in whole animal models. Finally, we discuss challenges, opportunities, and future perspectives.

Isolation and characterization of iron chelators from turmeric (Curcuma longa): selective metal binding by curcuminoids.

Iron overload disorders may be treated by chelation therapy. This study describes a novel method for isolating iron chelators from complex mixtures including plant extracts. We demonstrate the one-step isolation of curcuminoids from turmeric, the medicinal food spice derived from Curcuma longa. The method uses iron-nitrilotriacetic acid (NTA)-agarose, to which curcumin binds rapidly, specifically, and reversibly. Curcumin, demethoxycurcumin, and bisdemethoxycurcumin each bound iron-NTA-agarose with comparable affinities and a stoichiometry near 1. Analyses of binding efficiencies and purity demonstrated that curcuminoids comprise the primary iron binding compounds recovered from a crude turmeric extract. Competition of curcuminoid binding to the iron resin was used to characterize the metal binding site on curcumin and to detect iron binding by added chelators. Curcumin-Iron-NTA-agarose binding was inhibited by other metals with relative potency: (>90% inhibition) Cu2+ ~ Al3+ > Zn2+ ≥ Ca2+ ~ Mg2+ ~ Mn2+ (<20% inhibition). Binding was also inhibited by pharmaceutical iron chelators (desferoxamine or EDTA) or by higher concentrations of weak iron chelators (citrate or silibinin). Investigation of the physiological effects of iron binding by curcumin revealed that curcumin uptake by cultured cells was reduced >80% by addition of iron to the media; uptake was completely restored by desferoxamine. Ranking of metals by relative potencies for blocking curcumin uptake agreed with their relative potencies in blocking curcumin binding to iron-NTA-agarose. We conclude that curcumin can selectively bind toxic metals including iron in a physiological setting, and propose inhibition of curcumin binding to iron-NTA-agarose for iron chelator screening.

New insights into the antioxidant activity and cytotoxicity of arzanol and effect of methylation on its biological properties.

The heterodimeric phloroglucinyl pyrone arzanol (Arz) has raised considerable interest because of its antiviral, anti-inflammatory, and antioxidant activity. We have investigated the effect of methylation of the pyrone moiety on the antioxidant activity and cytotoxicity of Arz. This manoeuvre, that left the polyphenolic moiety unscathed, was nevertheless detrimental for antioxidant activity in both the cholesterol thermal degradation- and the Cu2+-induced liposome oxidation assays, providing evidence of structure-activity relationships that go beyond the preservation of the polyphenolic pharmacophore. The antioxidant activity of Arz was retained also in the Fe-NTA model of in vivo oxidative stress, with protective effect on the oxidative degradation of plasmatic lipids, unsaturated fatty acids and cholesterol. Both Arz and methylarzanol (Me-Arz) were devoid of toxic effect on colonic differentiated Caco-2 cells up to 100µM, but significantly reduced cancer Caco-2 cell viability at lower dosages. Arz could also selectively reduce viability of other cancer cell lines, [murine melanoma cells (B16F10 cells), human cervical carcinoma cells (HeLa cells)], suggesting that it can act as a selective modulator of cell processes typical of cancer cells. Taken together, our results qualify Arz as a lead structure for further in vivo investigation of its pharmacological potential.

Immobilization of enzymes on magnetic beads through affinity interactions.

The development of enzyme immobilization techniques that will not affect catalytic activity and conformation is an important research task. Affinity tags that are present or added at a specific position far from the active site in the structure of the native enzyme could be used to create strong affinity bonds between the protein structure and a surface functionalized with the complementary affinity ligand. These immobilization techniques are based on affinity interactions between biotin and (strept)avidin molecules, lectins and sugars, or metal chelate and histidine tag. Recent developments involve immobilization of tagged enzymes onto magnetic nanoparticles. These supports can improve the performance of immobilized biomolecules in analytical assay because magnetic beads provide a relative large numbers of binding sites for biochemical reactions resulting in faster assay kinetics. This chapter describes immobilization procedures of tagged enzymes onto various magnetic beads.

Dissolution of synthetic uranium dibutyl phosphate deposits in oxidizing and reducing chemical formulations.

Permanganate and nitrilotriacetic acid (NTA) based dilute chemical formulations were evaluated for the dissolution of uranium dibutyl phosphate (U-DBP), a compound that deposits over the surfaces of nuclear reprocessing plants and waste storage tanks. A combination of an acidic, oxidizing treatment (nitric acid with permanganate) followed by reducing treatment (NTA based formulation) efficiently dissolved the U-DBP deposits. The dissolution isotherm of U-DBP in its as precipitated form followed a logarithmic fit. The same chemical treatment was also effective in dissolving U-DBP coated on the surface of 304-stainless steel, while resulting in minimal corrosion of the stainless steel substrate material. Investigation of uranium recovery from the resulting decontamination solutions by ion exchange with a bed of mixed anion and cation resins showed quantitative removal of uranium.

Optimization of chelators to enhance uranium uptake from tailings for phytoremediation.

A greenhouse experiment was set up to investigate the ability of citric acid (CA), oxalic acid (OA), nitrilotriacetic acid (NTA) and EDTA for phytoremediation of uranium tailings by Indian mustard [Brassica juncea (L.) Czern. et Coss]. Uranium tailings were collected from Umra mining region and mixed with 75% of garden soil which yielded a 25:75 mixture. Prepared pots were divided into four sets and treated with following different concentrations - 0.1, 0.5, 2.5 and 12.5 mmol kg(-1) soil additions for each of the four chelators. Control pots which were not treated with chelators. Experiments were conducted in completely randomized block design with triplicates. The optimum concentrations of these chelators were found on the basis of biomass production, tolerance and accumulation potential. The data collected were expressed statistically. EDTA produced maximum growth depression whereas, minimum occurred in the case of NTA. Maximum U uptake (3.5-fold) in the roots occurred at 2.5 mmol of CA, while NTA proved to be the weakest for the same purpose. Severe toxicity in the form of reduced growth and plant death was recorded at 12.5 mmol of each chelator. Minimum growth inhibition produced by chelators occurred in NTA which was followed by OA, moderate in CA and maximum was traced in EDTA applications. Chelator strengthened U uptake in the present study follows the order: CA>EDTA>OA>NTA.

Influence of chelating agents on biogenic uraninite reoxidation by Fe(III) (Hydr)oxides.

Microbially mediated reduction of soluble U(VI) to U(IV) with subsequent precipitation of uraninite, UO(2(S)), has been proposed as a method for limiting uranium (U) migration. However, microbially reduced UO(2) may be susceptible to reoxidation by environmental factors, with Fe(III) (hydr)oxides playing a significant role. Little is known about the role that organic compounds such as Fe(III) chelators play in the stability of reduced U. Here, we investigate the impact of citrate, DFB, EDTA, and NTA on biogenic UO(2) reoxidation with ferrihydrite, goethite, and hematite. Experiments were conducted in anaerobic batch systems in PIPES buffer (10 mM, pH 7) with bicarbonate for approximately 80 days. Results showed EDTA accelerated UO(2) reoxidation the most at an initial rate of 9.5 µM day(-1) with ferrihydrite, 8.6 µM day(-1) with goethite, and 8.8 µM day(-1) with hematite. NTA accelerated UO(2) reoxidation with ferrihydrite at a rate of 4.8 µM day(-1); rates were less with goethite and hematite (0.66 and 0.71 µM day(-1), respectively). Citrate increased UO(2) reoxidation with ferrihydrite at a rate of 1.8 µM day(-1), but did not increase the extent of reaction with goethite or hematite, with no reoxidation in this case. In all cases, bicarbonate increased the rate and extent of UO(2) reoxidation with ferrihydrite in the presence and absence of chelators. The highest rate of UO(2) reoxidation occurred when the chelator promoted both UO(2) and Fe(III) (hydr)oxide dissolution as demonstrated with EDTA. When UO(2) dissolution did not occur, UO(2) reoxidation likely proceeded through an aqueous Fe(III) intermediate with lower reoxidation rates observed. Reaction modeling suggests that strong Fe(II) chelators promote reoxidation whereas strong Fe(III) chelators impede it. These results indicate that chelators found in U contaminated sites may play a significant role in mobilizing U, potentially affecting bioremediation efforts.

Allosteric effects of sulfonate anions on the rates of iron release from serum transferrin.

Serum transferrin is the protein that transports ferric ion through the bloodstream and is thus a potential target for iron chelation therapy. However, the release of iron from transferrin to low-molecular-weight chelating agents is usually quite slow. Thus a better understanding of the mechanism for iron release is important to assist in the design of more effective agents for iron removal. This paper describes the effect of sulfonate anions on the rates of iron removal from C-terminal monoferric transferrin by acetohydroxamic acid, deferiprone, nitrilotriacetic acid (NTA), and diethylenetriaminepentaacetic acid at 25°C in 0.1M N-(2-hydroxyethyl)piperazine-N'-(2-ethanesulfonic acid) (Hepes) buffer at pH 7.4. These ligands remove iron via a combination of pathways that show saturation and first order dependence on the ligand concentration. The kinetic effects of the anions methanesulfonate, methylenedisulfonate, and ethylenedisulfonate were evaluated. All these anions increase the overall rates of iron release, presumably by binding to an allosteric anion binding site on the protein. The two disulfonates produce a larger acceleration in iron release than the monosulfonate. More detailed studies using methylenedisulfonate show that this anion accelerates the rate of iron release via the saturation pathway. The addition of methylenedisulfonate results in the appearance of a large saturation pathway for iron release by NTA, which otherwise removes iron by a simple first-order process. The sulfonate group was selected for these studies because it represents an anionic functional group that can be covalently linked to a therapeutic ligand to accelerate iron release in vivo. The current studies indicate that the binding of the sulfonates to the allosteric site on the protein is quite weak, so that one would not expect a significant acceleration in iron release at clinically relevant ligand concentrations.

Facile fabrication of recyclable and active nanobiocatalyst: purification and immobilization of enzyme in one pot with Ni-NTA functionalized magnetic nanoparticle.

Ni-NTA functionalized iron oxide magnetic nanoparticle was synthesized and used to selectively immobilize a his-tagged enzyme from cell free extract as an active and recyclable nanobiocatalyst, where purification and immobilization of the target enzyme were accomplished in one pot.

Label-free femtomolar detection of target DNA by impedimetric DNA sensor based on poly(pyrrole-nitrilotriacetic acid) film.

An ultrahigh performance impedimetric DNA sensor is presented showing detection limits in the femtomolar range. This electrochemical setup was constructed initially by electrogeneration of poly(11-pyrrol-1-yl-undecanoic acid N(alpha'),N(alpha)-bis(carboxymethyl)-L-lysine amide) (poly(pyrrole-NTA)) film. The latter was then modified by the coordination of Cu(2+) ions onto the chelating NTA centers followed by the immobilization of the ssHIV-DNA previously modified by a polyhistidine tag by affinity binding. The immobilization of the DNA probe and hybridization with the complementary target ssHIV-DNA were investigated using fluorescence microscopy and quantified with quartz crystal microbalance experiments leading to DNA probe and duplex coverage of 1.7 x 10(-11) and 7.7 x 10(-12) mol cm(-2), respectively. The duplex formation was corroborated by amperometric measurements through the duplex labeling by a glucose oxidase. In the presence of hydroquinone as redox indicator, the DNA sensor was applied to the impedimetric detection of target DNA without a labeling step. A linear quantification of the HIV DNA target was carried out in the range 10(-15) to 10(-8) mol L(-1).

Quantification of trace elements in natural samples by electrospray ionization mass spectrometry with a size-exclusion column based on the formation of metal-aminopolycarboxylate complexes.

Metal ions were determined by ESI-MS in the negative ion mode as monovalent negative ions of their aminopolycarboxylic acid (APC) complexes, e.g., [Al(cydta)](-), [Pb(Hcydta)](-), where excess amounts of the APC agents were added to sample solutions. Among several APCs studied, we chose trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid (CyDTA) as the best chelating agent because of higher stabilities and higher sensitivities of the complexes. The ionization efficiency of these metal complexes was strongly affected by the presence of matrix salts, e.g., NaCl, KNO(3), and etc. Thus, a size exclusion column (Sephadex G-10) was used for the online separation of the metal-APC complexes from other matrix salts. This method was successfully applied to the quantitative analyses for total amounts of Al, Ni, Cu, Zn, and Pb in the biological certified reference materials, olive leaves (BCR-062) and plankton (BCR-414). The detection limits of the present methods for these elements were several to several ten nanomolar levels. Moreover, this approach was extended to determine ultratraces of fluoride based on the formation of the ternary complex of aluminum, fluoride and nitrilotriacetic acid (NTA), i.e., [AlF(nta)](-). Its detection limit was 10 nM and was 2 orders of magnitude better than that of a fluoride ion selective electrode method. This method was applied to determine fluoride in tap water, river water, and green tea samples.

Use of proliferation tests to evaluate the effects of complexing agents on beryllium toxicity.

Occupational exposure to beryllium may cause chronic beryllium disease (CBD), a granulomatous interstitial pneumonitis caused by a cell-mediated immune response with delayed hypersensitivity initiated by an electrostatic interaction with the MHC class II human leukocyte antigen (HLA). Increased research efforts focus on the development of a CBD treatment by chelation therapy. This work presents an in vitro evaluation of the beneficial effects of beryllium chelation with different organic substrates. We have used a standard beryllium lymphocyte proliferation test (BeLPT) adapted for mouse splenocytes. Three complexing agents, 4,5-dihydroxy-1,3-benzenedisulfonic acid (tiron), nitrilotripropionic acid (NTP) and nitrilotriacetic acid (NTA), were tested using different protocols of the splenocyte proliferation test (SPT). We studied their corrective effect (beryllium pre-exposed splenocytes), their protective effect (ligand pre-exposed splenocytes) and their combined effects at fixed Be:L ratio of 1:2, at fixed Be concentration and at fixed L concentration. We also studied the effect of tiron in preventing splenocyte sensitization to beryllium. All three complexing agents showed a corrective effect and proved efficient in the combined effects, except NTA in the fixed Be:L ratio. Only NTP and tiron showed a significant protection at lower beryllium concentrations, while NTA was not significant. Splenocytes pre-exposed to chelated beryllium did not show sensitization while splenocytes pre-exposed to beryllium were sensitized. We observed a strong correlation between the efficiency of the complexing agent and its affinity towards beryllium. Both tiron and NTP showed a similar affinity towards the beryllium ion that is 10(7) higher than that of NTA.

2-Aminobenzothiazole degradation by free and Ca-alginate immobilized cells of Rhodococcus rhodochrous.

2-Aminobenzothiazole (ABT) degradation was investigated using free and immobilized systems during photodegradation under solar light in the presence of Fe(III)-nitrilotriacetic acid (FeNTA), biodegradation by Rhodococcus rhodochrous, and during combined conditions. Ca-alginate hydrogel was chosen as a model matrix and some complementary studies were required to characterize this new system. R. rhodochrous metabolism in this type of environment was monitored by NMR spectroscopy. Neither change in intracellular pH values nor in ATP concentrations was observed by in vivo(31)P NMR, showing that no metabolic modification occurred between free and immobilized cells. (1)H NMR demonstrated that alginate was not used as carbon source by R. rhodochrous. After establishing the pre-treatment protocol by SPE to eliminate solubilised alginate, ABT adsorption on beads and degradation were studied. The same pathways of transformation were observed in suspended and immobilized cell systems. Considering the ABT adsorption phenomenon on alginate beads (8%), the efficiency of the two systems was found to be comparable although the degradation rate was slightly lower with immobilized cells. The most important result was the finding that the positive effect of FeNTA on ABT degradation with immobilized cells was similar to that observed previously with free cells. All these results show that mechanisms observed with free cells can be extrapolated to entrapped cells, i.e. under conditions much closer to those usually encountered in the environment.

Enhanced phytoextraction of uranium and selected heavy metals by Indian mustard and ryegrass using biodegradable soil amendments.

The applicability of biodegradable amendments in phytoremediation to increase the uptake of uranium (U), cadmium (Cd), chromium (Cr), copper (Cu), lead (Pb) and zinc (Zn) by Indian mustard (Brassica juncea) and ryegrass (Lolium perenne) was tested in a greenhouse experiment. Plants were cultivated during one month on two soils with naturally or industrially increased contaminant levels of U. Treatments with citric acid, NH4-citrate/citric acid, oxalic acid, S,S-ethylenediamine disuccinic acid (EDDS) or nitrilotriacetic acid (NTA) at a rate of 5 mmol kg(-1) dry soil caused increases in soil solution concentrations that were up to 18 times higher for U and up to 1570 times higher for other heavy metals, compared to the controls. Shoot concentrations increased to a much smaller extent. With EDDS, 19-, 34-, and 37-fold increases were achieved in shoots of Indian mustard for U, Pb and Cu, respectively. The increases in plant uptake of Cd, Cr and Zn were limited to a factor of four at most. Ryegrass generally extracted less U and metals than Indian mustard. Despite a marked increase of U and metal concentrations in shoots after addition of amendments, the estimated time required to obtain an acceptable reduction in soil contaminant concentrations was impractically long. Only for Cu and Zn in one of the studied soils, could the Flemish standards for clean soil theoretically be attained in less than 100 years.

[Construction expression and biologic activity of recombinant human sCR1 eucaryotic cell].

AIM: To express human soluble complement receptor type 1(sCR1)protein using ferment cell secreting type carrier and study the extraorgan biologic activity of recombinant human sCR1 fusion protein. METHODS: Total human RNA was extracted from peripheral blood. The full length cDNA of human sCR1 gene was obtained by RT-PCR and them, cloned into Pichia pastoris eukaryotic expression vector pPIC9k to construct the recombinant plasmid pPIC9k-sCR1 containing human sCR1.After identified by DNA sequencing, the recombinant plasmid pPIC9k-sCR1 was transformed into Pichia pastoris SMD1168. The ferment cell line of the recombinant sCR1 which was chosen by G418 resistance was identified by PCR, After methanol induction, the expressed protein products were verified by SDS-PAGE and Western blot, purified by Ni(2+)-NTA agarose affinity chromatography, and its biologic activity was identified. RESULTS: The obtained Pichia pastoris secretion type yeast carrier pPIC9k-sCR1 was chosen by G418 and identified by PCR to get a highly copied and integral recombinant ferment cell line. The recombinant human sCR1 fusion protein was expressed by yeast cells containing pPIC9k-sCR1 induced by methanol. It was a protein band about M(r) 31 000 in gel, which could be identified by CD35 of anti-sCR1 protein monoclonal antibody with Western blotting technique. The highly purified sCR1 fusion protein and its biologic activity were detected obtained by Ni(2+)-NTA agarose affinity chromatography. CONCLUSION: The recombinant human sCR1 fusion protein can be highly expressed in the Pichia pastoris expression system, which resembles the human natural protein's antigenicity and biologic activity.

Protective effect of Rumex patientia (English Spinach) roots on ferric nitrilotriacetate (Fe-NTA) induced hepatic oxidative stress and tumor promotion response.

In this communication, we document the antioxidant potential of ethanolic extract of Rumex patientia L. (Polygonaceae) roots and its chemopreventive effects against Fe-NTA mediated hepatic oxidative stress, hepatotoxicity and tumor promotion response. The extract exhibited high polyphenolic content, potent reducing power and significantly scavenged free radicals (including several reactive oxygen species (ROS) and reactive nitrogen species (RNS)). The extract also significantly and dose dependently protected against oxidative damage to lipids and DNA. These results indicated R. patientia root extract to exert a potent antioxidant activity in vitro. The efficacy of extract was also evaluated in vivo and it was found to exert a potent protective affect in acute oxidative tissue injury animal model: ferric nitrilotriacetate (Fe-NTA) induced hepatotoxicity in mice. Administration of Fe-NTA (9 mg/kg body weight, i.p.) to mice led to a significant oxidative stress and allied damage in liver tissues and induced hyperproliferation. A significant depletion was observed in GSH content and enzymes implicated in its metabolism. Attenuation also occurred in activities of other hepatic antioxidant enzymes including SOD, CAT, and GPX. Fe-NTA also incited hyperproliferation response elevating ornithine decarboxylase activity and [(3)H]-thymidine incorporation into DNA. Histopathological investigations and liver function tests (LFT) indicated Fe-NTA to cause extensive hepatic damage. However, prophylactic treatment with R. patientia root extract at a dose regimen of 100-200mg/kg body weight for a week not only restored hepatic antioxidant armory close to normal, but also significantly precluded oxidative damage restoring normal hepatic architecture and levels of hepatic damage markers. The data obtained in the present study illustrates R. patientia roots to possess potent antioxidant and free radical scavenging activities and thwart oxidative damage and hyperproliferation in hepatic tissues.

Degradation of 2-fluorophenol by the brown-rot fungus Gloeophyllum striatum: evidence for the involvement of extracellular Fenton chemistry.

Iron-containing liquid cultures of the brown-rot basidiomycete Gloeophyllum striatum degraded 2-fluorophenol. Two simultaneously appearing degradation products, 3-fluorocatechol and catechol, were identified by gas chromatography and mass spectrometry (GC-MS). Concomitantly, fluoride was produced at approximately 50% of the amount that theoretically could be achieved upon complete dehalogenation. Defluorination was strongly inhibited in the presence of either the hydroxyl radical scavenger mannitol or superoxide dismutase, as well as in the absence of iron. The addition of the natural iron chelator oxalate caused a clear but less extensive inhibition, whereas supplementation with the artificial iron chelator nitrilotriacetic acid increased fluoride production. Extracellular 2-fluorophenol degradation was evidenced by defluorination, observed upon addition of 2-fluorophenol to cell-free culture supernatants derived from iron-containing fungal cultures. Ultrafiltered culture supernatants oxidized methanol to formaldehyde, known as a product of the reaction of methanol with hydroxyl radical. In addition, G. striatum was found to produce metabolites extractable with ethyl acetate that are capable of reducing Fe3+. GC-MS analysis of such extracts revealed the presence of several compounds. The mass spectrum of a prominent peak matched those previously reported for 2,5-dimethoxyhydroquinone and 4,5-dimethoxycatechol, fungal metabolites implicated to drive hydroxyl radical production in Gloeophyllum. Taken together, these findings further support an extracellular Fenton-type mechanism operative during halophenol degradation by G. striatum.

Carbonic anhydrase inhibitors. A general approach for the preparation of water-soluble sulfonamides incorporating polyamino-polycarboxylate tails and of their metal complexes possessing long-lasting, topical intraocular pressure-lowering properties.

Reaction of polyamino-polycarboxylic acids or their dianhydrides with aromatic/heterocyclic sulfonamides possessing a free amino/imino/hydrazino/hydroxy group afforded mono- and bis-sulfonamides containing polyamino-polycarboxylic acid moieties in their molecule. The acids/anhydrides used in synthesis included IDA, NTA, EDDA, EDTA and EDTA dianhydride, DTPA and DTPA dianhydride, EGTA and EGTA dianhydride, and EDDHA, among others. All the newly prepared derivatives showed strong affinity toward isozymes I, II, and IV of carbonic anhydrase (CA). Metal complexes of the new compounds have also been prepared. Metal ions used in such preparations included di- and trivalent main-group and transition cations, such as Zn(II), Cu(II), Al(III), etc. Some of the new sulfonamides/disulfonamides obtained in this way, as well as their metal complexes, behaved as nanomolar CA inhibitors against isozymes II and IV, being slightly less effective in inhibiting isozyme I. Some of these sulfonamides as well as their metal complexes strongly lowered intraocular pressure (IOP) when applied topically, directly into the normotensive/glaucomatous rabbit eye, as 1-2% water solutions/suspensions. The good water solubility of these sulfonamide CA inhibitors, correlated with the neutral pH of their water solutions used in the ophthalmologic applications and the long duration of action of the IOP-lowering effect, makes them interesting candidates for developing novel types of antiglaucoma drugs devoid of serious topical side effects.