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Ligands containing the 1-methyl-3-hydroxy-2(1H)-pyridinone group (Me-3,2-HOPO) are powerful plutonium(IV) sequestering agents. The Ce(IV) complexes of bidentate and tetradentate HOPO ligands have been quantitatively studied as models for this sequestration. The complexes Ce(L1)4, Ce(L2)4, Ce(L3)2, and Ce(L4)2 (L1 = Me-3,2-HOPO; L2 = PR-Me-3,2-HOPO; L3 = 5LI-Me-3,2-HOPO; L4 = 5LIO-Me-3,2-HOPO) were prepared in THF solution from Ce(acac)4 and the corresponding ligand. The complex Ce(L4)2 was also prepared in aqueous solution by air oxidation of the Ce(III) complex [Ce(L4)2]-. Single-crystal X-ray diffraction analyses are reported for Ce(L1)(4)x2CHCl3 [P1 (no. 2), Z = 2, a = 9.2604(2) A, b = 12.1992(2) A, c = 15.9400(2) A, alpha = 73.732(1) degrees, beta = 85.041(1) degrees, gamma = 74.454(1) degrees], Ce(L3)2x2CH3OH [P2(1)/c (no. 14), Z = 4, a = 11.7002(2) A, b = 23.0033(4) A, c = 15.7155(2) A, beta = 96.149(1) degrees], Ce(L4)(2).2CH3OH [P1 (no. 2), Z = 2, a = 11.4347(2) A, b = 13.8008(2) A, c = 15.2844(3) A, alpha = 101.554(1) degrees, beta = 105.691(1) degrees, gamma = 106.746(1) degrees], and Ce(L4)2x4H2O [P2(1)/c (no. 14), Z = 4, a = 11.8782(1) A, b = 22.6860(3) A, c = 15.2638(1) A, beta = 96.956(1) degrees]. A new criterion, the shape measure S, has been introduced to describe and compare the geometry of such complexes. It is defined as [formula: see text], where m is the number of edges, delta i is the observed dihedral angle along the ith edge of delta (angle between normals of adjacent faces), theta i is the same angle of the corresponding ideal polytopal shape theta, and min is the minimum of all possible values. For these complexes the shape measure shows that the coordination geometry is strongly influenced by small changes in the ligand backbone or solvent. Solution thermodynamic studies determined overall formation constants (log beta) for Ce(L2)4, Ce(L3)2, and Ce(L4)2 of 40.9, 41.9, and 41.6, respectively. A thermodynamic cycle has been used to calculate these values from the corresponding formation constants of Ce(III) complexes and standard electrode potentials. From the formation constants and from the protonation constants of the ligands, extraordinarily high pM values for Ce(IV) are generated by these tetradentate ligands (37.5 for Ce(L3)2 and 37.0 for Ce(L4)2). The corresponding constants for Pu(IV) are expected to be substantially the same.

The synthesis, characterization, and metal-binding studies of chelate-functionalized dendrimers is reported. Salicylate, catecholate, and hydroxypyridinonate bidentate chelators have been coupled to the surface of both poly(propyleneimine) (Astramol) and poly(amidoamine) (Starburst, PAMAM) dendrimers up to the fifth generation (64 endgroups). A general method has been developed for the facile and high quality chromatographic purification of poly(propyleneimine) and poly(amidoamine) dendrimer derivatives. One- and two-dimensional (TOCSY) 1H NMR experiments and electrospray ionization mass spectrometry (ESI-MS) have confirmed the exhaustive coupling of these chelators to the primary amine functionalities of the dendrimers. Spectrophotometric titrations were used to investigate the metal binding ability of these macrochelates. Spectral analysis shows that ferric iron binding to these ligands is localized to the chelating endgroups. The ability of these dendritic polymers to bind large numbers of metal ions may lead to applications as metal sequestering agents for waste remediation technologies.

New ligands based on hydroxypyridinonate (HOPO) and other bidentate ligands are explored as iron(III) sequestering agents. These are based on the N,N',N"-tris[(3-hydroxy-1-methyl- 2-oxo-1,2-didehydropyrid-4-yl)-carboxamidoethyl]amine (TREN-Me-3,2-HOPO) platform in which one Me-3,2-HOPO ligand group is substituted with either a 2-hydroxyisophthalamide (TREN-Me-3,2-HOPOIAM) or a 2,3-dihydroxyterephthalamide (TREN-Me-3,2-HOPOTAM) moiety. The ferric complexes have been prepared and structurally characterized by X-ray diffraction: Fe[TREN-Me-3,2-HOPOIAM] crystallizes in the monoclinic space group C2/c with cell parameters a = 18.1186(3) A, b = 17.5926(2) A, c = 25.0476(2) A, beta = 98.142(1) degrees, Z = 8. Fe[TREN-Me-3,2-HOPOTAM]- crystallizes in the monoclinic space group C2/c with cell parameters a = 31.7556(12) A, b = 14.0087(6) A, c = 22.1557(9) A, beta = 127.919(1) degrees, Z = 8. The aqueous coordination chemistry of these ligands with both the ferric and ferrous redox states of iron has been examined using spectroscopic and electrochemical methods, giving log formation constants of 26.89(3) (beta 110), 31.16(6) (beta 111) for the ferric TREN-Me-3,2-HOPOIAM complexes and 33.89(2) (beta 110), 38.45(2) (beta 111) for the ferric TREN-Me-3,2-HOPOTAM complexes. For the reduced (ferrous) complexes values of 10.03(9) (beta 110) and 13.7(2) (beta 110) were observed for the Fe[TREN-Me-3,2-HOPOIAM]- and Fe[TREN-Me-3,2-HOPOTAM]2- complexes, respectively. These data provide a complete description of metal-ligand speciation as a function of pH and of redox activity. The ligands described in this work are part of a new class of heteropodate ligands which exploit the various chelating properties of several binding units within a single tripodal ligand and allow for systematic variation of the properties for medical or other applications.

While iron release from enterobactin-mediated iron transport occurs primarily via an esterase that destroys the siderophore, other catechol siderophores that are not susceptible to hydrolysis act as bacterial growth factors. Elucidating the structures of protonated ferric enterobactin may reveal the pathway by which synthetic analogues fulfill bacterial iron requirements. In order to more completely model this potential delivery pathway for ferric iron, as well as to understand the pH dependent structural dynamics of ferric enterobactin, two ligands, (2-hydroxybenzoyl-2-aminoethyl)-bis(2,3-dihydroxybenzoyl-2-aminoethyl)amine (TRENCAMSAM) and (2-hydroxy-3-methoxybenzoyl-2-aminoethyl)-bis(2,3-dihydroxybenzoyl-2- aminoethyl)amine (TRENCAM(3M)SAM), have been synthesized as models for monoprotonated enterobactin. The coordination chemistry of these ligands with Fe3+ and Al3+ has been investigated. Fe[TRENCAMSAM]2- crystallizes in the triclinic space group P1: Z = 1, a = 11.3307(6) A, b = 12.5479(7) A, c = 15.5153(8) A, alpha = 94.513(1) degree, beta = 105.867(1) degree, gamma = 94.332(1) degree. The structure is a two-metal two-ligand dimer supported by mu-oxo bridges from two catecholate moieties. Al[TRENCAMSAM]2- crystallizes in the triclinic space group P1: Z = 2, a = 9.1404(2) A, b = 13.3570(1) A, c = 15.5950(1) A, alpha = 95.711(1) degree, beta = 104.760(1) degree, gamma = 92.603(1) degree. The complex is a monomer with a five-coordinate, square-pyramidal aluminum cation. Al[TRENCAM(3M)SAM]2- crystallizes in the monoclinic space group C2/m: Z = 8, a = 34.244(2) A, b = 11.6206(6) A, c = 21.9890(12) A, beta = 101.478(1) degree. The complex is also a monomer, but with a highly distorted five-coordinate, square-pyramidal aluminum cation coordination sphere. At high pH these complexes do not display a salicylate mode of binding; however, at low pH Al[TRENCAMSAM]2- converts to protonated Al[H3TRENCAMSAM]+, which is a six-coordinate, tris-salicylate complex. Al[H3TRENCAMSAM]+ crystallizes in the triclinic space group P1: Z = 2, a = 11.5475(4) A, b = 12.1681(4) A, c = 12.5094(4) A, alpha = 109.142(1) degree, beta = 104.327(1) degree, gamma = 103.636(1) degree. This is the first catecholamide enterobactin analogue that has been structurally characterized in both a catecholate and salicylate mode of coordination.

Bidentate hydroxypyridinone chelators effectively complex and facilitate excretion of trivalent iron. To test the hypothesis that hexadentate chelators are more effective than bidentate chelators at low concentrations, urinary and biliary Fe excretions were determined in Fe-loaded rats before and after administration of a bidentate chelator, Pr-(Me-3,2-HOPO), or its hexadentate analogue, TREN-(Me-3,2-HOPO). The bidentate chelator slightly increased biliary Fe excretion in Fe-loaded rats after IV (90 micromol/kg) and PO (90 or 270 micromol/kg) administration, but chelation efficiency did not exceed 1%. The hexadentate chelator markedly increased biliary Fe excretion, achieving overall chelation efficiencies of 14% after IV administration of 30 micromol/kg and 8 or 3% after PO (30 or 90 micromol/kg) administration. The hexadentate chelator was significantly more effective than the bidentate chelator after IV injection and oral dosing. In chelator-treated Fe-loaded or saline-injected rats,90% of the excreted Fe was in the bile. Oral TREN-(Me-3,2-HOPO), given to non-Fe-loaded rats, did not appreciably change Fe output, indicating that there was little Fe depletion in the absence of Fe overload. These results support the hypothesis that greater Fe chelation efficiency can be achieved with hexadentate than with bidentate chelators at lower, and presumably safer, concentrations. The results also demonstrate that TREN-(Me-3, 2-HOPO) is a promising, orally effective, Fe chelator.

The effectiveness of the siderophore analogues DFO-HOPO (a hydroxypyridone derivative of desferrioxamine) and 3,4,3-LIHOPO (a linear tetrahydroxypyridinone) for the decorporation of 238Pu and 241Am from rat was studied. (1) Dosage-effect relationship. A similar treatment effect on Pu was achieved by single s.c. injection of 30 mumol kg-1 or by oral administration of 100 mumol kg-1 of either of the two ligands, provided the oral dose was administered earlier. In general, LIHOPO was more effective than DFO-HOPO: retention of Pu in the liver and bones was reduced by LIHOPO to10% of control values. No increase in renal retention of the actinides was observed. Whilst DFO-HOPO did not affect Am retention, a substantial reduction was achieved by LIHOPO. Removal effectiveness for injected LIHOPO on Pu was higher than that on Am, especially in the bones and after low ligand doses. Orally administered small doses of LIHOPO, however, mobilized more Am than Pu, both from the liver and the bone. (2) Time-effect relationship. The effectiveness of the injected ligands for Pu decreased exponentially with the time between exposure and treatment. With DFO-HOPO, the calculated half-times for decrease of mobilized fractions of Pu from the bone and liver were 5 and 12 h respectively. The effect of LIHOPO on Pu decreased much more slowly, with a half-time of 3-4 weeks. For instance, a single injection of 30 mumol kg-1 LIHOPO at 10 days post-Pu removed 30 and 50% activity from the bone and liver respectively. The removal effect of LIHOPO for Am in the liver decreased with time in the same way as for Pu but the mobilized fractions of skeletal and renal Am decreased from the first day with a half-time of only 8 and 4 days respectively.

The yeast Rhodotorula pilimanae produces the dihydroxamate siderophore rhodotorulic acid (RA) in prodigious amounts when starved for iron. Synthetic dihydroxamate analogs of RA have been prepared in which the diketopiperazine ring of RA is replaced by a simple chain of n methylene groups. It is found that R. pilimanae is able to accumulate iron using these achiral complexes, as well as from simple monohydroxamate analogs, at rates comparable to those of RA. While the Fe2RA3 complex does not enter the cell, there is a receptor system whose geometric requirements for siderophore recognition have been probed using analogs. In contrast to mono- or dihydroxamate ligands, the trihydroxamate siderophores such as ferrioxamine B are completely ineffective at delivering iron to R. pilimanae. This is ascribed to the greater stability of these complexes, which blocks release of the Fe(III) in a ligand exchange process that is required for uptake. To explore whether this ligand exchange involves redox catalysis, Ga(III) was substituted for Fe(III). The gallium was taken up at rates near those of iron and were also energy-dependent, as determined by metabolic inhibition with KCN.

Although the ferrioxamines are an important and well-characterized class of siderophores produced by several species of Nocardia, Streptomyces, Micromonospora, Arthrobacter, Chromobacterium, and Pseudomonas, no studies of the mechanism of ferrioxamine-mediated iron uptake have been performed for an organism which produces the siderophore. This is the first report of metal transport in Streptomyces pilosus mediated by the native ferrioxamines B, D1, D2, and E. 55Fe accumulation in these ferrioxamines was dependent on metabolic energy and was a saturable process with increasing complex concentration. The apparent Km for [55Fe]ferrioxamine B uptake was approximately 0.2 microM. Both chromic desferriferrioxamine B and [67Ga]desferriFerrioxamine B were transported at rates similar to those of the 55Fe complexes: this implies that no decomplexation or reduction of the metal ion is required for transport, since the chromic complexes are kinetically inert and the gallium complexes have no stable divalent state as a possible reduction product. In addition, isomers of inert chromic desferriferrioxamine B complexes were used to probe the stereospecificity of the ferrioxamine uptake system. The chromic complexes were separated into three fractions by cationic exchange chromatography and assigned as two cis and a (mixture of) trans geometrical isomer(s) by their visible spectra. [55Fe]ferrioxamine B uptake was equally inhibited by each isomer, suggesting that no differentiation between cis and trans geometrical isomers occurs. In the presence of chromic desferriferrioxamine B isomers, the uptake rates for 55Fe-labeled ferrioxamines E, D1, and D2 were even more strongly reduced than was that for [55Fe]ferrioxamine B itself. From these results we conclude that all the ferrioxamines tested are transported into the cells by the same uptake system.

Synthetic analogs of enterochelin (enterobactin) were tested for their ability to support the growth of Escherichia coli K-12 under iron-limiting conditions. The cyclic compound MECAM [1,3,5-N.N'; N"-tris-(2,3-dihydroxybenzoyl)-triamino-methylbenzene] and its N-methyl derivative Me3MECAM promoted growth, whereas the 2,3-dihydroxy-5-sulfonyl derivatives MECAMS and Me3MECAMS were inactive. The same results were obtained with TRIMCAM [1,3,5-tris(2,3-dihydroxybenzoylcarbamido)-benzene] and TRIMCAMS (the 2,3-dihydroxy-5-sulfonyl derivative of TRIMCAM). However, the sulfonic acid-containing linear compound LICAMS [1,5,10-N,N', N"-tris(5-sulfo-2,3-dihydroxybenzoyl)-triaza-decane] supported growth. In contrast, LIMCAMC, in which the sulfonyl groups at the five position of LICAMS are replaced by carboxyl groups at the four position, was inactive. The uptake of the active analogs required the functions specified by the fepB, fesB, and tonB genes. Surprisingly, growth promotion of mutants lacking the enterochelin receptor protein in the outer membrane was observed. Only MECAM protected cells against colicin B (which kills cells after entering at the enterochelin uptake sites) and transported Fe3+ at about half the enterochelin rate.

Rhodotorulic acid (RA), a dihydroxamate siderophore produced by Rhodotorula pilimanae, forms 3:2 complexes with ferric and chromic ions (M2RA3) at pH 7. Kinetically inert chromic complexes of RA have been separated into geometrical isomers and for the first time partially resolved into optical isomers. The three isomers delta-cis, delta-trans, and lambda-trans were characterized by their visible and circular dichroism spectra. Inhibition by both delta-isomers of radiolabeled ferric RA uptake in R. pilimanae was equally effective. However the lambda-cis isomer was significantly less effective as an inhibitor. Concentration-dependent uptake kinetics were performed with ferric RA and the ferric complex of synthetic enantio-RA, which form predominantly delta and lambda complexes, respectively. The lambda-enantio-Fe2RA3 was 50% less effective in supplying iron to R. pilimanae than was Fe2RA3. An additional synthetic analog of RA, which lacks a carbonyl group at the diketopiperazine ring, exhibited the same uptake rates as ferric RA. We conclude that stereoselective recognition of optical isomers takes place during iron uptake mediated by RA and that this recognition primarily involves the right-handed delta coordination "propellor" of the metal center and its adjacent functionalities.

The effect of imposing constraints on the parameters of a least-squares refinement is considered, and a general equation is presented that relates the derivatives for the calculated, unconstrained parameters to those of the constrained parameters.

Macromolecules containing four sulfonated catecholy (2,3-dihydroxybenzoyl) groups are effective for decorporation of newly acquired Pu(IV). However, multiple injections in mice and single injections in dogs of 30 mumole/kg of 3,4,3-LICAM(S), the most effective sulfonated poly(catechoylamide) ligand, indicated that it would be toxic, so the ligand structure was modified. Each ligand was injected into mice (30 mumole/kg, intraperitoneally) 1 hr after an intravenous injection of 238Pu(IV) citrate, and mice were killed 24 hr after the Pu injection. Excreta and tissues were analyzed for Pu. (a) The number of catechoyl groups per molecule was reduced to suppress affinity for Fe(III). Net excretion (treated - control) of 55% of the injected Pu was promoted by tetrameric 3,4,3-LICAM(S), 51% by trimeric 3,4-LICAM(S), 22% by dimeric 2-LICAM(S), and 7.4% by the monomer, Tiron. (b) A mesitylene platform was substituted for the linear backbone. Net Pu excretion promoted by MECAM(S), a structurally less flexible trimer, was only 26%, and excretion was delayed. (c) A carboxyl substituent on the catechoyl groups reduced the acidity and hydrophilicity of the ligands. Tetrameric 3,4,3-LICAM(C) promoted 63% net Pu excretion, and one-third of that was fecal. The Pu contents of liver and skeleton were 33 and 44% of their respective 1-hr control values--compared to 51 and 44%, respectively, for CaNa3-DTPA. Mice given 30 mumole/kg of 3,4,3-LICAM(C) 20 times in 4 weeks showed no ill effects. (d) Large N-terminal alkane substituents added to 3,4,3-LICAM(C) increased ligand lipophilicity, hindered Pu chelation, and delayed excretion.

Rhodotorulic acid (RA), a dihydroxamate siderophore produced by Rhodotorula pilimanae, forms 3:2 complexes with ferric and chromic ions (M2RA3) at pH 7. Kinetically inert chromic complexes of RA have been separated into geometrical isomers and for the first time partially resolved into optical isomers. The three isomers delta-cis, delta-trans, and lambda-trans were characterized by their visible and circular dichroism spectra. Inhibition by both delta-isomers of radiolabeled ferric RA uptake in R. pilimanae was equally effective. However the lambda-cis isomer was significantly less effective as an inhibitor. Concentration-dependent uptake kinetics were performed with ferric RA and the ferric complex of synthetic enantio-RA, which form predominantly delta and lambda complexes, respectively. The lambda-enantio-Fe2RA3 was 50% less effective in supplying iron to R. pilimanae than was Fe2RA3. An additional synthetic analog of RA, which lacks a carbonyl group at the diketopiperazine ring, exhibited the same uptake rates as ferric RA. We conclude that stereoselective recognition of optical isomers takes place during iron uptake mediated by RA and that this recognition primarily involves the right-handed delta coordination "propellor" of the metal center and its adjacent functionalities.

The effect of the intravenous administration of the synthetic siderophore LI-CAM-C on the biodistribution of Ga-67 was investigated. The ligand was found capable of the in vivo complexing with Ga-67, which hastened the renal clearance of the nuclide. The gallium concentration was decreased in all organs, with the exception of the liver and spleen, where we suggest that hydroxide precipitates interfere with gallium sequestration by LICAM-C. The gallium in abscess tissue was only slightly affected, giving rise to an increase in the Ga-67 abscess-to-soft-tissue concentration ratio when LICAM-C is administered. Dosimetry calculations show that the siderophore decreases the radiation burden from Ga-67 citrate. The advantage of clinical application of LICAM-C are discussed.

Isopropyl N-substituted tricatecholamide analogs of enterobactin have been found to form gallium and indium complexes with very high stability constants and to exhibit in vivo characteristics significantly different from gallium- or indium-transferrin and EDTA. The 3,4-DiP-LICAMS and TiP-MECAMS complexes were found to clear primarily through the kidneys, whereas the less polar 3,4-DiP-LICAM complex was eliminated through the liver. The rationale for developing new metal-binding analogs with larger organic groups attached to the amide nitrogens is discussed.