Interaction of macrocyclic gadolinium-based MR contrast agents with Type I collagen. Equilibrium and kinetic studies.

The interactions of gadoterate meglumine, gadobutrol, gadoteridol and Gd(HB-DO3A) with bovine Type I collagen were investigated by ultrafiltration and dialysis. The affinity of the four agents to collagen is similar. However, the maximum adsorbed amount of GdIII-complexes decreases in the following order: gadoterate meglumine > gadobutrol > gadoteridol > Gd(HB-DO3A). Calculations with the open three-compartment model reveal that the structural homologs gadoteridol and Gd(HB-DO3A) have a lower adsorption onto collagen, which may explain the less prolonged in vivo retention of gadoteridol observed in soft tissues of rats.

Comparison of the equilibrium, kinetic and water exchange properties of some metal ion-DOTA and DOTA-bis(amide) complexes.

The 1,7-diacetate-4,10-diacetamide substituted 1,4,7,10-tetraazacyclododecane structural unit is common to several responsive Magnetic Resonance Imaging (MRI) contrast agents (CAs). While some of these complexes (agents capable of sensing fluctuations in Zn2+, Ca2+ etc. ions) have already been tested in vivo, the detailed physico-chemical characterization of such ligands have not been fully studied. To fill this gap, we synthesized a representative member of this ligand family possessing two acetate and two n-butylacetamide pendant side-arms (DO2A2MnBu = 1,4,7,10-tetraazacyclodoecane-1,7-di(acetic acid)-4,10-di(N-butylacetamide)), and studied its complexation properties with some essential metal and a few lanthanide(III) (Ln(III)) ions. Our studies revealed that the ligand basicity, the stability of metal ion complexes, the trend of stability constants along the Ln(III) series, the formation rates of the Ln(III) complexes and the exchange rate of the bound water molecule in the Gd(III) complex fell between those of Ln(DOTA)- and Ln(DOTA-tetra(amide))3+ complexes (DOTA = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid, DOTAM = 1,4,7,10-tetrakis(carbamoylmethyl)-1,4,7,10-tetraazacyclododecane). The only exception is the stability of Cu(DO2A2MnBu) which was found to be only slightly lower than that of Cu(DOTA)2- (log KCuL = 19.85 vs. 21.98). This is likely reflects exclusive coordination of the negatively charged acetate donor atoms to the Cu2+ ion forming an octahedral complex with the amides remaining uncoordinated. The only anomaly observed during the study was the rates of acid assisted dissociation of the Ln(III) complexes, which occur at a rate similar to those observed for the Ln(DOTA)- complexes. These data indicate that even though the Ln(DO2A2MnBu)+ complexes have lower thermodynamic stabilities, their kinetic inertness should be sufficient for in vivo use.

Effect of the Nature of Donor Atoms on the Thermodynamic, Kinetic and Relaxation Properties of Mn(II) Complexes Formed With Some Trisubstituted 12-Membered Macrocyclic Ligands.

During the past few years increasing attention has been devoted to Mn(II) complexes as possible substitutes for Gd(III) complexes as contrast agents in MRI. Equilibrium (log KMnL or pMn value), kinetic parameters (rates and half-lives of dissociation) and relaxivity of the Mn(II) complexes formed with 12-membered macrocyclic ligands were studied. The ligands were selected in a way to gain information on how the ligand rigidity, the nature of the donor atoms in the macrocycle (pyridine N, amine N, and etheric O atom), the nature of the pendant arms (carboxylates, phosphonates, primary, secondary and tertiary amides) affect the physicochemical parameters of the Mn(II) complexes. As expected, decreasing the denticity of DOTA (to afford DO3A) resulted in a drop in the stability and inertness of [Mn(DO3A)]- compared to [Mn(DOTA)]2-. This decrease can be compensated partially by incorporating the fourth nitrogen atom into a pyridine ring (e.g., PCTA) or by replacement with an etheric oxygen atom (ODO3A). Moreover, the substitution of primary amides for acetates resulted in a noticeable drop in the stability constant (PC3AMH), but it increased as the primary amides (PC3AMH) were replaced by secondary (PC3AMGly) or tertiary amide (PC3AMPip) pendants. The inertness of the Mn(II) complexes behaved alike as the rates of acid catalyzed dissociation increased going from DOTA (k1 = 0.040 M-1s-1) to DO3A (k1 = 0.45 M-1s-1). However, the rates of acid catalyzed dissociation decreased from 0.112 M-1s-1 observed for the anionic Mn(II) complex of PCTA to 0.0107 M-1s-1 and 0.00458 M-1s-1 for the cationic Mn(II) complexes of PC3AMH and PC3AMPip ligands, respectively. In spite of its lower denticity (as compared to DOTA) the sterically more hindered amide complex ([Mn(PC3AMPip)]2+) displays surprisingly high conditional stability (pMn = 8.86 vs. pMn = 9.74 for [Mn(PCTA)]-) and excellent kinetic inertness. The substitution of phosphonates for the acetate pendant arms (DOTP and DO3P), however, resulted in a noticeable drop in the conditional stability as well as dissociation kinetic parameters of the corresponding Mn(II) complexes ([Mn(DOTP)]6- and [Mn(DO3P)]4-) underlining that the phosphonate pedant should not be considered as a suitable building block for further ligand design while the tertiary amide moiety will likely have some implications in this respect in the future.

Equilibrium Thermodynamics, Formation, and Dissociation Kinetics of Trivalent Iron and Gallium Complexes of Triazacyclononane-Triphosphinate (TRAP) Chelators: Unraveling the Foundations of Highly Selective Ga-68 Labeling.

In order to rationalize the influence of FeIII contamination on labeling with the 68Ga eluted from 68Ge/68Ga-generator, a detailed investigation was carried out on the equilibrium properties, formation and dissociation kinetics of GaIII- and FeIII-complexes of 1,4,7-triazacyclononane-1,4,7-tris(methylene[2-carboxyethylphosphinic acid]) (H6TRAP). The stability and protonation constants of the [Fe(TRAP)]3- complex were determined by pH-potentiometry and spectrophotometry by following the competition reaction between the TRAP ligand and benzhydroxamic acid (0.15 M NaNO3, 25°C). The formation rates of [Fe(TRAP)] and [Ga(TRAP)] complexes were determined by spectrophotometry and 31P-NMR spectroscopy in the pH range 4.5-6.5 in the presence of 5-40 fold HxTRAP(x-6) excess (x = 1 and 2, 0.15 M NaNO3, 25°C). The kinetic inertness of [Fe(TRAP)]3- and [Ga(TRAP)]3- was examined by the trans-chelation reactions with 10 to 20-fold excess of HxHBED(x-4) ligand by spectrophotometry at 25°C in 0.15 M NaCl (x = 0,1 and 2). The stability constant of [Fe(TRAP)]3- (logKFeL = 26.7) is very similar to that of [Ga(TRAP)]3- (logKGaL = 26.2). The rates of ligand exchange reaction of [Fe(TRAP)]3- and [Ga(TRAP)]3- with HxHBED(x-4) are similar. The reactions take place quite slowly via spontaneous dissociation of [M(TRAP)]3-, [M(TRAP)OH]4- and [M(TRAP)(OH)2]5- species. Dissociation half-lives (t1/2) of [Fe(TRAP)]3- and [Ga(TRAP)]3- complexes are 1.1 × 105 and 1.4 × 105 h at pH = 7.4 and 25°C. The formation reactions of [Fe(TRAP)]3- and [Ga(TRAP)]3- are also slow due to the formation of the unusually stable monoprotonated [*M(HTRAP)]2- intermediates [*logKGa(HL) = 10.4 and *logKFe(HL) = 9.9], which are much more stable than the [*Ga(HNOTA)]+ intermediate [*logKGa(HL) = 4.2]. Deprotonation and transformation of the monoprotonated [*M(HTRAP)]2- intermediates into the final complex occur via OH--assisted reactions. Rate constants (kOH) characterizing the OH--driven deprotonation and transformation of [* Ga(HTRAP)]2- and [*Fe(HTRAP)]2- intermediates are 1.4 × 105 M-1s-1 and 3.4 × 104 M-1s-1, respectively. In conclusion, the equilibrium and kinetic properties of [Fe(TRAP)] and [Ga(TRAP)] complexes are remarkably similar due to the close physico-chemical properties of FeIII and GaIII-ions. However, a slightly faster formation of [Ga(TRAP)] over [Fe(TRAP)] provides a rationale for a previously observed, selective complexation of 68GaIII in presence of excess FeIII.

Improved Efficacy of Synthesizing *MIII-Labeled DOTA Complexes in Binary Mixtures of Water and Organic Solvents. A Combined Radio- and Physicochemical Study.

Typically, the synthesis of radiometal-based radiopharmaceuticals is performed in buffered aqueous solutions. We found that the presence of organic solvents like ethanol increased the radiolabeling yields of [68Ga]Ga-DOTA (DOTA = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacatic acid). In the present study, the effect of organic cosolvents [ethanol (EtOH), isopropyl alcohol, and acetonitrile] on the radiolabeling yields of the macrocyclic chelator DOTA with several trivalent radiometals (gallium-68, scandium-44, and lutetium-177) was systematically investigated. Various binary water (H2O)/organic solvent mixtures allowed the radiolabeling of DOTA at a significantly lower temperature than 95 °C, which is relevant for the labeling of sensitive biological molecules. Simultaneously, much lower amounts of the chelators were required. This strategy may have a fundamental impact on the formulation of trivalent radiometal-based radiopharmaceuticals. The equilibrium properties and formation kinetics of [M(DOTA)]- (MIII= GaIII, CeIII, EuIII, YIII, and LuIII) complexes were investigated in H2O/EtOH mixtures (up to 70 vol % EtOH). The protonation constants of DOTA were determined by pH potentiometry in H2O/EtOH mixtures (0-70 vol % EtOH, 0.15 M NaCl, 25 °C). The log K1H and log K2H values associated with protonation of the ring N atoms decreased with an increase of the EtOH content. The formation rates of [M(DOTA)]- complexes increase with an increase of the pH and [EtOH]. Complexation occurs through rapid formation of the diprotonated [M(H2DOTA)]+ intermediates, which are in equilibrium with the kinetically active monoprotonated [M(HDOTA)] intermediates. The rate-controlling step is deprotonation (and rearrangement) of the monoprotonated intermediate, which occurs through H2O (*M(HL) kH2O) and OH- (*M(HL) kOH) assisted reaction pathways. The rate constants are essentially independent of the EtOH concentration, but the M(HL) kH2O values increase from CeIII to LuIII. However, the log KM(HL)H protonation constants, analogous to the log KH2 value, decrease with increasing [EtOH], which increases the concentration of the monoprotonated M(HDOTA) intermediate and accelerates formation of the final complexes. The overall rates of complex formation calculated by the obtained rate constants at different EtOH concentrations show a trend similar to that of the complexation rates determined with the use of radioactive isotopes.

Exploiting the Proton Exchange as an Additional Route to Enhance the Relaxivity of Paramagnetic MRI Contrast Agents.

The relaxivity of Gd(HP-DO3A) was studied as a function of pH and buffer composition in order to identify the main factors of the observed relaxation enhancement due to the exchange of the coordinated hydroxyl proton. It was established that the paramagnetic relaxation time, T1M, of the coordinated hydroxyl proton is about 50% shorter than that of the protons in the coordinated water molecule. The control of the p K of the coordinated alcoholic -OH moiety in the ligand is fundamental to utilize the proton exchange enhanced relaxivity under physio/pathologic conditions. A new derivative of Gd(HP-DO3A) was synthesized by replacing the -CH3 group with a -CF3 moiety. In this complex, the -OH group becomes more acidic. Consequently, the maximum contribution of the proton exchange to the relaxivity is shifted to a lower pH region with the fluorinated ligand.

Equilibrium, Kinetic and Structural Properties of Gallium(III) and Some Divalent Metal Complexes Formed with the New DATAm and DATA5m Ligands.

The development of 68 Ge/68 Ga generators has made the positron-emitting 68 Ga isotope widely accessible and raised interest in new chelate complexes of Ga3+ . The hexadentate 1,4-di(acetate)-6-methyl[amino(methyl)acetate]perhydro-1,4-diazepane (DATAm ) ligand and its bifunctional analogue, 1,4-di(acetate)-6-pentanoic acid[amino(methyl)acetate]perhydro-1,4-diazepane (DATA5m ), rapidly form complexes with 68 Ga in high radiochemical yield. The stability constants of DATAm and DATA5m complexes formed with Ga3+ , Zn2+ , Cu2+ , Mn2+ and Ca2+ have been determined by using pH potentiometry, spectrophotometry (Cu2+ ) and 1 H and 71 Ga NMR spectroscopy (Ga3+ ). The stability constants of Ga(DATAm ) and Ga(DATA5m ) complexes are slightly higher than those of Ga(AAZTA). The species distribution calculations indicated the predominance of Ga(L)OH mixed-hydroxo complexes at physiological pH. The 1 H and 71 Ga NMR spectroscopy studies provided information about the coordinated functional groups of ligands and on the kinetics of exchange between the Ga(L) and Ga(L)OH complexes. The transmetalation reactions between the Ga(L) complexes and Cu2+ citrate (6

The role of equilibrium and kinetic properties in the dissociation of Gd[DTPA-bis(methylamide)] (Omniscan) at near to physiological conditions.

[Gd(DTPA-BMA)] is the principal constituent of Omniscan, a magnetic resonance imaging (MRI) contrast agent. In body fluids, endogenous ions (Zn(2+), Cu(2+), and Ca(2+)) may displace the Gd(3+). To assess the extent of displacement at equilibrium, the stability constants of DTPA-BMA(3-) complexes of Gd(3+), Ca(2+), Zn(2+), and Cu(2+) have been determined at 37 °C in 0.15 M NaCl. The order of these stability constants is as follows: GdL≈CuL>ZnL≫CaL. Applying a simplified blood plasma model, the extent of dissociation of Omniscan (0.35 mM [Gd(DTPA-BMA)]) was found to be 17% by the formation of Gd(PO4), [Zn(DTPA-BMA)](-) (2.4%), [Cu(DTPA-BMA)](-) (0.2%), and [Ca(DTPA-BMA)](-) (17.7%). By capillary electrophoresis, the formation of [Ca(DTPA-BMA)](-) has been detected in human serum spiked with [Gd(DTPA-BMA)] (2.0 mM) at pH 7.4. Transmetallation reactions between [Gd(DTPA-BMA)] and Cu(2+) at 37 °C in the presence of citrate, phosphate, and bicarbonate ions occur by dissociation of the complex assisted by the endogenous ligands. At physiological concentrations of citrate, phosphate, and bicarbonate ions, the half-life of dissociation of [Gd(DTPA-BMA)] was calculated to be 9.3 h at pH 7.4. Considering the rates of distribution and dissociation of [Gd(DTPA-BMA)] in the extracellular space of the body, an open two-compartment model has been developed, which allows prediction of the extent of dissociation of the Gd(III) complex in body fluids depending on the rate of elimination of the contrast agent.

Solution structures, stabilities, kinetics, and dynamics of DO3A and DO3A-sulphonamide complexes.

The Gd(3+)-DO3A-arylsulphonamide (DO3A-SA) complex is a promising pH-sensitive MRI agent. The stability constants of the DO3A-SA and DO3A complexes formed with Mg(2+), Ca(2+), Mn(2+), Zn(2+), and Cu(2+) ions are similar, whereas the logKLnL values of Ln(DO3A-SA) complexes are 2 orders of magnitude higher than those of the Ln(DO3A) complexes. The protonation constant (log KMHL) of the sulphonamide nitrogen in the Mg(2+), Ca(2+), Mn(2+), Zn(2+), and Cu(2+) complexes is very similar to that of the free ligand, whereas the logKLnHL values of the Ln(DO3A-SA) complexes are lower by about 4 logK units, indicating a strong interaction between the Ln(3+) ions and the sulphonamide N atom. The Ln(HDO3A-SA) complexes are formed via triprotonated *Ln(H3DO3A-SA) intermediates which rearrange to the final complex in an OH(-)-assisted deprotonation process. The transmetalation reaction of Gd(HDO3A-SA) with Cu(2+) is very slow (t1/2 = 5.6 × 10(3) h at pH = 7.4), and it mainly occurs through proton-assisted dissociation of the complex. The (1)H and (13)C NMR spectra of the La-, Eu-, Y-, and Lu(DO3A-SA) complexes have been assigned using 2D correlation spectroscopy (COSY, EXSY, HSQC). Two sets of signals are observed for Eu-, Y-, and Lu(DO3A-SA), showing two coordination isomers in solution, that is, square antiprismatic (SAP) and twisted square antiprismatic (TSAP) geometries with ratios of 86-14, 93-7, and 94-6%, respectively. Line shape analysis of the (13)C NMR spectra of La-, Y- , and Lu(DO3A-SA) gives higher rates and lower activation entropy values compared to Ln(DOTA) for the arm rotation, which indicates that the Ln(DO3A-SA) complexes are less rigid due to the larger flexibility of the ethylene group in the sulphonamide pendant arm. The fast isomerization and the lower activation parameters of Ln(DO3A-SA) have been confirmed by theoretical calculations in vacuo and by using the polarizable continuum model. The solid state X-ray structure of Cu(H2DO3A-SA) shows distorted octahedral coordination. The coordination sites of Cu(2+) are occupied by two ring N- and two carboxylate O-atoms in equatorial position. The other two ring N-atoms complete the coordination sphere in axial positions. The solid state structure also indicates that a carboxylate O atom and the sulphonamide nitrogen are protonated and noncoordinated.

Dissociation kinetics of open-chain and macrocyclic gadolinium(III)-aminopolycarboxylate complexes related to magnetic resonance imaging: catalytic effect of endogenous ligands.

The kinetics of the metal exchange reactions between open-chain Gd(DTPA)(2-) and Gd(DTPA-BMA), macrocyclic Gd(DOTA)(-) and Gd(HP-DO3A) complexes, and Cu(2+)  ions were investigated in the presence of endogenous citrate, phosphate, carbonate and histidinate ligands in the pH range 6-8 in NaCl (0.15 M) at 25 °C. The rates of the exchange reactions of Gd(DTPA)(2-) and Gd(DTPA-BMA) are independent of the Cu(2+) concentration in the presence of citrate and the reactions occur via the dissociation of Gd(3+)  complexes catalyzed by the citrate ions. The HCO(3)(-)/CO(3)(2-) and H(2)PO(4)(-) ions also catalyze the dissociation of complexes. The rates of the dissociation of Gd(DTPA-BMA), catalyzed by the endogenous ligands, are about two orders of magnitude higher than those of the Gd(DTPA)(2-). In fact near to physiological conditions the bicarbonate and carbonate ions show the largest catalytic effect, that significantly increase the dissociation rate of Gd(DTPA-BMA) and make the higher pH values (when the carbonate ion concentration is higher) a risk-factor for the dissociation of complexes in body fluids. The exchange reactions of Gd(DOTA)(-) and Gd(HP-DO3A) with Cu(2+) occur through the proton assisted dissociation of complexes in the pH range 3.5-5 and the endogenous ligands do not affect the dissociation rates of complexes. More insights into the interaction scheme between Gd(DTPA-BMA) and Gd(DTPA)(2-) and endogenous ligands have been obtained by acquiring the (13)C NMR spectra of the corresponding diamagnetic Y(III)-complexes, indicating the increase of the rates of the intramolecular rearrangements in the presence of carbonate and citrate ions. The herein reported results may have implications in the understanding of the etiology of nephrogenic systemic fibrosis, a rare disease that has been associated to the administration of Gd-containing agents to patients with impaired renal function.

Determination of gadolinium-based magnetic resonance imaging contrast agents by micellar electrokinetic capillary chromatography.

MEKC with DAD was applied to detect six Gd-based contrasting agents (CAs) (Gd-DTPA-BMA (Omniscan), Gd-HPDO3A (ProHance), Gd-DOTA (Dotarem), Gd-AAZTA, Gd-BOPTA (Multihance) and Gd-DTPA (Magnevist)) commonly used in MRI diagnostics. The achieved LODs ranged between 0.40 and 20 µM and the optimized method gave excellent precision, especially when two internal standards were applied (less than 0.34 RSD% for migration time). The MEKC technique made it possible to determine the CAs in urine and serum samples of patients having a therapeutic dose. Due to the SDS content of the running buffer, the serum samples can be directly injected to analyze Gd-based CAs without interference of high protein content.

Kinetics of the exchange reactions between Gd(DTPA)2-, Gd(BOPTA)2-, and Gd(DTPA-BMA) complexes, used as MRI contrast agents, and the triethylenetetraamine-hexaacetate ligand.

The kinetics of ligand exchange reactions occurring between the Gd(DTPA), Gd(BOPTA), and Gd(DTPA-BMA) complexes, used as contrast agents in MRI, and the ligand TTHA, have been studied in the pH range 6.5-11.0 by measuring the water proton relaxation rates at 25 °C in 0.15 M NaCl. The rates of the reactions are directly proportional to the concentration of TTHA, indicating that the reactions take place with the direct attack of the H(i)TTHA((6-i)-) (i = 0, 1, 2 and 3) species on the Gd(3+) complexes, through the formation of ternary intermediates. The rates of the exchange reactions of the neutral Gd(DTPA-BMA) increase when the pH is increased from 6.5 to 9, because the less protonated H(i)TTHA((6-i)-) species can more efficiently attack the Gd(3+) complex. The rates of the exchange reactions of [Gd(DTPA)](2-) and [Gd(BOPTA)](2-) also increase from pH 8.5 to 11, but from 6.5 to 8.5 an unexpected decrease was observed in the reaction rates. The decrease has been interpreted by assuming the validity of general acid catalysis. The protons from the H(i)TTHA((6-i)-) species (i = 2 and 3) can be transferred to the coordinated DTPA or BOPTA in the ternary intermediates when the dissociation of the Gd(3+) complexes occurs faster. The kinetic inertness of Gd(DTPA), Gd(BOPTA), and Gd(DTPA-BMA) differs very considerably; the rates of the ligand exchange reactions of Gd(DTPA-BMA), thus the rates of its dissociation, are 2 to 3 orders of magnitude higher than those of Gd(DTPA) and Gd(BOPTA). The rates of the ligand exchange reactions increase with increasing concentration of the endogenous citrate, phosphate, or carbonate ions at a pH of 7.4, but the effect of citrate and phosphate is negligible at their physiological concentrations. The increase in the reaction rates at the physiological concentration of the carbonate ion is significant (20-60%), and the effect is the largest for the Gd(DTPA-BMA) complex.

Dramatic increase of selectivity for heavy lanthanide(III) cations by tuning the flexibility of polydentate chelators.

Two novel octadentate ligands have been synthesized by attaching two terminal iminodiacetic groups to either 1,4-diazepane (BCAED) or piperazine (BCAEP) as central scaffold. The introduction of the seven- or six-membered ring into the ligand backbone is expected to modify their overall flexibility and then to affect the stability of the corresponding lanthanide(III) complexes. In this work, thermodynamic stability data are determined for the formation of the complexes of BCAED and BCAEP with La(3+), Nd(3+), Eu(3+), Gd(3+), Ho(3+), and Lu(3+). The ligand BCAED shows a strong binding affinity for Lu(3+) (logK = 20.99), moderate for Gd(3+) (logK = 17.15) and rather weak for La(3+) (logK = 12.77). Thus, the variation of logK across the Ln series assumes the remarkable value of 8.22, the largest so far reported. This points to a predominant role of a suitable size match between the metal ion and the ligand cavity, determined by its structure and flexibility. The ligand BCAEP forms less stable complexes with lanthanide(III) cations although it retains a good selectivity (DeltalogK(La-Lu) = 5.66). The Gd(III) complexes have been investigated in aqueous solution by measuring their relaxivity as a function of pH, at 20 MHz and 25 degrees C. The results can be interpreted very well in terms of the species distribution curves calculated from the thermodynamic data and indicate that in these complexes Gd(3+) is octacoordinated, without any bound water molecule. This coordination geometry is maintained in the solid state as shown by the X-ray crystal structure of [Na(H(2)O)(2)][Gd(BCAED)] where the metal ion is at the center of a bicapped-trigonal prism. Finally, the (13)C NMR spectra (9.4 T, 25 degrees C) of the diamagnetic La(3+), Y(3+), and Lu(3+) complexes show that a pronounced stereochemical rigidity is associated with the thermodynamically more stable complexes.

Equilibrium and kinetic properties of the lanthanoids(III) and various divalent metal complexes of the heptadentate ligand AAZTA.

The heptadentate ligand 1,4-bis(hydroxycarbonylmethyl)-6-[bis(hydroxycarbonylmethyl)]amino-6-methylperhydro-1,4-diazepine (AAZTA) and its derivatives were recently reported to give stable complexes with Gd(3+) with superior efficiency as MRI contrast agents. Nevertheless, only preliminary data are available on the coordination behavior of this interesting ligand. In this work, thermodynamic and kinetic stability data are determined for the formation of complexes with AAZTA and the lanthanoid metal ions, and other divalent metal ions of interest for this application. The AAZTA ligand binds the lanthanoid ions with log K(ML) values of 17.53-21.85 with its affinity steadily increasing from La(3+) to Lu(3+), suggesting that the seven-membered skeleton is better suited to accommodate smaller metal ions. Even though the denticity is lower, the stability of the heavier lanthanoid complexes is comparable to those of the classical ligand diethylenetriaminepentaacetic acid (DTPA). The transmetalation reactions of [Gd(AAZTA)](-) with Cu(2+) and Eu(3+) predominantly occur through proton-assisted dissociation of the complex. The role of the direct attack of Cu(2+) or Eu(3+) in the exchange reactions is limited, although the formation of dinuclear complexes decreases the proton-assisted dissociation. Near physiological conditions, [Gd(AAZTA)](-) is significantly more inert than [Gd(DTPA)](2-), allowing its potentially safe use as contrast agent in magnetic resonance imaging.

Synthesis, potentiometric, kinetic, and NMR Studies of 1,4,7,10-tetraazacyclododecane-1,7-bis(acetic acid)-4,10-bis(methylenephosphonic acid) (DO2A2P) and its complexes with Ca(II), Cu(II), Zn(II) and lanthanide(III) ions.

A cyclen-based ligand containing trans-acetate and trans-methylenephosphonate pendant groups, H 6DO2A2P, was synthesized and its protonation constants (12.6, 11.43, 5.95, 6.15, 2.88, and 2.77) were determined by pH-potentiometry and (1)H NMR spectroscopy. The first two protonations were shown to occur at the two macrocyclic ring N-CH 2-PO 3 (2-) nitrogens while the third and fourth protonations occur at the two phosphonate groups. In parallel with protonation of the two -PO 3 (2-) groups, the protons from the NH (+)-CH 2-PO 3 (2-) are transferred to the N-CH 2-COO (-) nitrogens. The stability constants of the Ca (2+), Cu (2+), and Zn (2+) (ML, MHL, MH 2L, and M 2L) complexes were determined by direct pH-potentiometry. Lanthanide(III) ions (Ln (3+)) form similar species, but the formation of complexes is slow; so, "out-of-cell" pH-potentiometry (La (3+), Eu (3+), Gd (3+), Y (3+)) and competitive spectrophotometry with Cu(II) ion (Lu (3+)) were used to determine the stability constants. By comparing the log K ML values with those of the corresponding DOTA (H 4DOTA = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) and DOTP (H 8DOTP = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetramethylenephosphonic acid) complexes, the order DOTA < DO2A2P < DOTP was found for all the metal ion complexes examined here with the exception of the Ca (2+) complexes, for which the order is reversed. The relaxivity of Gd(DO2A2P) decreases between pH 2 and 7 but remains constant in the pH range of 7 < pH < 12 ( r 1 = 3.6 mM (-1) s (-1)). The linewiths of the (17)O NMR signals of water in the absence and presence of Gd(DO2A2P) (at pH = 3.45 and 8.5) between 274 and 350 K are practically the same, characteristic of a q = 0 complex. Detailed kinetic studies of the Ce (3+) and Gd (3+) complexes with DO2A2P showed that complex formation is slow and involves a high stability diprotonated intermediate Ln(H 2DO2A2P)*. Rearrangement of the diprotonated intermediate into the final complex is an OH (-) assisted process but, unlike formation of Ln(DOTA) complexes, rearrangement of Ln(H 2DO2A2P)* also takes place spontaneously likely as a result of transfer of one of the protons from a ring nitrogen to a phosphonate group. The order of the OH (-) assisted formation rates of complexes is DOTA > DO2A2P > DOTP while the order of the proton assisted dissociation rates of the Gd (3+) complexes is reversed, DOTP > DO2A2P > DOTA. (1)H and (13)C NMR spectra of Eu(DO2A2P) and Lu(DO2A2P) were assigned using two-dimensional correlation spectroscopy (2D COSY), heteronuclear multiple quantum coherence (HMQC), heteronuclear chemical shift correlation (HETCOR), and exchange spectroscopy (EXSY) NMR methods. Two sets of (1)H NMR signals were observed for Eu(DO2A2P) characteristic of the presence of two coordination isomers in solution, a twisted square antiprism (TSAP) and a square antiprism (SAP), in the ratio of ~93% and ~7%, respectively. Line shape analysis of the (1)H NMR spectra of Lu(DO2A2P) gave lower activation parameters compared to La(DOTP) for interconversion between coordination isomers. This indicates that the Ln(DO2A2P) complexes are less rigid probably due to the different size and spatial requirements of the carboxylate and phosphonate groups.

Potentiometric and relaxometric properties of a gadolinium-based MRI contrast agent for sensing tissue pH.

The pH-sensitive contrast agent, GdDOTA-4AmP (Gd1) has been successfully used to map tissue pH by MRI. Further studies now demonstrate that two distinct chemical forms of the complex can be prepared depending upon the pH at which Gd(3+) is mixed with ligand 1. The desired pH-sensitive form of this complex, referred to here as a Type II complex, is obtained as the exclusive product only when the complexation reaction is performed above pH 8. At lower pH values, a second complex is formed that, by analogy with an intermediate formed during the preparation of GdDOTA, we tentatively assign to a Type I complex where the Gd(3+) is coordinated only by the appended side-chain arms of 1. The proportion of Type I complex formed is largely determined by the pH of the complexation reaction. The magnitude of the pH-dependent change in the relaxivity of Gd1 was found to be less than earlier reported (Zhang, S.; Wu, K.; Sherry, A. D. Angew. Chem., Int. Ed. 1999, 38, 3192), likely due to contamination of the earlier sample by an unknown amount of Type I complex. Examination of the nuclear magnetic relaxation dispersion and relaxivity temperature profiles, coupled with information from potentiometric titrations, shows that the amphoteric character of the phosphonate side chains enables rapid prototropic exchange between the single bound water of the complex with the bulk water thereby giving Gd1 a unique pH-dependent relaxivity that is quite useful for the pH mapping of tissues by MRI.

Synthesis and Characterization of DOTA-(amide)(4) Derivatives: Equilibrium and Kinetic Behavior of Their Lanthanide(III) Complexes.

Lanthanide complexes of tetraamide derivatives of DOTA are of interest today because of their application as chemical exchange saturation transfer (CEST) agents for magnetic resonance imaging (MRI). The protonation constants of some simple tetraamide derivatives of DOTA and the stability constants of the complexes formed with some endogenous metal ions, namely Mg(2+), Ca(2+), Cu(2+), Zn(2+), and lanthanide(III) ions, have been studied. These complexes were found to be considerably less stable than the corresponding [M(DOTA)](2-) complexes, largely due to the lower basicity of the tetraamide ligands. The Mg(2+) and Ca(2+) complexes are well described by formation of only ML species at equilibrium while the Zn(2+) and Cu(2+) complexes exhibit one and two additional deprotonation steps above a pH of around 6, respectively. The extra deprotonation that occurs at high pH for the [Zn{DOTA-(amide)(4)}](2+) complexes has been assigned to an amide deprotonation by (1)H NMR spectroscopy. The first deprotonation step for the Cu(2+) complexes was traced to formation of the ternary hydroxo complexes ML(OH) (by UV/Vis spectrophotometry) while the second step corresponds to deprotonation of an amide group to form ML(OH)H(-1)-type complexes. The trends in the stability constants of the [Ln{DOTA-(amide)(4)}](3+) complexes follow similar trends with respect to ion size as those reported previously for the corresponding [Ln(DOTA)](-) complexes, but again, the stability constants are about 10-11 orders of magnitude lower. A kinetic analysis of complex formation has shown that complexes are directly formed between a Ln(3+) cation and fully deprotonated L without formation of a protonated intermediate. [Ln{DOTA-(MeAm)(4)}](3+) complex formation occurs at a rate that is two to three orders of magnitude slower than those of the corresponding [Ln(DOTA)](-) complexes, while the variation in complex formation rates with Ln(3+) ion size is opposite to that observed for the corresponding [Ln(DOTA)](-) complexes. The Ce(3+) and Eu(3+) complexes of DOTA-(MeAm)(4) are kinetically inert with respect to acid-catalyzed dissociation, which suggests that these complexes may potentially be safe for use in vivo.

Equilibria and structure of the lanthanide(III)-2-hydroxy-1,3-diaminopropane-N,N,N',N'-tetraacetate complexes: formation of alkoxo-bridged dimers in solid state and solution.

The complex formed between 1,3-diamino-2-hydroxypropane-N,N,N',N'-tetraacetic acid (H4L-OH) and Nd3+ at pH 7.5 was found to be a dinuclear dimer in the solid state by X-ray crystallography. In the complex K4[Nd2(L-O)2(H2O)2].14H2O each ligand is coordinated to both Nd3+ atoms with an iminodiacetate group (the Nd3+-Nd3+ distance is 3.9283(8) A). The alcoholic OH groups are deprotonated, and the alkoxo oxygens are coordinated to both Nd3+ in a bridging position. The Nd3+ ions are nine-coordinated with one water molecule per Nd(III) ion in the inner sphere. The complex K4[Nd2(L-O)2(H2O)2].14H2O has an inversion center, and the space group is P1. Two of the K+ counterions are six-coordinated, while the other two K+ ions are eight-coordinated; polar polymeric water-K+ layers are formed between the apolar ligand layers via the bridging water molecules. The dinuclear dimer complexes are also present in aqueous solution. The proton relaxivities of the Gd3+ complex decrease with the increase of pH, and at pH > 6, the low relaxivity values indicate the probable absence of H2O in the inner sphere and the predominance of the eight-coordinated dimer species [Gd2(L-O)2].4- The results of ESI-TOF MS studies of the complexes of La3+, Nd3+, and Lu3+ proved the formation of dinuclear dimers in dilute (0.25 mM) solutions. pH-potentiometric titrations indicate the formation of complexes with 1:1 (Ln(L-OH)-, Ln(HL-OH), and Ln2(L-O)24-) and 2:1 (Ln2(L-O)+) metal-to-ligand ratios. The stability constants of the Ln(L-OH)- species increase from La3+ (log K = 10.19) to Lu3+ (log K = 14.08). The alcoholic OH group of the Ln(L-OH)- species dissociates at unusually low pH values. The pH range of dissociation shifts to lower and lower pH's with the increasing atomic number of the lanthanides. This pH range is about 4-7 for the La3+ complex and 1-4 for the Lu3+ complex. The results of 1D and 2D 1H and 13C NMR studies of the La3+ complex, the number and multiplicity of signals, and the values of coupling constants are in agreement with the dinuclear dimer structure of the complex in solution.

Equilibrium and NMR studies on GdIII, YIII, CuII and ZnII complexes of various DTPA-N,N''-bis(amide) ligands. Kinetic stabilities of the gadolinium(III) complexes.

Three DTPA-derivative ligands, the non-substituted DTPA-bis(amide) (L(0)), the mono-substituted DTPA-bis(n-butylamide) (L(1)) and the di-substituted DTPA-bis[bis(n-butylamide)] (L(2)) were synthesized. The stability constants of their Gd3+ complexes (GdL) have been determined by pH-potentiometry with the use of EDTA or DTPA as competing ligands. The endogenous Cu2+ and Zn2+ ions form ML, MHL and M(2)L species. For the complexes CuL(0) and CuL(1) the dissociation of the amide hydrogens (CuLH(-1)) has also been detected. The stability constants of complexes formed with Gd3+, Cu2+ and Zn2+ increase with an increase in the number of butyl substituents in the order ML(0) < ML(1) < ML(2). NMR studies of the diamagnetic YL(0) show the presence of four diastereomers formed by changing the chirality of the terminal nitrogens of their enantiomers. At 323 K, the enantiomerization process, involving the racemization of central nitrogen, falls into the fast exchange range. By the assignment and interpretation of 1H and 13C NMR spectra, the fractions of the diastereomers were found to be equal at pH = 5.8 for YL(0). The kinetic stabilities of GdL(0), GdL(1) and GdL(2) have been characterized by the rates of the exchange reactions occurring between the complexes and Eu3+, Cu2+ or Zn2+. The rates of reaction with Eu3+ are independent of the [Eu3+] and increase with increasing [H+], indicating the rate determining role of the proton assisted dissociation of complexes. The rates of reaction with Cu2+ and Zn2+ increase with rising metal ion concentration, which shows that the exchange can take place with direct attack of Cu2+ or Zn2+ on the complex, via the formation of a dinuclear intermediate. The rates of the proton, Cu2+ and Zn2+ assisted dissociation of Gd3+ complexes decrease with increasing number of the n-butyl substituents, which is presumably the result of steric hindrance hampering the formation or dissociation of the intermediates. The kinetic stabilities of GdL(0) and GdL(1) at pH = 7.4, [Cu2+] = 1 x 10(-6) M and [Zn(2+)] = 1 x 10(-5) M are similar to that of Gd(DTPA)2-, while the complex GdL2 possesses a much higher kinetic stability.

Copper(II) complexes of some N-substituted bis(aminomethyl)phosphinate ligands. An integrated EPR study of microspeciation and coordination modes by the two-dimensional simulation method.

Copper(II) complexes of bis(aminomethyl)phosphinic acid (L1), bis(N-glycino-N-methyl)phosphinic acid (L2), bis(N-benzylglycino-N-methyl)phosphinic acid (L3), bis(l-prolino-N-methyl)phosphinic acid (L4) and bis(iminodicarboxymethyl-N-methyl)phosphinic acid (L5) were studied in aqueous solution by pH-potentiometric and electron paramagnetic resonance (EPR) spectroscopic methods. The EPR spectrum packages recorded at various ligand-to-metal concentration ratios and pH's were analyzed (after matrix rank analysis by the method of residual intensities as a complementary method) by the two-dimensional computer simulation method, which simultaneously determines the formation constants and the EPR parameters of the various (micro)species. L1 forms mono and bis complexes in different protonation states; for the other ligands, the mono complexes are always prevalent. For steric reasons, the formation of CuL is shifted to increasingly higher pH regions in the sequence L2, L3 and L4. CuLH was identified for L3, L4 and L5, and also CuLH(2) for L4 and L5. Cu(2)L(2) was found in small amounts for L3 and L4, while it predominates at pH>4 for L5. For L5, Cu(2)L(2)H(2) was also detected. For the ligands that form dimeric metal complexes in equimolar solution or at a ligand excess, Cu(2)L is formed at a metal ion excess. Ligation of the phosphinate O was suggested by indirect proofs in the protonated complexes of L1. For the ligands L2, L3 and L4, the copper(II) coordination in various species in different protonation states is reminiscent of that in the mono and bis complexes of simple amino acids. For the bis(aminomethyl)phosphinates, however, the cis positions of the amino groups in CuL are ensured by the structure of the ligand, and the isomers differ from each other in the (equatorial or axial) position of the second carboxylate group.