Insight into the formation of N-nitrosodimethylamine in metformin products.

An effective analytical method for the quantification of N-nitrosodimethylamine (NDMA) using a liquid chromatography coupled with tandem mass spectrometry was developed and applied to a process optimization study of the production of metformin film coated tablets in order to identify the key factors behind the NDMA formation in metformin products. The method uses a linear gradient elution with mobile phases 0.1 % formic acid in water for chromatography and methanol for chromatography and a column Acquity UPLC HSS T3 1.8 µm. The use of the tandem mass spectrometry in a positive ion mode with an atmospheric pressure chemical ionization allows for the use of an isotopically labelled internal standard and an external calibration standard. The method was validated according to the guidelines of International Council for Harmonization in terms of limit of detection and quantification, linearity, precision, accuracy and method selectivity. To further justify the effectiveness of the method, a comparison between two laboratories was performed using a linear regression testing. Both methods give comparable results. 469 samples of both metformin active pharmaceutical ingredient and film coated tablets were analysed and the key factors behind NDMA formation were identified. Hypotheses explaining the mechanism were formulated and confronted with measurements and scientific literature. Protective measures to prevent NDMA contamination in metformin products were drawn.

Gadolinium(III) complexes of mono- and diethyl esters of monophosphonic acid analogue of DOTA as potential MRI contrast agents: solution structures and relaxometric studies.

Two new macrocyclic DOTA-like chelates containing one phosphonate pendant arm were synthesised as potential contrast agents for MRI (magnetic resonance imaging). The chelates bind to the lanthanide(III) in an octadentate manner, via four nitrogen atoms, three carboxylate and one phosphonate oxygen atoms. Solution structures of [Ln(do3ap(OEt2))(H(2)O)] and [Ln(do3ap(OEt))(H(2)O)](-) were studied using (31)P and (1)H NMR spectroscopy and SAP (square-antiprismatic)/TSAP (twisted square-antiprismatic) isomerism was observed. Depending on the nature of the lanthanide(III) ion, the lanthanide(III) complexes of H(4)do3ap(OEt) are present in solution as up to four different diastereoisomers observable with NMR. The TSAP isomer is the most abundant at the beginning of the lanthanide series and, with a decrease of the ionic radius of lanthanide(III) ions, both TSAP and SAP forms were observed. A second interconversion (SAP<-->TSAP') becomes important at the end of the series (TSAP' means the TSAP species without a coordinated water molecule). The remaining axial coordination site is occupied by one water molecule for the Gd(3+)-complex. The calculated fraction of the TSAP isomer in the gadolinium(III) complexes increases in the order [Gd(DOTA)(H(2)O)](-) < [Gd(do3ap(OEt2))(H(2)O)] < [Gd(do3ap(OEt))(H(2)O)](-) < [Gd(do3ap)(H(2)O)](2-). Gadolinium(III) complexes of phosphorus-containing chelates, generally, have the advantage of a relatively fast water exchange rate due to a greater sterical demand of the phosphorus acid moiety and of the presence of the second-sphere water shell, which also contributes to the overall relaxivity. The [Gd(do3ap(OEt2))(H(2)O)] and [Gd(do3ap(OEt))(H(2)O)](-) complexes were studied by variable-temperature (17)O NMR and (1)H NMRD. The experimental data were evaluated simultaneously with commonly used equations based on Solomon-Bloembergen-Morgan approximation, extended by a contribution of the second coordination sphere. The water exchange rates were found to be strongly dependent on the TSAP/SAP isomeric ratio and the overall charge of the complex: the monoanionic [Gd(do3ap(OEt))(H(2)O)](-) complex with TSAP molar fraction equal to 0.36 has the water exchange rate of 20 x 10(6) s(-1) (tau(M) = 50 ns) while neutral [Gd(do3ap(OEt2))(H(2)O)] complex with TSAP molar fraction 0.28 has an exchange rate equal to 4.4 x 10(6) s(-1) (tau(M) = 227 ns).

PAMAM dendrimeric conjugates with a Gd-DOTA phosphinate derivative and their adducts with polyaminoacids: the interplay of global motion, internal rotation, and fast water exchange.

A series of dendrimeric conjugates based on a PAMAM (polyamidoamine) backbone with macrocyclic Gd-DO3A-P(ABn) complexes (monophosphinated analogue of DOTA) was prepared. The chelates were covalently attached to the G1-, G2-, and G4-PAMAM dendrimers through a thiourea linker in high loads (>90%). The prepared conjugates G1-(Gd-DO3A-P(BnN{CS}))(8), G2-(Gd-DO3A-P(BnN{CS}))(16), and G4-(Gd-DO3A-P(BnN{CS}))(59) showed relaxivities of 10.1, 14.1, and 18.6 s(-)(1) mM(-)(1) at 20 MHz and 37 degrees C and pH = 7.5, respectively. A variable-pH study (range 2-12) revealed up to 30% increase in the relaxivity at low pH for the G2-(Gd-DO3A-P(BnN{CS}))(16) conjugate. As confirmed by (1)H NMR titration of the unmodified G2 dendrimer, this is due to protonation of core tertiary amines leading to a more open and rigid structure. The variable-temperature (17)O NMR and (1)H NMRD relaxometric studies confirmed that the relaxivity is not controlled by water exchange but by rotational dynamics. A multiparametrical data evaluation using the Lipari-Szabo approach revealed that the water residence lifetime, (298)tau(M), for the conjugates studied was ca. 45-70 ns, which is longer than the value found for the monomeric model compound Gd-DO3A-P(ABn) (16 ns) but short enough so as not to limit the relaxivity. The global rotational correlation time, (298)tau(Rg), varied from 1.5 to 3.1 ns and seemed to indicate a sufficiently slow molecular tumbling to achieve the high relaxivities measured; however, the rigidity factor S(2) (approximately 0.26), describing the internal flexibility, was far from optimum. The overall relaxivity was significantly increased (e.g. by a factor of 1.8 for the G1-(Gd-DO3A-P(BnN{CS}))(8) conjugate) when a positively charged polyaminoacid like poly(Arg) or poly(Lys) was added to the conjugate solutions. The electrostatic interactions partially "freeze" the internal mobility of the conjugate and also slow down global motion. This assumption was confirmed by an evaluation of (1)H relaxometric data obtained for the G2-(Gd-DO3A-P(BnN{CS}))(16)-poly(Lys)(59) adduct. Importantly, it was proved that the adduct formation did not hamper the water exchange process.

Relaxometric and solution NMR structural studies on ditopic lanthanide(III) complexes of a phosphinate analogue of DOTA with a fast rate of water exchange.

A novel "ditopic" ligand containing two monophosphinate triacetate DOTA-like units linked by a thiourea bridge has been synthesized and its complexes with Ln3+ ions (Ln = Y, Eu, Gd, Dy) investigated by NMR spectroscopy and relaxometry. The presence of one water molecule in the first coordination sphere has been determined by the measurement of the dysprosium(III)-induced 17O NMR shifts. The 1H and 31P NMR spectra of the Eu(III) derivative indicate a higher abundance of the fast-exchanging twisted square antiprismatic (m) isomer than the isomeric square antiprismatic (M; m/M = 3:2) complex. The analysis of the 89Y and 13C T1 NMR relaxation times in the Gd(III)/Y(III) mixed complex have provided useful structural information. Values of ca. 6.3 and 8.2 A for the Gd...Y and Gd...C distances, respectively, have been estimated which indicate a rather compact solution structure. This result finds support in the value of the relaxivity whose increase (at 20 MHz and 298 K) on passing from the monomeric (5.7 s(-1) mM(-1)) to the ditopic complex (8.2 s(-1) mM(-1)) can be attributed to the doubling of the inner-sphere term following the doubling of the molecular size. The structural and dynamic relaxivity-controlling parameters were assessed by a simultaneous fitting of the variable temperature 17O NMR and 1H NMRD relaxometric data. The mean water residence lifetime (298tauM) has been found to be 53 ns, one of the shortest values reported for ditopic complexes. The reorientational correlation time is two times longer (298tauR = 183 ps) than the corresponding value of the parent monomeric Gd(III) complex, thus supporting the view of a limited degree of internal rotation. The possible influence of magnetic Gd-Gd coupling has been excluded by a comparison of the 1H NMRD profiles of the homodinuclear Gd(III)/Gd(III) and the heterodinuclear Gd(III)/Y(III) complexes.

Three in one: TSA, TSA', and SA units in one crystal structure of a yttrium(III) complex with a monophosphinated H4dota analogue.

Compound [Y(Hdo3aP(ABn))(H2O)][Y(Hdo3aP(ABn))].6H2O.iPrOH, where H4do3aP(ABn) is 1,4,7,10-tetraazacyclododecane-1,4,7-triacetic-10-methyl(4-aminobenzylphosphinic) acid, a monophosphinate analogue of H4dota, was prepared in the solid state and studied using X-ray crystallography. In contrast to all single-crystal structures of complexes of H4dota-like ligands published before, three distinct units were found in one cell: two species adopting a twisted-square-antiprismatic configuration with (TSA) and without (TSA') a coordinated water molecule and one isomer with a square-antiprismatic configuration (SA). In addition, this is the first complex with the H4dota-like ligand for which the structures of three possible species were determined in the solid state.

A bisphosphonate monoamide analogue of DOTA: a potential agent for bone targeting.

A new macrocyclic DOTA-like ligand (BPAMD) for bone imaging and therapy containing a monoamide bis(phosphonic acid) bone-seeking group was designed and synthesized. Its lanthanide(III) complexes were prepared and characterized by 1H and 31P NMR spectroscopy. The Gd(III)-BPAMD complex was investigated in detail by 1H and 17O relaxometric studies to inspect parameters relevant for its potential application as an MRI contrast agent. Sorption experiments were conducted with Gd(III) and Tb(III) complexes using hydroxyapatite (HA) as a model of bone surface. Very effective uptake of the Gd-BPAMD complex by the HA surface was observed in NMR experiments. Radiochemical studies with the (160Tb-BPAMD)-HA system proved the sorption to be remarkably fast and strong on one hand and fully reversible on the other hand. The strong (Gd-BPAMD)-HA interaction was also supported by 1H NMRD measurements in the presence of a hydroxyapatite slurry, which showed an increase of the rotational correlation time upon adsorption of the complex on the HA surface, resulting in a significant relaxivity enhancement. The amide-bis(phosphonate) moiety is the only factor responsible for the binding of the complex to HA.

Crystal structures of lanthanide(III) complexes with cyclen derivative bearing three acetate and one methylphosphonate pendants.

A series of lanthanide(III) complexes formulated as M[Ln(Hdo3ap)].xH(2)O (M = Li or H and Ln = Tb, Dy, Er, Lu, and Y) with the monophosphonate analogue of H(4)dota, 1,4,7,10-tetraazacyclododecane-1,4,7-triacetic-10-methylphosphonic acid (H(5)do3ap), was prepared in the solid state and studied using X-ray crystallography. All of the structures show that the (Hdo3ap)(4-) anion is octadentate coordinated to a lanthanide(III) ion similarly to the other H(4)dota-like ligands, i.e., forming O(4) and N(4) planes that are parallel and have mutual angle smaller than 3 degrees . The lanthanide(III) ions lie between these planes, closer to the O(4) base than to the N(4) plane. All of the structures present the lanthanide(III) complexes in their twisted-square-antiprismatic (TSA) configuration. Twist angles of the pendants vary in the range between -24 and -30 degrees, and for each complex, they lie in a very narrow region of 1 degree. The coordinated phosphonate oxygen is located slightly above (0.02-0.19 Angstroms) the O(3) plane formed with the coordinated acetates. A water molecule was found to be coordinated only in the terbium(III) and neodymium(III) complexes. The bond distance Tb-O(w) is unusually long (2.678 Angstroms). The O-Ln-O angles decrease from 140 degrees [Nd(III)] to 121 degrees [Lu(III)], thus confirming the increasing steric crowding around the water binding site. A comparison of a number of structures of Ln(III) complexes with DOTA-like ligands shows that the TSA arrangement is flexible. On the other hand, the SA arrangement is rigid, and the derived structural parameters are almost identical for different ligands and lanthanide(III) ions.

Dendrimeric Gd(III) complex of a monophosphinated DOTA analogue: optimizing relaxivity by reducing internal motion.

A marked increase of relaxivity has been observed upon rigidifying the internal frame of Gd-containing PAMAM dendrimers: the effect has been attained by either protonation of the dendrimer or by forming supramolecular adducts with cationic polyaminoacids.

Lanthanide(III) complexes of a mono(methylphosphonate) analogue of H4dota: the influence of protonation of the phosphonate moiety on the TSAP/SAP isomer ratio and the water exchange rate.

A monophosphonate analogue of H4dota, 1,4,7,10-tetraazacyclododecane-4,7,10-tris(carboxymethyl)-1-methylphosphonic acid (H5do3aP), and its complexes with lanthanides were synthesized. Multinuclear NMR studies reveal that, in aqueous solution, lanthanide(III) complexes of the ligand exhibit structures analogous to those of H4dota complexes. Thus, the central ion is nine-coordinate, surrounded by four nitrogen atoms, three acetate and one phosphonate oxygen atoms, and one water molecule in an apical position. For complexes of H5do3aP with Ln(III) ions in the middle of the series, the abundance of the desired twisted square-antiprismatic (TSAP) isomer is higher than for the corresponding H4dota complexes. The TSAP/square-antiprismatic (SAP) isomer ratio is highly sensitive to protonation of the phosphonate group: a higher abundance of the TSAP isomer was found in acidic solutions. The microscopic protonation constants of the TSAP isomers are higher than those of the SAP isomers. The presence of one water molecule in the first coordination sphere of the complexes in the pH region studied (pH 2.5-7.0) is confirmed by 17O NMR spectroscopy. The results of a simultaneous fit of variable-temperature 17O NMR relaxation data and 1H NMRD profiles show that the residence time of water (tauM) in the Gd(III) complex is much smaller than for [Gd(dota)(H2O)]-. The exchange rate appears to be dependent on the pH of the solution. The values of tauM are 37, 40, and 14 ns at pH 2.5, 4.7, and 7.0, respectively. These observations can be explained by an extensive second-sphere hydrogen-bonding network that varies with the state of protonation of the phosphonate moiety. Upon protonation of the complex, the second-sphere hydration probably becomes more ordered, which may result in a decrease in penetrability and an increase in tauM. The relaxivity of the Gd(III) complex is almost independent of the pH and is equal to 4.7 s(-1) mM(-1) (20 MHz, pH 7 and 37 degrees C). The solid-state structure was determined for the Nd(III) complex. It crystallizes as the TSAP isomer and the unit cell contains two independent molecules of the complex with different Nd-O(water) bond lengths of 2.499 and 2.591 A.

Synthesis of a bifunctional monophosphinic acid DOTA analogue ligand and its lanthanide(III) complexes. A gadolinium(III) complex endowed with an optimal water exchange rate for MRI applications.

A new bifunctional octa-coordinating ligand containing an aminobenzyl moiety, DO3APABn (H4DO3APABn = 1,4,7,10-tetraazacyclododecane-4,7,10-triacetic-1-{methyl[(4-aminophenyl)methyl]phosphinic acid}), has been synthesized. Its lanthanide(III) complexes contain one water molecule in the first coordination sphere. The high-resolution 1H and 31P spectra of [Eu(H2O) (DO3APABn)]- show that the twisted square-antiprismatic form of the complexes is more abundant in respect to the corresponding Eu(III)-DOTA complex. The 1H NMRD and variable-temperature 17O relaxation measurements of [Gd(H2O)(DO3APABn)]- show that the water residence time is short (298tauM = 16 ns) and falls into the optimal range predicted by theory for the attainment of high relaxivities once this complex would be endowed by a slow tumbling rate. The relaxivity (298r1 = 6.7 mM(-1) s(-1) at 10 MHz) is higher than expected as a consequence of a significant contribution from the second hydration sphere. These results prompt the use of [Gd(H2O)(DO3APABn)]- as a building block for the set-up of highly efficient macromolecular MRI contrast agents.

Lanthanide(III) complexes of a pyridine N-oxide analogue of DOTA: exclusive M isomer formation induced by a six-membered chelate ring.

The presence of a six-membered chelate ring involving a pyridine N-oxide moiety induces exclusive M isomer formation throughout the whole lanthanide series endowed with a fast water exchange in the case of the Gd(III) complex.