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Novel 1,10-phenanthroline-2,9-bistriazoles derivatives have been synthesized by copper-catalyzed azide/alkyne cycloaddition reactions and assessed for their ability to bind and stabilize G-quadruplex (G4) structures. Ten novel compounds were evaluated using Förster resonance energy transfer (FRET) melting, circular dichroism (CD), and fluorescence spectroscopy on several G4 sequences. Biophysical characterization led to the identification of compounds 4 a, 4 b, and 5 b as good G4 ligands of KRAS G4 sequences. The impact on cell viability of all derivatives was also assessed, revealing weak effects. However, compound 2 a exhibited cytotoxicity activity on A549 and H1299 cancer cells and low cytotoxicity towards MRC-5 non-malignant cells MRC-5 not connected with its G4-binding ability. Flow cytometry showed that 2 a induced a cell viability decrease in S and G2/M phases for A549 and H1299; thus, more studies should be performed to explore the proteins involved in cell cycle regulation., (© 2024 Wiley-VCH GmbH.)

Due to the relevance of lactic acidosis in cancer, several therapeutic strategies have been developed targeting its production and/or regulation. In this matter, inhibition approaches of key proteins such as lactate dehydrogenase or monocarboxylate transporters have showed promising results, however, metabolic plasticity and tumor heterogeneity limits their efficacy. In this study, we explored the anticancer potential of a new strategy based on disturbing lactate permeability independently of monocarboxylate transporters activity using a small molecule ionophore named Lactrans-1. Derived from click-tambjamines, Lactrans-1 facilitates transmembrane lactate transportation in liposome models and reduces cancer cell viability. The results showed that Lactrans-1 triggered both apoptosis and necrosis depending on the cell line tested, displaying a synergistic effect in combination with first-line standard chemotherapeutic cisplatin. The ability of this compound to transport outward lactate anions was confirmed in A549 and HeLa cells, two cancer cell lines having distinct rates of lactate production. In addition, through cell viability reversion experiments it was possible to establish a correlation between the amount of lactate transported and the cytotoxic effect exhibited. The movement of lactate anions was accompanied with intracellular pH disturbances that included basification of lysosomes and acidification of the cytosol and mitochondria. We also observed mitochondrial swelling, increased ROS production and activation of oxidative stress signaling pathways p38-MAPK and JNK/SAPK. Our findings provide evidence that enhancement of lactate permeability is critical for cellular pH homeostasis and effective to trigger cancer cell death, suggesting that Lactrans-1 may be a promising anticancer therapy., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Piperazines and diazepines are examples of nitrogen heterocycles present in many marketed drugs highlighting their importance in the discovery of novel bioactive compounds. However, their synthesis often faces challenges, including complex functionalization and lengthy reaction sequences. Multicomponent reactions, notably the Ugi reaction, have emerged as powerful tools to address these hurdles. Here, we have demonstrated the possibility of using the combination of arylglyoxals and carboxylic acids tethered to nonprotected deactivated amines as a powerful strategy for the synthesis of complex fused heterocycles. The limited nucleophilic character of the amino group of the anthranilic acid, indole-2-carboxylic acid, pyrrole-2-carboxylic acid or N -phenylglycine has allowed the use of these compounds in the Ugi reaction without triggering competitive reactions. The additional functional group present in the resulting Ugi adduct can be leveraged in different post-condensation strategies to easily generate multiple fused nitrogen heterocycles including benzodiazepinone and piperazinone cores., (Copyright © 2024, Gómez-Ayuso et al.)

Six novel click-tambjamines (1-6) bearing an alkyl chain of varying length linked to the imine moiety have been formulated in nanostructured lipid carriers (NLCs) to evaluate their transmembrane anion transport activity both when free ( i.e. , not encapsulated) and nanoformulated. Nanostructured lipid carriers (NLCs) are an example of drug delivery systems (DDSs) that stand out because of their versatility. In this work we show that NLCs can be used to efficiently formulate highly lipophilic anionophores and experiments conducted in model liposomes reveal that these formulations are adequate to deliver anionophores without compromising their transport activity. This result paves the way to facilitate the study of highly lipophilic anionophores and their potential use as future drugs.

An excessive production of lactate by cancer cells fosters tumor growth and metastasis. Therefore, targeting lactate metabolism and transport offers a new therapeutic strategy against cancer, based on dependency of some cancer cells for lactate as energy fuel or as oncogenic signal. Herein we present a family of anionophores based on the structure of click-tambjamines that have proved to be extremely active lactate carriers across phospholipid membranes. Compound 1, the most potent lactate transmembrane carrier, was studied in HeLa cells. The use of a monocarboxylate transporters (MCTs) inhibitor proved that 1 is an active lactate transporter in living cells, confirming the results obtained in phospholipid vesicles. Moreover, an additive effect of compound 1 with cisplatin was observed in HeLa cells. Identification of active lactate anionophores working in living cells opens up ways to exploit this class of compounds as molecular tools and drugs addressing dysregulated lactate metabolism., Competing Interests: A patent application (P202330370) related to this work has been submitted., (© 2023 The Authors.)

After more than three decades of extensive investigations on supramolecular polymers, strategies for self-limiting growth still remain challenging. Herein, we exploit a new V-shaped monomer design to achieve anticooperatively formed oligomers with superior robustness and high luminescence. In toluene, the monomer-oligomer equilibrium is shifted to the monomer side, enabling the elucidation of the molecular packing modes and the resulting (weak) anticooperativity. Steric effects associated with an antiparallel staircase organization of the dyes are proposed to outcompete aromatic and unconventional B-F⋅⋅⋅H-N/C interactions, restricting the growth at the stage of oligomers. In methylcyclohexane (MCH), the packing modes and the anticooperativity are preserved; however, pronounced solvophobic and chain-enwrapping effects lead to thermally ultrastable oligomers. Our results shed light on understanding anticooperative effects and restricted growth in self-assembly., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Quality control in the food industry is of the upmost importance from the food safety, organoleptic and commercial viewpoints. Accordingly, the development of in situ , rapid, and costless analytical tools is a valuable task in which we are working. Regarding this point, the copper content of grape must has to be determined by wineries along the wine production process. For this purpose, grape must samples are sent to laboratories where the copper content is measured usually by flame atomic absorption spectrometry or by inductively coupled plasma mass spectrometry. We herein propose a straightforward, rapid, and inexpensive methodology based both on a film-shaped colorimetric polymer sensor and a smartphone method that at the same time can be used by unskilled personnel. The sensory polymer films change their color upon dipping them on the grape must, and the color evolution is analyzed using the digital color parameters of a picture taken to the film with a smartphone. Furthermore, the analytical procedure is automatically carried out by a smartphone app. The limit of detection of copper of the polymer sensor is 0.08 ppm. Following this approach, 18 production samples coming from the French Groupe ICV company were studied. The copper content of the samples was analyzed by the usual procedure carried out by the company (flame atomic absorption spectrometry) and by the method proposed in this work, ranging this content from 0.41 to 6.08 ppm. The statistical study showed that the results of both methods are fully consistent, showing the validity of the proposed method for the determination of copper in grape must within the frame of wine production wineries and industries.

We report on an innovative method to measure the Zn(II) concentration in commercial pet food samples, both wet and dry food. It is based on a colorimetric sensory polymer prepared from commercial monomers and 0.5 % of a synthetic monomer having a quinoline sensory core (N-(8-(2-azidoacetamido)quinolin-5-yl)methacrylamide). We obtained the sensory polymer as crosslinked films by thermally initiated bulk radical polymerization of the monomers of 100 μm thickness, which we punched into Ø6 mm sensory discs. The immersion of the discs in water solutions containing Zn(II) turned the fluorescence on, allowing for the titration of this cation using the G parameter of a digital picture taken to the discs. The limits of detection and quantification were 29 and 87 µg/L, respectively. Furthermore, we measured the concentration of Zn(II) even in the presence of other cations, detecting no significant interferences. Thus, in a further step, we obtained the concentration of Zn(II) from 15 commercial pet food samples, ranging from 19 to 198 mg/kg, following a simple extraction procedure and contacting the extractant with our sensory discs. These results were contrasted with that obtained by ICP-MS as a reference method., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

G-quadruplex (G4) structures are non-canonical DNA/RNA secondary structures able to form within guanine rich nucleic acids sequences. They are present in several regions of the human genome including gene promoters, untranslated sequences, and telomeres. Due to their biological relevance G4 structures are considered important drug targets, in particular for anticancer therapies, leading to the development of G4 stabilizing small molecules. Telomeric regions have received special attention in this field since they can fold into several distinct intramolecular G-quadruplexes topologies. Herein, we report the synthesis of 2,9-disubstituted-1,10-phenanthroline derivatives and their ability to stabilize different intramolecular telomeric G4 sequences. We evaluated ligand-induced stabilization, selectivity and specificity of ligands using Förster Resonance Energy Transfer (FRET) melting experiments and circular dichroism (CD). In addition, we assessed the cytotoxicity of ligands against two cancer cell lines (A549 and H1299) and one healthy cell line (NHDF)., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

A new family of squaramide-based anionophores (L1-L8) have been synthesised and fully characterised with the aim to investigate the effect of indolyl substituents on their anion binding and transmembrane transport properties. L1, L2, L6, and L8, bearing a 7-indolyl/indol-7-yl moiety as the substituent, were found to be the most efficient of the series in binding chloride with high stability constants. L1, L6, and L8 were also found to be the most potent anionophores of the series, able to mediate transmembrane anion transport. In particular, L6 bearing the 3,5-bis(trifluoromethyl)phenyl group was found to be the most active transporter, and its efficiency as an anionophore/anion transporter was favourably compared with that of their symmetrically-substituted squaramide analogues L9 and L10, previously reported in the literature.

The diastereoselective synthesis of two families of pyrrolopiperazine-2,6-diones is presented. These compounds were prepared by one-pot Ugi/nucleophilic substitution/N-acylation/debenzoylation/(elimination) sequences. This novel route provides straightforward access to a wide variety of pyrrolopiperazine-2,6-diones with high chemical yields and complete diastereoselectivities. The proposed synthetic strategy poses a significant improvement compared to the syntheses of pyrrolopiperazine-2,6-diones previously described, as it allows introduction of different substituents to the C4 position and the diastereoselective generation of a new stereogenic center on the bridgehead carbon (C8a).

Overcoming resistance is one of the most challenging features in current anticancer therapy. Autophagy is a cellular process that confers resistance in some advanced tumors, since it enables cancer cells to adapt to stressful situations, such as anticancer treatments. Hence, the inhibition of this cytoprotective autophagy leads to tumor cells sensitization and death. In this regard, we designed a novel potent anionophore compound that specifically targets lysosomes, called LAI-1 (late-stage autophagy inhibitor-1), and evaluated its role in blocking autophagy and its potential anticancer effects in three lung cancer cell lines from different histological subtypes. Compared to other autophagy inhibitors, such as chloroquine and 3-Methyladenine, the LAI-1 treatment induced more potent anticancer effects in all tested cancer cells. LAI-1 was able to efficiently target and deacidify lysosomes, while acidifying cytoplasmic pH. Consequently, LAI-1 efficiently blocked autophagy, indicated by the increased LC3-II/I ratio and p62/SQSTM1 levels. Moreover, no colocalization was observed between autophagosomes, marked with LC3 or p62/SQSTM1, and lysosomes, stained with LAMP-1, after the LAI-1 treatment, indicating the blockage of autophagolysosome formation. Furthermore, LAI-1 induced cell death by activating apoptosis (enhancing the cleavage of caspase-3 and PARP) or necrosis, depending on the cancer cell line. Finally, LAI-1 sensitized cancer cells to the first-line chemotherapeutic agent cisplatin. Altogether, LAI-1 is a new late-stage autophagy inhibitor that causes lysosomal dysfunction and the blockage of autophagolysosome formation, as well as potently induces cancer cell death and sensitization to conventional treatments at lower concentrations than other known autophagy inhibitors, appearing as a potential new therapeutic approach to overcome cancer resistance.

The antitumor activity of imidazolium salts is highly dependent upon their lipophilicity that can be tuned by the introduction of different hydrophobic substituents on the nitrogen atoms of the imidazolium ring of the molecule. Taking this into consideration, we have synthesized and characterized a series of tripodal imidazolium salts derived from L-valine and L-phenylalanine containing different hydrophobic groups and tested them against four cancer cell lines at physiological and acidic pH. At acidic pH (6.2) the anticancer activity of some of the tripodal compounds changes dramatically, and this parameter is crucial to control their cytotoxicity and selectivity. Moreover, several of these compounds displayed selectivity against the control healthy cell line higher than four. The transmembrane anion transport studies revealed moderate transport abilities suggesting that the observed biological activity is likely not the result of just their transport activity. The observed trends in biological activity at acidic pH agree well with the results for the CF leakage assay. These results strongly suggest that this class of compounds can serve as potential chemotherapeutic agents.

The use of arylglyoxal as starting material in Passerini and Ugi reactions affords β-ketoamides. This has allowed to study keto-enol tautomerism in these systems and assess the way in which the presence of acyloxy or aminoacyl groups bound to the C2 position affects such tautomerism, and to investigate the reactivity of both the enol and carbonyl forms. In this work we also prove the versatility of the Passerini reaction, since depending on the conditions to which the corresponding adducts are subjected different products of synthetic interest can be obtained.

Prodiginines and tambjamines are anion-selective ionophores capable of facilitating the transport of anions across the plasma membrane in mammalian cells. One of the potential applications of these anionophores is the possibility of employing them as a substitutive therapy for pathologies involving anion channels, as in cystic fibrosis. We have studied the interaction of a large anion as gluconate with three prodiginine- and two tambjamine-like compounds. Apparent dissociation constants for the chloride, iodide and gluconate complexes were estimated from iodide influx experiments in mammalian cells exposed to different extracellular anion combinations. Our experiments indicate that gluconate is not transported by the prodiginines, leaving the anionophores free to transport chloride and iodide. Conversely, gluconate would be transported to some extent by the tambjamines, competing with halides for the anionophores, and consequently reducing their flux. This might be related to the different structural features of both families of compounds. These data have important implications for the selection of impermeable anions in the analysis of the anionophore mechanism., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Herein we present the synthesis of two ligands containing two di(2-picolyl)amine (DPA) units linked by either a 1,1'-(pyridine-2,6-diyl)bis(3-ethylurea) (L1) or a 1,1'-(1,3-phenylene)bis(3-ethylurea) (L2) spacer. The corresponding binuclear Cu II and Zn II complexes were prepared and isolated. The X-ray structures of the L1 ligand and the [Cu 2 L1Cl 2 ] 2+ complex evidence an unusual cis/trans conformation of one of the urea groups stabilized by an intramolecular hydrogen bond with the nitrogen atom of the pyridyl spacer. The Cu II complexes form rather strong ternary complexes with phosphorylated anions. The [Cu 2 L1] 4+ complex presents a rather high affinity for pyrophosphate (logK 11  = 8.19 at pH 7, 25 °C), while [Cu 2 L2] 4+ stands out because of its strong binding to AMP 2- (logK 11  = 9.3 at pH 7, 25 °C). The interaction of the Cu II complexes with deoxyribonucleic acid from calf thymus (ct-DNA) was monitored using circular dichroism (CD) and luminescence spectroscopies. These studies revealed a quite strong interaction of the complexes with ct-DNA (K b  = (6.4 ± 0.7) × 10 3 for [Cu 2 L1] 4+ and K b  = (6.3 ± 1.0) × 10 3 for [Cu 2 L2] 4+ ). Competition experiments carried out in the presence of methyl green and BAPPA (N 1 ,N 3 -Bis(4-amidinophenyl)propane-1,3-diamine) as major and minor groove competitors, respectively, confirm that the interaction of both complexes with DNA takes place through the minor groove, in agreement with docking studies. The [Cu 2 L2] 4+ complex is quite efficient in promoting the cleavage of the double-stranded pUC19 plasmid DNA, by favoring the conversion of the supercoiled form to the nicked form following a hydrolytic mechanism., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

A novel class of transmembrane anion carriers, the click-tambjamines, display remarkable anionophoric activities in model liposomes and living cells. The versatility of this building block for the generation of molecular diversity offers promise to develop future drugs based on this design.

Cystic fibrosis (CF) is a genetic disease characterized by the lack of cystic fibrosis transmembrane conductance regulator (CFTR) protein expressed in epithelial cells. The resulting defective chloride and bicarbonate secretion and imbalance of the transepithelial homeostasis lead to abnormal airway surface liquid (ASL) composition and properties. The reduced ASL volume impairs ciliary beating with the consequent accumulation of sticky mucus. This situation prevents the normal mucociliary clearance, favouring the survival and proliferation of bacteria and contributing to the genesis of CF lung disease. Here, we have explored the potential of small molecules capable of facilitating the transmembrane transport of chloride and bicarbonate in order to replace the defective transport activity elicited by CFTR in CF airway epithelia. Primary human bronchial epithelial cells obtained from CF and non-CF patients were differentiated into a mucociliated epithelia in order to assess the effects of our compounds on some key properties of ASL. The treatment of these functional models with non-toxic doses of the synthetic anionophores improved the periciliary fluid composition, reducing the fluid re-absorption, correcting the ASL pH and reducing the viscosity of the mucus, thus representing promising drug candidates for CF therapy., Competing Interests: E.C., O.M. and R.Q. are inventors of a patent application (PCT/EP2019/057696) which protects the composition and use of the compounds described in the present study. The other authors declare no conflict of interest.

The synthesis of three novel families of pyrrolo[2,1- c ][1,4]benzodiazepine-5-ones is described. The compounds were prepared according to a three-step sequence, involving an Ugi reaction, building of the pyrrolo nucleus, and reduction-cyclization to the corresponding diazepine. Depending on the amine employed in the synthesis of the Ugi adducts, different unsaturation degrees could be obtained in the pyrrolo ring (saturated or with endo or exo unsaturations), a key feature determining their biological activity, as it affects the affinity of the pyrrolobenzodiazepines toward DNA and thus their cytotoxicity. This synthetic methodology represents a significant improvement with respect to those described in the literature so far, as it uses inexpensive and commercially available starting materials without needing derivatization or the use of protecting groups.

Highly active transmembrane anion transporters have demonstrated their activity against antibiotic-resistant and clinically relevant bacterial strains. This type of compound offers promise as a strategy to develop novel antibacterial agents.

Background and Purpose: Cystic fibrosis (CF) is a lethal autosomal recessive genetic disease that originates from the defective function of the CF transmembrane conductance regulator (CFTR) protein, a cAMP-dependent anion channel involved in fluid transport across epithelium. Because small synthetic transmembrane anion transporters (anionophores) can replace the biological anion transport mechanisms, independent of genetic mutations in the CFTR, such anionophores are candidates as new potential treatments for CF., Experimental Approach: In order to assess their effects on cell physiology, we have analysed the transport properties of five anionophore compounds, three prodigiosines and two tambjamines. Chloride efflux was measured in large uni-lamellar vesicles and in HEK293 cells with chloride-sensitive electrodes. Iodide influx was evaluated in FRT cells transfected with iodide-sensitive YFP. Transport of bicarbonate was assessed by changes of pH after a NH 4 + pre-pulse using the BCECF fluorescent probe. Assays were also carried out in FRT cells permanently transfected with wild type and mutant human CFTR., Key Results: All studied compounds are capable of transporting halides and bicarbonate across the cell membrane, with a higher transport capacity at acidic pH. Interestingly, the presence of these anionophores did not interfere with the activation of CFTR and did not modify the action of lumacaftor (a CFTR corrector) or ivacaftor (a CFTR potentiator)., Conclusion and Implications: These anionophores, at low concentrations, transported chloride and bicarbonate across cell membranes, without affecting CFTR function. They therefore provide promising starting points for the development of novel treatments for CF., (© 2019 The British Pharmacological Society.)

We report a series of Cu(II) and Zn(II) complexes with different ligands containing a dipicolyl unit functionalized with urea groups that may contain or not a phenylboronic acid function. These complexes were designed for the recognition of phosphorylated anions through coordination to the metal ion reinforced by hydrogen bonds involving the anion and NH groups of urea. The complexes were isolated and several adducts with pyrophosphate were characterized using Xray diffraction measurements. Coordination of one of the urea nitrogen atoms to the metal ion promoted the hydrolysis of the ligands containing 1,3-diphenylurea units, while ligands bearing 1-ethyl-3-phenylurea groups did not hydrolyze significantly at room temperature. Spectrophotometric titrations, combined with ¹H and 31 P NMR studies, were used in investigating the binding of phosphate, pyrophosphate (PPi), and nucleoside 5'-polyphosphates (AMP, ADP, ATP, CMP, and UMP). The association constants determined in aqueous solution (pH 7.0, 0.1 M MOPS) point to a stronger association with PPi, ADP, and ATP as compared with the anions containing a single phosphate unit. The [CuL⁴] 2+ complex shows important selectivity for pyrophosphate (PPi) over ADP and ATP., Competing Interests: The authors declare no conflict of interest. The founding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, and in the decision to publish the results.

Pyrrolo[2,1-c][1,4]benzodiazepines (PBDs) and other benzo-fused N-heterocycles constitute privileged structures found in numerous bioactive compounds. Thus, developing simple and selective syntheses to furnish these derivatives from easily accessible starting materials is an important and challenging goal. In this work, novel pyrrolobenzodiazepine and pyrroloquinazoline derivatives have been synthesized following a common two step synthetic strategy. This strategy involves a one-pot Ugi/cyclization sequence followed by a reduction with spontaneous thermocontrolled cyclization. The control of the temperature in this second step allows fully selective access to either pyrrolo[2,1-c][1,4]benzodiazepine-3-ones 6 or pyrrolo[2,1-b]quinazolines 7. Density functional theory (DFT) calculations have been carried out to rationalize this reactivity, identifying the kinetic and thermodynamic reaction products and offering insights into the cyclization pathways. These synthetic methodologies show the versatility of the Ugi reaction as a tool in the synthesis of heterocyclic compounds with a pseudopeptidic skeleton.

The ditopic receptor L3 [1-(2-((7-(4-(tert-butyl)benzyl)-1,4,7,10-tetraazacyclododecan-1-yl)methyl)phenyl)-3-(3-nitrophenyl)urea] containing a macrocyclic cyclen unit for Cu(ii)-coordination and a urea moiety for anion binding was designed for recognition of metal salts. The X-ray structure of [CuL3(SO 4 )] shows that the sulfate anion is involved in cooperative binding via coordination to the metal ion and hydrogen-bonding to the urea unit. This behaviour is similar to that observed for the related receptor L1 [1-(2-((bis(pyridin-2-ylmethyl)amino)methyl)phenyl)-3-(3-nitrophenyl)urea], which forms a dimeric [CuL1(μ-SO 4 )] 2 structure in the solid state. In contrast, the single crystal X-ray structure of [ZnL3(NO 3 ) 2 ] contains a 1 : 2 complex (metal : anion) where one anion coordinates to the metal and the other is hydrogen-bonded to the urea group. Spectrophotometric titrations performed for the [CuL3(OSMe 2 )] 2+ complex indicate that this system is able to bind a wide range of anions with an affinity sequence: MeCO 2 - > Cl - > H 2 PO 4 - > Br - > NO 2 - > HSO 4 - > NO 3 - . Lipophilic analogues of L1 and L3 extract CuSO 4 and CuCl 2 from water into chloroform with high selectivity over the corresponding Co(ii), Ni(ii) and Zn(ii) salts.

The ability of Cu(II) and Zn(II) complexes of the ditopic receptor H2L [1-(2-((bis(pyridin-2-ylmethyl)amino)methyl)phenyl)-3-(3-nitrophenyl)urea] for anion recognition is reported. In the presence of weakly coordinating anions such as ClO4(-), the urea group binds to the metal ion (Cu(II) or Zn(II)) through one of its nitrogen atoms. The study of the interaction of the metal complexes with a variety of anions in DMSO shows that SO4(2-) and Cl(-) bind to the complexes through a cooperative binding involving simultaneous coordination to the metal ion and different hydrogen-bonding interactions with the urea moiety, depending on the shape and size of the anion. On the contrary, single crystal X-ray diffraction studies show that anions such as NO3(-) and PhCO2(-) form 1:2 complexes (metal/anion) where one of the anions coordinates to the metal center and the second one is involved in hydrogen-bonding interaction with the urea group, which is projected away from the metal ion. Spectrophotometric titrations performed for the Cu(II) complex indicate that this system is able to bind a wide range of anions with an affinity sequence: MeCO2(-) ∼ Cl(-) (log K11 > 7) > NO2(-) > H2PO4(-) ∼ Br(-) > HSO4(-) > NO3(-) (log K11 < 2). In contrast to this, the free ligand gives much weaker interactions with these anions. In the presence of basic anions such as MeCO2(-) or F(-), competitive processes associated with the deprotonation of the coordinated N-H group of the urea moiety take place. Thus, N-coordination of the urea unit to the metal ion increases the acidity of one of its N-H groups. DFT calculations performed in DMSO solution are in agreement with both an anion-hydrogen bonding interaction and an anion-metal ion coordination collaborating in the stabilization of the metal salt complexes with tetrahedral anions.