Andreia Leite

Novel 3-hydroxy-4-pyridinonato oxidovanadium(IV) complexes to investigate structure/activity relationships.

A previous evaluation of the insulin-like activity of three 3-hydroxy-4-pyridinonato oxidovanadium(IV) complexes raised questions about structure/activity relationships, namely the influence of the hydrophilic/lipophilic balance of the complex and the capacity of the ligand to stabilize the +4 oxidation state of vanadium ion, on achieving an positive effect. To address these questions, we synthesized six new oxidovanadium(IV) complexes with variable hydrophilic/lipophilic balance, obtained by introducing different substituents on the nitrogen atom, and used two 3-hydroxy-4-pyrones as starting reagents to provide methyl and ethyl groups in the ortho position of the ring. For the new and previously reported complexes, we studied the oxidation-reduction properties and insulin-like activity in terms of inhibitory effect on Free fatty acid (FFA) release in isolated rat adipocytes. The results obtained show that only one of the complexes, Bis(3-hydroxy-1(H)-2-methyl-4-pyridonato)oxidovanadium(IV), VO(mpp)(2), exhibits a significantly greater capacity to inhibit FFA release than VOSO(4) and consequently is worthy to be considered for further studies. The establishment of structure activity relationships was not attainable but this study brings new information about the influence of some properties of the compounds on the achievement of an insulin-like effect. The results reveal that: (i) the oxidation-reduction cycles of the complexes are identical; (ii) the presence of more lipophilic substituents on the nitrogen atom does not enhance insulin-like properties; (iii) a high solubility in water proved to be not sufficient for a positive activity in inhibiting FFA release; (iv) a small molecular size may be an important property for reaching the right targets.

Investigation of the insulin-like properties of zinc(II) complexes of 3-hydroxy-4-pyridinones: Identification of a compound with glucose lowering effect in STZ-induced type I diabetic animals

Results from an investigation in an in vivo model of STZ-induced diabetic rats demonstrate that compound bis(1,2-dimethyl-3-hydroxy-4(1H)-pyridinonate)zinc(II), Zn(dmpp)(2), significantly lowers the blood glucose levels of individuals, thus showing evidence of glucose lowering activity. The compound was selected from a set of eight zinc(II) complexes of 3-hydroxy-4-pyridinones with diverse lipophilicity that were prepared and characterized in our laboratory. Assessment of insulin-like activity of the complexes was firstly performed in vitro by measuring the inhibition of FFA release in isolated rat adipocytes. The results indicate that compounds bis(2-methyl-3-hydroxy-4-pyridinonate)zinc(II), Zn(mpp)(2) and Zn(dmpp)(2) display significantly higher activity than that of the respective positive control thus suggesting its selection for in vivo tests. Safety evaluation of the active zinc(II) compounds was performed in freshly isolated rat hepatocytes. The results support that cell viability is not significantly different from the control set after 1 and 2h of incubation with both zinc(II) complexes.

Effect of tris(3-hydroxy-4-pyridinonate) iron(III) complexes on iron uptake and storage in soybean (Glycine max L.).

Iron deficiency chlorosis (IDC) is a serious environmental problem affecting the growth of several crops in the world. The application of synthetic Fe(III) chelates is still one of the most common measures to correct IDC and the search for more effective Fe chelates remains an important issue. Herein, we propose a tris(3-hydroxy-4-pyridinonate) iron(III) complex, Fe(mpp)3, as an IDC corrector. Different morphological, biochemical and molecular parameters were assessed as a first step towards understanding its mode of action, compared with that of the commercial fertilizer FeEDDHA. Plants treated with the pyridinone iron(III) complexes were significantly greener and had increased biomass. The total Fe content was measured using ICP-OES and plants treated with pyridinone complexes accumulated about 50% more Fe than those treated with the commercial chelate. In particular, plants supplied with compound Fe(mpp)3 were able to translocate iron from the roots to the shoots and did not elicit the expression of the Fe-stress related genes FRO2 and IRT1. These results suggest that 3,4-HPO iron(III) chelates could be a potential new class of plant fertilizing agents.

1,3-Dipolar cycloadditions with meso-tetraarylchlorins – site selectivity and mixed bisadducts

The 1,3-dipolar cycloaddition reaction of meso-tetraarylporphyrins with nitrones gives isoxazolidine-fused chlorins. Depending on the substitution pattern on the meso-aryl groups and the nitrone, the chlorins can be obtained in high yields (up to 91%). Bacteriochlorin-type bisadducts are also obtained, although in low yield, from the reaction of meso-tetrakis(pentafluorophenyl)porphyrin with N-methyl, N-cyclohexyl or N-benzyl nitrone. The structure of a bis(N-benzyl isoxazolidine-fused) bacteriochlorin was determined by single-crystal X-ray diffraction and rationalized by DFT calculations. To further explore the nature of site selectivity in the formation of bisadducts, isomeric mixed bacteriochlorins and isobacteriochlorins were synthesized by two complementary routes that involved the sequential addition of two types of 1,3-dipoles to the porphyrin macrocycle: a nitrone and an azomethine ylide. The photophysical properties of the mixed bacteriochlorins were evaluated and the results were compared with the reported data for other meso-tetraarylbacteriochlorins.

Tuning the Anti(myco)bacterial Activity of 3-Hydroxy-4-pyridinone Chelators through Fluorophores

Controlling the sources of Fe available to pathogens is one of the possible strategies that can be successfully used by novel antibacterial drugs. We focused our interest on the design of chelators to address Mycobacterium avium infections. Taking into account the molecular structure of mycobacterial siderophores and considering that new chelators must be able to compete for Fe(III), we selected ligands of the 3-hydroxy-4-pyridinone class to achieve our purpose. After choosing the type of chelating unit it was also our objective to design chelators that could be monitored inside the cell and for that reason we designed chelators that could be functionalized with fluorophores. We didn’t realize at the time that the incorporation a fluorophore, to allow spectroscopic detection, would be so relevant for the antimycobacterial effect or to determine the affinity of the chelators towards biological membranes. From a biophysical perspective, this is a fascinating illustration of the fact that functionalization of a molecule with a particular label may lead to a change in its membrane permeation properties and result in a dramatic change in biological activity. For that reason we believe it is interesting to give a critical account of our entire work in this area and justify the statement “to label means to change”. New perspectives regarding combined therapeutic approaches and the use of rhodamine B conjugates to target closely related problems such as bacterial resistance and biofilm production are also discussed.

EPR and 51V NMR studies of prospective anti-diabetic bis(3-hydroxy-4-pyridinonato)oxidovanadium(IV) complexes in aqueous solution and liposome suspensions

We report EPR and 51V NMR characterization of vanadium(IV/V) species originating from bis(3-hydroxy-4-pyridinonato)oxidovanadium(IV) complexes in aqueous solution at pH 7.4, under aerobic conditions and along time. Since complexes possess variable hydrophilic/lipophilic balance and consequently different solubilities in water we also performed a parallel study in POPC liposome suspensions. For less soluble complexes we used DMSO as a co-solvent. We identified three groups of complexes with different solubilities in water. Compounds 1 and 2 are soluble enough to produce 1.5 mM solutions in MOPS buffer. Compounds 3 and 4 exhibit intermediate solubility but studies in MOPS buffer and POPC liposomes demonstrate that solubilisation is considerably enhanced in liposome suspensions. Compounds 5 and 6 exhibit low water solubility, which is not sufficiently enhanced in liposome suspensions thus constraining the use of DMSO as a co-solvent. Analysis of the EPR spectra of bis(3-hydroxy-4-pyridinonato)oxidovanadium(IV) complexes shows that upon dissolution a single species is present in solution, [VOL2]. In the presence of air [VOL2] is oxidized to three species, [VO2L2]−, [VO2L] and V1 (H2VO4−) as characterized by the 51V NMR spectra. Addition of sodium ascorbate reduces the V(V) species to the original [VOL2] complex. The results demonstrate that the use of DMSO lowers the rate of oxidation of the [VOL2] complex. For DMSO percentages higher than 10% two distinct EPR signals are observable and EPR spin-Hamiltonian parameters confirm the presence of [VOL2] in two different solvation environments, which confirms the participation of DMSO in the first solvation sphere of the complex.

Synthesis and coordination studies of 5-(4′-carboxyphenyl)-10,15,20-tris(pentafluorophenyl)porphyrin and its pyrrolidine-fused chlorin derivative

The introduction of a carboxylate function into porphyrins allows a variety of modifications, including coordination and conjugation, which are central to enhance the efficiency of macrocycles in photonic materials and biological applications. Herein, a synthetic strategy to obtain 5-(4′-carboxyphenyl)-10,15,20-tris(pentafluorophenyl)porphyrin and its pyrrolidine-fused chlorin derivative was developed by the 1,3-dipolar cycloaddition of a carbomethoxyphenyl substituted porphyrin with an azomethine ylide, followed by hydrolysis under thermal acidic conditions. The 1,3-dipolar cycloaddition of the carbomethoxyphenyl porphyrin with N-methyl nitrone was also performed to give an isomeric mixture of isoxazolidine-fused chlorins, revealing lower selectivity and lower yields; in addition, retrocycloaddition of the isoxazolidine-fused chlorins was observed under the hydrolysis conditions. The resulting carboxyphenyl macrocycles were characterized using 1H and 19F NMR, ESI-MS and SC-XRD for 5-(4′-carboxyphenyl)-10,15,20-tris(pentafluorophenyl)porphyrin. In order to study the influence of coordination to a metal ion on the electronic properties of carboxyphenyl substituted porphyrins, a series of metal complexes of 5-(4′-carboxyphenyl)-10,15,20-tris(pentafluorophenyl)porphyrin and its pyrrolidine-fused chlorin derivative were synthesized by microwave-mediated metallation with Fe(III), Cu(II) and Zn(II) salts. EPR spectroscopy was particularly relevant to the characterization of the Cu(II) complexes of both macrocycles and to study the coordination chemistry of these ligands with Cu(II) ions.

Uncovering novel 3-hydroxy-4-pyridinone metal ion complexes with potential anti-inflammatory properties.

Ligands of the 3-hydroxy-4-pyridinone (3,4-HPO) type, with one (Hmpp) or two methyl groups (Hdmpp), have been reported to possess biomedical, chemical and analytical applications. In this first screening study aiming to uncover new promising agents to mitigate the oxidative damage highly present in several metabolic disorders, such as diabetes mellitus, we assessed the potential of twelve 3,4-HPO metal ion complexes to modulate oxidative burst in human neutrophils. Metal ion 3,4-HPO complexes of Ni, Fe, V, Co, Cu and Zn were synthesized and tested up to 15μM. Among all the compounds, [Cu(mpp)2] and [Cu(dmpp)2] exhibited the highest scavenging capacity against superoxide radical (O2(-)) (IC50=0.36±0.07 and 0.30±0.06μM, respectively) and against hypochlorous acid (HOCl) (IC50=0.6±0.3 and 0.4±0.1μM, respectively). In the particular case of O2(-), [Fe(mpp)3] and [Fe(dmpp)3] (both at 15μM) presented 35% and 22% of inhibition, respectively, while all the other compounds were neither able to scavenge O2(-) nor stimulate its production. Regarding the scavenging capacity against hydrogen peroxide (H2O2), all the compounds showed low efficiency (from 6-39%). Finally, with exception of [VO(mpp)2] and [VO(dmpp)2], all compounds exhibited scavenging activity against HOCl (39-81%) and the most efficient compounds were Cu(II) and Zn(II) complexes. Thus, these preliminary results uncover promising new metal ion complexes, inhibitors of neutrophils oxidative burst, with potential anti-inflammatory properties, which may be seen as an useful strategy for further studies in the treatment of a number of diseases where oxidative damage is a serious issue.