Bis(picolinato) complexes of vanadium and zinc as potential antidiabetic agents: synthesis, structural elucidation and in vitro insulin-mimetic activity study

Vanadium(V) compounds with 3,5-difluoropicolinic acid (HpicFF) and 3-hydroxypicolinic acid (H2hypic) and zinc(II) compounds with HpicFF, H2hypic and 4-methylhydroxypicolinic acid (HpicCH2OH) in the presence or absence of pyridine (py), 4-(dimethylamino)pyridine (DMAP), and 1,10-phenanthroline (phen) have been synthesized and characterized. The crystal structures of NH4[VO2(picFF)2] 1.6H2O (3 1.6H2O), NH4[VO2(hypic)2] H2O (4 H2O), [Zn(picFF)2(H2O)2] (5), [Zn(picFF)2(py)2] py (6 py), [Zn(picFF)2(DMAP)2] =3H2O (7 =3H2O), [Zn(picFF)2(phen)] 2CHCl3 (8 2CHCl3), [Zn(Hhypic)2(MeOH)2] (10), [Zn(Hhypic)2(DMAP)(H2O)] (11) and [Zn(Hhypic)2(phen)] (12) were determined by X-ray crystallography. The spatial arrangement of the vanadium(V) complex 3 possesses carboxylate oxygen atoms in a mutual trans orientation while crystal structure 4 represents the first crystallographic evidence for the formation of an isomer with two picolinato nitrogen atoms in a mutual trans position. Two different spatial arrangements were found in the zinc(II) complexes with trans (5, 10 and 11) and cis (6, 7, 8 and 12) octahedral geometry. Insulin-mimetic activity of the selected VO, VO2 and Zn II complexes was studied by in vitro inhibition of the free fatty acid (FFA) release from isolated rat adipocytes treated with epinephrine. All the studied metal complexes showed insulin-mimetic activity. Vanadium complexes 1–4 exhibit activities similar to VOSO4 with the Hhypic complexes being more active than the picFF ones. The zinc complexes also exhibited some insulin-mimetic activity, with the Hhypic complex (9) being more active than the other two, however, the insulin-mimetic activity of these complexes did not show more potent activity than ZnSO4.


Introduction
According to the estimations of the World Health Organization high blood glucose level represents a risk, that is globally the third most important factor for premature mortality, after high blood pressure and the use of tobacco. 1 The worldwide number of patients suffering from diabetes mellitus (DM) has been estimated to be 415 million in 2015 representing B8.8% of adult population and is expected to increase to 642 million by 2040 when B10% of adult population will be affected. 1 DM is a metabolic disorder characterized by abnormal glucose levels with a tendency to hyperglycaemia that occurs due to the decrease of insulin function and/or level. Undiagnosed hyperglycaemia represents a severe risk since it can lead to serious damage of several body systems. DM is generally classified as type 1, type 2, and gestational diabetes. Type 1 DM is a consequence of an absolute lack of insulin due to autoimmune destruction of pancreatic b cells and as such is not a lifestylerelated condition. Type 2 DM representing the majority of diabetic patients (B90%) is caused by insulin resistance or degradation of secreted insulin and is a life-style related disease. Ageing populations and modern lifestyle, i.e. reduced physical activity, dietary changes, obesity, increasing urbanization, and increased stress are believed to be the main reason for the prevalence of DM. 1,2 The treatment for type 1 DM is subcutaneous injections of insulin, since oral administration of insulin is not efficient. Primary treatments for type 2 DM are dietary control and regular exercise and after the development of the disease oral hypoglycaemic drugs are used. However, they are often associated with undesirable side effects or a drop in efficacy is observed after prolonged use 3 and patients have to start using insulin injections.
Zinc and vanadium compounds possess various biological roles in living organisms [3][4][5] and are thus among the suitable candidates in the search for novel orally active chemotherapeutics as an alternative for the existing hypoglycaemic drugs. Zinc is essential in growth and development and plays an important role in hundreds of metalloenzymes and in thousands of metalloproteins. 3 Over 75% of the zinc content in the human body is located in skeletal muscle and bone tissue. 4 Zinc deficiency can lead to several diseases including diabetes and the recommended dietary allowance is 8 mg kg À1 for women and 11 mg kg À1 for men. 4 Zinc compounds exhibit a wide variety of pharmacological properties, such as anticancer, 6 antioxidant, 7 antibacterial and antimicrobial activities, 8 as well as effects in the treatment of Alzheimer disease. 9 Biological activities of vanadium compounds, e.g., as antitumor, antiparasitic (Chagas disease), and osteogenic agents have also been described. 10 The first report regarding the influence of vanadium on diabetic patients dates back to 1899 when Lyonett and Martin observed reduced levels of glucose in the urine of patients using sodium metavanadate. 11,12 In 1980 and in 1992 it was found that vanadium and zinc salts, respectively, mimic the action of insulin. 3,12 Those findings gave impetus to the extensive research on zinc and vanadium coordination compounds where organic carriers were found to substantially increase the bioavailability and reduce the administrated doses needed. 13 Several classes of very promising complexes consist of Zn and V species among which the ethylmaltolato vanadium complex has entered the phase IIa clinical trials. Further studies on this compound as an antidiabetic agent were abandoned due to renal problems, however, it is currently studied as a therapeutic for the prevention, stoppage, and reparation of secondary tissue injuries. 13f Picolinato complexes were also found to possess strong antidiabetic action. Several derivatives of the picolinato scaffold have been prepared, such as 6-methylpicolinate, 6-ethylpicolinate, halopicolinates, 3-hydroxypicolinate, and also species formed by monoesters and amides of 2,5-dipicolinic acid, all exhibiting insulin-mimetic properties. 3,14 Additionally, complexes with several other ligand systems have been studied during the last few years 15 among which complexes with antioxidants such as allixin, S-allixin-Nmethyl and pyrrolidine-N-dithiocarbamato ligands were found to be particularly active. 16 All these recent efforts have been oriented towards the synthesis of new compounds with high bioavailability and low toxicity since the main obstacle with the ethylmaltolato vanadium complex was the high dose necessary to achieve the therapeutic effect. 13f Recently it has been established on selected zinc complexes by noninvasive methods for analyzing the biodistribution of complexes that high antidiabetic activity correlates with a long retention time of complexes in the blood. 17 According to Rehder, 10a ligands containing the pyrone, the pyridinone or the pyridine-carboxylate scaffold are seen as the best candidates with a sufficiently long physiological half-life to recombine with metal in the body and thus to decrease potential toxicity.
Herein, we report ten vanadium and zinc compounds formed by 3,5-difluoropicolinic acid (HpicFF), 3-hydroxypicolinic acid (H 2 hypic) and 4-(hydroxymethyl)picolinic acid (HpicCH 2 OH). Structural characterization of nine complexes was carried out by X-ray diffraction analysis. Insulin-mimetic activity of the selected complexes was studied by an in vitro inhibition of the FFA release from isolated rat adipocytes treated with epinephrine.

X-ray structure determination
Crystals suitable for X-ray analysis were obtained from compound solutions or filtered reaction mixtures at room temperature by slow evaporation of the solvents over a few days. Single-crystal X-ray diffraction data were collected on either a Nonius Kappa CCD diffractometer or an Agilent Technologies SuperNova Dual diffractometer with an Atlas detector using monochromated Mo-Ka radiation (l = 0.71073 Å) or Cu-Ka radiation (l = 1.54184 Å) at room temperature or at 150 K. The data were processed using DENZO or CrysAlis Pro. 20 The structures were solved by direct methods using the program SHELXS-97 or SIR97 and refined on F 2 using full-matrix least-squares procedures with SHELX2014. 21 All non-hydrogen atoms were refined anisotropically. The hydrogen atoms on carbon atoms and OH groups were treated as riding atoms in geometrically idealized positions. Hydrogen atoms attached to the nitrogen atom of ammonium cations, water and methanol oxygen atoms were readily located from difference Fourier maps and were refined fixing the bond lengths and isotropic temperature factors as U iso (H) = 1.5U eq (N,O) unless otherwise noted. In 3Á1.6H 2 O water oxygen atoms O8, O9 and O10 were refined with a 0.25 : 0.20 : 0.14 occupancy ratio and ISOR instruction was used on them for proper refinement. Hydrogen atoms on the O7-O10 atoms in 3Á1.6H 2 O and on the O13 and O14 atoms in 7Á2/3H 2 O were not found in the difference Fourier maps and were not included in the refinement. In 8Á2CHCl 3 the chloroform solvate molecule is disordered over two positions in a refined ratio of 0.60 : 0.40, and DFIX, SADI, SIMU and DELU instructions were used on the solvate molecule for the appropriate refinement. Crystallographic data are listed in Table S1 (ESI †).

In vitro insulin-mimetic activity
The procedure for the in vitro test was adopted from the literature. 22 Male Wistar rats were sacrificed under anesthesia with ether. The adipose tissues were removed, chopped with scissors, and digested with collagenase for 60 min at 37 1C in Krebs-Ringer bicarbonate buffer (120 mM NaCl, 1.27 mM CaCl 2 , 1.2 mM MgSO 4 , 4.75 mM KCl, 1.2 mM KH 2 PO 4 , 24 mM NaHCO 3 : pH 7.4), containing 2% BSA (bovine serum albumin). The obtained adipocytes were then separated from undigested tissues by filtration through nylon mesh and washed three times. The metal complexes which were dissolved in ultrapure water at 500 mM (final conc.) and glucose solution (final conc.; 5 mM) was separately added to the isolated adipocytes, and the resulting suspension was pre-incubated at 37 1C for 30 min. Finally, epinephrine solution (final conc.; 10 mM) was added to the above suspension, and the resulting solutions were incubated at 37 1C for 180 min. The reactions were stopped by soaking in ice water, and the mixtures were centrifuged at 3000 rpm for 10 min. FFA (free fatty acids) levels in the outer solution of the cells were determined using an FFA kit (NEFA C-test WAKO, Wako Pure Chemicals).
All animal experiments in the present study were approved by the Experimental Animal Research of Kyoto Pharmaceutical University (KPU) and were performed according to the Guideline for Animal Experimentation of KPU.

Results and discussion
Synthesis and IR spectroscopy Oxidovanadium(IV), V IV O, compounds 1 and 2 were synthesized from VOSO 4 Á5H 2 O and HpicFF or H 2 hypic as described previously. 18 Dioxidovanadate(V) and V V O 2 , compounds were prepared from ammonium metavanadate and the corresponding picolinic acid analogues, HpicFF (3) or H 2 hypic (4), in a 1 : 2 ratio in methanol or water/methanol, respectively. Zinc(II) bis(3,5-difluoropicolinato) and bis(3-hydroxypicolinato) complexes were prepared in two steps. In the first step the white powder of the zinc bis(3,5-difluoropicolinato) compound (5) and the zinc bis(3-hydroxypicolinato) compound (9) were prepared in a water/methanol mixture or in water from zinc acetate dihydrate and HpicFF or H 2 hypic acid in a 1 : 2 ratio. In the second step 5 or 9 was suspended in an organic solvent and then the neutral N-donor ligands py (6Ápy), DMAP (7Á 2 = 3 H 2 O, 11) or phen (8ÁCHCl 3 , 12) were added. The N-donor ligands were selected in order to make a structural comparison with previously published zinc complexes with the 5-cyanopicolinato (picCN) organic carrier where a cis-octahedral arrangement was observed in py and 4-aminopyridine (4apy) complexes and also a square-pyramidal geometry was observed in the 4apy complex in contrast to the [Zn(picCN) 2 (H 2 O) 2 ] complex possessing a trans arrangement of the ligands in the coordination sphere. 23 The reaction of 5 with py in toluene/methanol yielded crystals 6Ápy, however, when these crystals were dried in air, the pyridine was lost and the complex decomposed transforming into the compound 5 as was observed also in the analogous system with the picCN ligand. 23 The reaction of 9 with DMAP yielded complex 11 even though the ratio 1 : 2 was used. A small amount of crystals of 10 with methanol molecules bonded to the metal center were obtained during the unsuccessful synthesis of 11 in methanol. For comparison, the reaction of [Zn(picCN) 2 (H 2 O) 2 ] with py in methanol yielded only the starting complex, the substitution of the water by py or MeOH as the ligand was not observed. 23 The zinc bis(4-(hydroxymethyl)picolinato) compound 13Á0.5H 2 O was prepared in methanol in an analogous way as were compounds 5 and 9. Although several attempts were made to structurally characterize the zinc bis(4-(hydroxymethyl)picolinato) species in the presence or absence of neutral N-donors we were not able to obtain crystals of 13Á0.5H 2 O or other adducts suitable for X-ray structural determination. IR spectroscopy of 3 and 4 shows the presence of N-H stretching vibrations of the ammonium cation in the range between 3176 and 3052 cm À1 in both compounds and stretching bands of the VO 2 + cation were observed between 897 and 853 cm À1 . The IR spectra of vanadium compounds 3 and 4 and zinc compounds 5-9 and 11-13 show antisymmetric, n asym (CO 2 ), and symmetric, n sym (CO 2 ), stretching vibrations of the carboxylato group in the ranges of 1666-1598 cm À1 and 1392-1300 cm À1 , respectively. The difference D, a useful parameter revealing the possible coordination mode of the carboxylato ligands, has values greater than 200 cm À1 for each of these complexes indicating a monodentate coordination of the carboxylate group. 24 Structure of the complexes  Fig. 1 and 3, and selected bond distances and angles are summarized in Tables 1 and 2. Compound 3Á1.6H 2 O crystallizes in the tetragonal space group P4 2 /n and the asymmetric unit contains a complex dioxidovanadate(V) anion, an ammonium cation, one water molecule with full occupancy and 0.6 water molecule disordered over three positions. Compound 4 crystallizes in the monoclinic space group C2/c and the asymmetric unit contains one half of the complex anion, one half of the ammonium cation and one half of the water molecule due to the two-fold rotation axis.
Ammonium cations and complex anions in the crystal structure of 3Á1.6H 2 O are connected into the infinite belt along the c axis. The ammonium cation is connected to three adjacent [VO 2 (picFF) 2 ] À complex anions where the hydrogenbond acceptors are the O1 carboxyl atom of one complex anion, the O3 carboxyl atom of the second complex anion and the O5 oxido oxygen atom of the VO 2 + unit of the third complex anion.
The fourth hydrogen-bonding of ammonium cation is with the O7 atom of the water molecule ( Fig. S1 and Table S2, ESI †). Additionally, C6-H6Á Á ÁO2 and C12-H12Á Á ÁO4 interactions between pyridyl rings and carbonyl moieties stabilize the belt formation. Belts are connected to the supramolecular structure via C4-H4Á Á ÁO5 interactions between the pyridyl rings and oxido atoms and C10-H10Á Á ÁO4 interactions between the pyridyl rings and carbonyl moieties (Table S2, ESI †). Channels along the c axis are occupied by water molecules (Fig. 2). There are no significant pÁ Á Áp interactions present. Vanadium atom in 4 is also six-coordinated having an octahedral geometry similar to 3 with a cis arrangement of the oxido oxygen atoms of      Ammonium cations, complex anions and solvate water in the crystal structure of 4ÁH 2 O form hydrogen-bonded infinite 2D layers along the ab plane (Fig. S2, ESI †). The ammonium cation is connected to three adjacent [VO 2 (Hhypic) 2 ] À complex anions through two bifurcated hydrogen-bonds N2-H2AÁ Á ÁO1 and N2-H2AÁ Á ÁO2 with carboxylato units of two complex anions and through two N2-H2BÁ Á ÁO4 hydrogen bonds with both oxido atoms of VO 2 + unit of the third complex anion (Table S3, ESI †). The solvate water molecule is involved in O5-H5AÁ Á ÁO4 hydrogen-bonding with the oxido atoms of two adjacent VO 2 + units. The C6-H6Á Á ÁO2 interaction between the pyridyl ring and carbonyl oxygen atom of the adjacent complex anion further stabilize the layer formation. Layers are connected into the supramolecular structure along the c axis via N2-H2BÁ Á ÁO3 interactions between the ammonium cation and the hydroxyl group as well as C4-H4Á Á ÁO4 interactions between the pyridyl ring and the oxido atoms of the VO 2 + unit and by p-p interactions with a Cg3Á Á ÁCg3 centroid-to-centroid distance of 3.736(3) Å ( Fig. S3 and  3 ). Crystals of compounds 5-8 were grown from the reaction mixtures after a few days of slow evaporation of the solvents. In air the crystals of 8Á2CHCl 3 gradually lose solvate molecules and transform into solvate-free 8. Diagrams of the structures of complexes 5-8 are shown in Fig. 4-7, and selected bond distances and angles are summarized in Tables 3-6.
The complex 5 crystallizes in the triclinic space group P% 1. The asymmetric unit contains one half of the complex with the zinc atom sitting on the inversion center. The zinc atom is octahedrally coordinated with two bidentate picFF ligands in the equatorial position in a trans arrangement and with two water molecules in the axial positions. The distance between the zinc and carboxyl oxygen (Zn1-O1 = 2.0541(12) Å) is shorter than the bond between zinc and water (Zn1-O3 = 2.2092(14) Å). The Zn1-N1 distance is 2.1020(14) Å. The angles between the trans oriented atoms are 1801 due to the center of symmetry. The chelate angle of the picFF ligand O1-Zn1-N1 is 79.49(5)1, the angle between the water and carboxyl oxygen atom O1-Zn1-O3 is 91.16(5)1, and the angle between the water and picolinato nitrogen atom O3-Zn1-N1 is 89.93(5)1.
The crystal structure of 5 is stabilized by hydrogen bonding between a coordinated water molecule acting as a hydrogen bond donor and a carbonyl atom acting as a hydrogen bond

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acceptor forming centrosymmetric hydrogen bonding with a R 2 4 (8) ring motif. The water molecule is also involved in hydrogen bonding with the fluorine atom of the adjacent molecule forming a R 2 1 (6) ring motif. These interactions form hydrogenbonded layers parallel to the ab plane ( Fig. 4 (b) and Table S5, ESI †). The layers are connected into the supramolecular structure along the c axis via C-HÁ Á ÁF interactions and p-p interactions with a Cg3Á Á ÁCg3 centroid-to-centroid distance of 3.6302(10) Å (Tables S5 and S6,  The supramolecular structure of 6Ápy is stabilized by various C-HÁ Á ÁO and C14-H14Á Á ÁF2 hydrogen bonding interactions forming layers along the bc axis ( Fig. S4 and Table S7, ESI †). Pyridine solvate molecules are part of these layers and are connected through C15-H15Á Á ÁN5 hydrogen bonding and pÁ Á Áp interactions with a Cg4Á Á ÁCg7 centroid-to-centroid distance of 4.170(3) Å. The layers are connected into the supramolecular structure through pÁ Á Áp interactions with a Cg3Á Á ÁCg5 centroidto-centroid distance of 4.0362(17) Å between the picolinate and pyridine moieties that are part of adjacent complexes (Table S8, ESI †). Symmetry code: (i) Àx + 1, Ày + 1, Àz + 1.  . For clarity only one out of three crystallographically independent molecules is shown. Thermal ellipsoids are shown with 30% probability level at 293 K.  The compound 7Á 2 = 3 H 2 O crystallizes as a hydrate in the orthorhombic space group Pbca with the formula [Zn(picFF) 2 (DMAP) 2 ]Á 2 = 3 H 2 O. The asymmetric unit contains three crystallographically independent complex molecules and two water solvate molecules. All three independent complexes have a distorted octahedral geometry, with a cis-octahedral arrangement of DMAP ligands on the zinc central atom analogous to 6Ápy. The geometric parameters for all three complexes are very similar (Table 5) In the crystal structure of 8Á2CHCl 3 the complex molecules are connected into the infinite belt along the c axis through C4-H4Á Á ÁO2 interactions and chloroform solvate molecules are connected to the complex molecules through bifurcated C13-H13Á Á ÁO1/O2 interactions with the carboxylato groups. The pÁ Á Áp stacking interactions between adjacent phen moieties with a Cg6Á Á ÁCg17 centroid-to-centroid distance of 3.706(3) Å connect the belts into a three-dimensional structure (Fig. S6 (11) and [Zn(Hhypic) 2 (phen)] (12). A small amount of the crystals of 10 was obtained during the unsuccessful synthesis of 11 when using methanol as the solvent. The complex 10 crystallizes in the monoclinic space group C2/c. The asymmetric unit contains one half of the complex with the zinc atom sitting on the inversion center. The zinc atom is octahedrally coordinated with two bidentate Hhypic ligands in the equatorial position in trans arrangement and with two methanol molecules in the axial positions (Fig. 8). The distance between zinc and the carboxyl oxygen (Zn1-O1 = 2.0750(10) Å) is shorter than the bond between zinc and the methanol oxygen atom (Zn1-O4 = 2.1930(11) Å). The Zn1-N1 distance is 2.0765(14) Å. The chelate angle of the Hhypic ligand O1-Zn1-N1 is 80.73(4)1, the angle between the methanol and carboxyl oxygen atoms O1-Zn1-O4 is 93.85(4)1, and the angle between the methanol oxygen atom and the picolinato nitrogen atom O4-Zn1-N1 is 90.03(5)1 ( Table 7).
The crystal structure of 10 is stabilized by hydrogen bonding between a coordinated methanol molecule acting as a hydrogen bond donor and a carbonyl atom of an adjacent complex acting  (4) Angle (1) as a hydrogen bond acceptor forming centrosymmetric hydrogen bonding with a R 2 2 (12) ring motif and a chain along the b axis is formed. These chains are further connected into an infinite layer along the bc plane through the formation of C-HÁ Á ÁO hydrogen bonding, with the picolinic moiety as a hydrogen bond donor and the methanol molecule as a hydrogen bond acceptor ( Fig. S7 and Table S9, ESI †). There are no significant pÁ Á Áp interactions present.
The compound 11 crystallizes in the orthorhombic space group Pnna with the formula [Zn(Hhypic) 2 (DMAP)(H 2 O)]. The asymmetric unit contains one half of the complex with a zinc atom, DMAP and water sitting on the two-fold axis. The zinc atom is octahedrally coordinated with two bidentate Hhypic ligands in the equatorial position in trans arrangement and with one DMAP and one water molecule in the axial positions (Fig. 9). The distance between the zinc and carboxyl oxygen (Zn1-O1 = 2.1028(11) Å) is shorter than the bond between zinc and the water oxygen atom (Zn1-O4 = 2.1578(19) Å). The Zn1-N1 distance is 2.1350(14) Å and the DMAP is coordinated to the zinc with a Zn1-N2 distance of 2.161(2) Å. The chelate angle of the Hhypic ligand O1-Zn1-N1 is 79.18(5)1, the angle between the water and carboxyl oxygen atoms O1-Zn1-O4 is 88.62(3)1, and the angle between the water oxygen atom and picolinato nitrogen atom O4-Zn1-N1 is 86.21(3)1 ( Table 8).

Structural comparison
Valuable structural information is usually obtained by a solidstate X-ray diffraction study, however, when the compounds are introduced into a biological system certain chemical and biological changes can occur. EPR spectroscopy was found to be a powerful tool for the study of the solution structure and (bio)transformations of vanadium(IV) species. 25 Lately, we have been studying V IV O compounds with picolinic acid and pyrazine derivatives concerning their structures and interactions with serum proteins. 18 For compounds 1 and 2 three isomers were observed by EPR in an organic solvent or in aqueous solution. The H 2 hypic was found to be the strongest ligand and the HpicFF the weakest one and thus different biotransformations at physiological pH in the presence of apo-transferrin and human serum albumin were observed. 18 For the study of pharmacological activity all such generated species have to be taken into account and it is a challenge to obtain the observed solution-state species also in the solid state. In this respect, we were able to establish the first crystallographic evidence for the formation of an isomer of a vanadyl bis(picolinato) complex with the two pyridine nitrogen atoms in a mutual trans position previously observed only in the solution-state; the known solid-state structures possess a common arrangement with the two equatorial pyridine nitrogen atoms in a mutual cis position. 18 We have extended our research on the diverse structural characteristics and supramolecular topologies of potential antidiabetic vanadium species 23,26 also on vanadium(V) and zinc(II) compounds in order to study the flexibility of the coordination sphere and possible spatial arrangements in these species as well.
In  23 and in two compounds with 5-cyanopicolinato and 3-aminopyrazine-2-carboxylato ligands 23 possessing nitrogen atoms coordinated to the V(V) metal center trans to the oxido oxygen atoms as observed also in 3. The only report on structure (II) is the prototypical complex NH 4 [VO 2 (pic) 2 ]ÁH 2 O prepared by Melchior et al. 29 with one picolinato nitrogen atom coordinated to the V metal center trans to the oxido oxygen and the other picolinato nitrogen atom coordinated in a trans position to the carboxyl oxygen atom. There were no reports on isomer (III) in the CSD and the 3-hydroxypicolinato complex 4 is thus the first crystallographic evidence for the formation of an isomer of a V V O 2 bis(picolinato) complex with the two pyridine nitrogen atoms in a mutual trans position. It has to be pointed out that 3-hydroxypicolinic acid possesses diverse ligation modes and can act Fig. 11 Asymmetric unit of [Zn(Hhypic) 2 (phen)] (12). Thermal ellipsoids are shown with 50% probability level at 150 K.  19 and also in 5, 10, and 11. Structure B with a cis arrangement was found in six entries where the neutral ligand L is imidazole, pyridine, 4-aminopyridine or phenanthroline and was found also in 6-8 and 12. 23,33 For each additional two cis spatial arrangements for the octahedral environment was reported only one example, cis-[Zn(pic) 2 39 exhibit the trigonalbipyramidal geometry E with the nitrogen donor atoms in an axial position and one complex [Zn(Hpycc) 2 (H 2 O)] (H 2 pycc = pyridine-5-carboxylato-2-carboxylic acid) 40 exhibits the trigonalbipyramidal geometry F with the carboxylic oxygen atoms in an axial position. The square-pyramidal geometry G was reported only in two complexes [Zn(bupyc) 2 (H 2 O)] (Hbupyc = 5-butylpyridine-2carboxylic acid) 41 and [Zn(picCN) 2 (4apy)] (HpicCN = 5-cyanopicolinic acid, 4apy = 4-aminopyridine). 23 In previously published zinc complexes with the 5-cianopicolinato (picCN) organic carrier in the presence of the 4apy ligand in a 1 : 2 ratio the cis-octahedral complex [Zn(picCN) 2 (4apy) 2 ] was obtained from the apolar solvent system and the square-pyramidal complex [Zn(picCN) 2 (4apy)] was obtained from methanol. 23 On the other hand, the reaction of the starting compound [Zn(Hypic) 2 (H 2 O) 2 ] (9) with two equivalents of DMAP in methanol gave small amount of crystals 10 with methanol molecules bonded to metal center and the same reaction in chloroform/toluene/methanol solvent mixture gave complex 11 as the product. These findings show the rich structural diversity and an important influence of the solvent system.

In vitro insulin-mimetic activity
The inhibitory effects on free fatty acid (FFA) release from the isolated rat adipocytes treated with epinephrine were measured on selected vanadium and zinc complexes with sufficient solubility in water in order to determine the insulin-like activities. For these in vitro experiments seven vanadium and zinc complexes with 3,5-difluoropicolinato, 3-hydroxypicolinato and 4-(hydroxymethyl)picolinato ligands were found to have suitable solubility in water. Two of them were the vanadium(IV) complexes [VO(picFF) 2 (H 2 O)] (1) and Na[VO(Hhypic) 3 ]ÁH 2 O (2) which were synthesized as described previously 18 and two were their vanadium(V) analogues (3,4). The zinc(II) complexes selected were 5, 9 and 13. The effects of the vanadium and zinc complexes on the FFA release compared with those of the inorganic compounds VS (VOSO 4 ) and ZS (ZnSO 4 ) are summarized in Fig. 12 and 13, respectively. All the studied complexes show insulin-mimetic activity. Vanadium complexes 1-4 exhibit activities similar to VOSO 4 with Hhypic complexes (2,4) being more active than the picFF ones. According to previous findings 13a V IV O compounds generally exhibit higher activity than V V O 2 species and this observation can be attributed to the different redox behavior of the vanadium species, 42 the different mechanism of cellular uptake 43 or the different speciation in the cytosol, 44 and it has been also proposed that [V IV O(OH) 3 ] À could be the active species. 5,10 In our study the picFF complex with V IV O indeed exhibits slightly higher activity than the V V O 2 complex, while in the Hhypic complexes the oxidation state of the metal center has no significant influence on the inhibition of FFA. Zinc complexes also exhibited some insulin-mimetic activity, with the Hhypic complex (9) being more active than the other two, however, the insulin-mimetic activity of these complexes did not show more potent activity than ZnSO 4 .

Conclusion
Two vanadium(V) compounds with 3,5-difluoropicolinato acid (HpicFF) and 3-hydroxypicolinato acid (H 2 hypic), four zinc(II) compounds with HpicFF in the presence of water, pyridine (py), 4-(dimethylamino)pyridine (DMAP), and 1,10-phenanthroline (phen) and two zinc(II) compounds with H 2 hypic in the presence of DMAP and phen as well as a zinc(II) compound with HpicCH 2 OH have been prepared and characterized. The crystal structure of NH 4 [VO 2 (picFF) 2 ]Á1.6H 2 O (3Á1.6H 2 O) possesses a spatial arrangement with the carboxylate oxygen atoms in a mutual trans orientation although the carboxylate oxygen atoms in the prototypical complex NH 4 (11) and [Zn(Hhypic) 2 (phen)] (12) two different spatial arrangements were found with trans (5, 10 and 11) and cis (6, 7, 8 and 12) octahedral geometry. The presence of the DMAP ligand seems not to play a crucial role in the spatial arrangement since the geometry in 7 is cis and in 11 it is trans. Attempts were made to prepare the analogue of 11 with the py ligand for structural comparison with 6, however, we were not able to obtain crystals suitable for XRD analysis. As already observed for [Zn(picCN) 2 (py) 2 ] also 6 loses all pyridine molecules on standing in air and is transformed to the water complex 5. The insulinmimetic activity of V IV O (1, 2), V V O 2 (3,4) and the Zn II complexes (5, 9 and 13) was studied by an in vitro inhibition of the free fatty acid (FFA) release from isolated rat adipocytes treated with epinephrine. All selected metal complexes showed the insulinmimetic activity. Vanadium complexes 1-4 exhibit activities similar to VOSO 4 with H 2 hypic complexes being more active than the picFF ones. Zinc complexes also exhibited some insulin-mimetic activity, with the H 2 hypic complex (9) being more active than the other two (5, 13).

Conflicts of interest
There are no conflicts of interest to declare.