Synthesis and in vitro Bio-activity Evaluation of N4-benzyl Substituted 5-Chloroisatin-3-thiosemicarbazones as Urease and Glycation Inhibitors.

A series of fifteen N4-benzyl substituted 5-chloroisatin-3-thiosemicarbazones 5a-o were synthesized and screened mainly for their antiurease and antiglycation effects. Lemna aequinocitalis growth and Artemia salina assays were carried out to determine their phytotoxicity and cytotoxicity potential. All the compounds proved to be extremely effective urease inhibitors, demonstrating enzyme inhibition much better than the reference inhibitor, thiourea (IC50 values 1.31 ± 0.06 to 3.24 ± 0.15 vs. 22.3 ± 1.12 µM). On the other hand, eight out of fifteen compounds tested, i.e. 5b, 5c, 5h-k, 5m and 5n were found to be potent glycation inhibitors. Of these, five viz. 5c, 5h-j and 5n proved to be exceedingly efficient, displaying glycation inhibition greater than the reference inhibitor, rutin (IC50 values 114.51 ± 1.08 to 229.94 ± 3.40 vs. 294.5 ± 1.5 µM).

Synthesis, X-ray molecular structure, biological evaluation and molecular docking studies of some N4-benzyl substituted 5-nitroisatin-3-thiosemicarbazones.

A series of fifteen N4-benzyl substituted 5-nitroisatin-3-thiosemicarbazones 5a-o was synthesized and evaluated for urease inhibitory, phytotoxic and cytotoxic influences. All the compounds proved to be highly potent inhibitors of the enzyme, showing inhibitory activity (IC50=0.87±0.25-8.09±0.23µM) much better than the reference inhibitor, thiourea (IC50=22.3±1.12µM) and may thus act as persuasive leads for further studies. In phytotoxicity assay, twelve out of fifteen thiosemicarbazones tested i.e. 5a-e, 5g, 5i and 5k-o appeared to be active, exhibiting weak or non-significant (5-35%) growth inhibition at the highest tested concentrations (1000 or 500µg/mL). In contrast, only one compound i.e. 5i was active in the brine shrimp (Artemia salina) lethality bioassay, demonstrating cytotoxic activity with LD50 value 2.55×10-5M. Molecular docking studies of compounds 5a-o were also performed to identify their probable binding modes in the active site of the enzyme.

Improved physicochemical characteristics of artemisinin using succinic acid.

Artemisinin (ARMN) is a potent antimalarial drug, which is effective against multidrug resistant strains of Plasmodium falciparum and produces rapid recovery even in patients with cerebral malaria. Being poorly soluble in water, artemisinin is incompletely absorbed after oral intake due to poor dissolution characteristics in the intestinal fluids. To enhance these properties, solid dispersions of artemisinin with succinic acid (SUC) were prepared using drug-carrier ratios 1 : 1, 1 : 4, 1 : 6, 1 : 8 and 1 : 10 by solvent evaporation and freeze drying methods. These solid dispersions were characterized by differential scanning calorimetery (DSC), Fourier transform infrared spectroscopy (FTIR), x-ray diffraction patterns (XRD), phase solubility and dissolution kinetics evaluated by applying zero order, first order, Higuchi, and Korsmeyer-Peppas models. Physical mixtures produced significantly higher aqueous solubility and rate of dissolution as compared to artemisinin alone. The dissolution profiles of all formulations followed Higuchi model and exhibited diffusion-controlled release of drug. Solvent evaporation method (SLVPs) exhibited improved solubility and freeze dried solid dispersions (FDSDs) produced highest solubility but stability constant was opposite. ARMN and SUC both were found completely crystalline as shown by their XRD patterns. Physical mixtures (PMs) showed reduced intensity in their XRD patterns while solid dispersions by SLVPs exhibited twice reduced intensity and much displaced angles, whereas FDSDs showed synergistic effects in some of ARMN and SUC peaks. DSC thermograms of FDSDs at drug-carrier ratios 1 : 1-1 : 4 showed lower melting temperature and enthalpy change (deltaH) values than respective SLVPs, whereas at higher ratios, a reverse was true. SLVPs showed displaced methyl stretching bands at lower drug-carrier ratios and exhibited O-H stretching characteristic bands of SUC at higher drug-carrier ratios. In addition, carbonyl group and C-O stretching vibrations characteristic of SUC (1307 cm(-1)) appeared prominently compared to PMs, whereas C-O stretching characteristic bands of ARMN disappeared at higher ratios. FDSDs exhibited distinct nature of bonding compared to respective SLVPs and PMs.

5-Nitroisatin-derived thiosemicarbazones: potential antileishmanial agents.

A series of 29 previously reported N(4)-substituted 5-nitroisatin-3-thiosemicarbazones 2-30 has been screened for leishmanicidal potential. Compounds 2-4, 7, 8, 10-13, 15-19, 21, 23, 24, 26, 28 and 30 exhibited good to excellent antileishmanial activities with IC50 values ranging from 0.44 ± 0.02 to 32.38 ± 0.66 µg/mL. Of these, 5, 7, 19 and 28 proved to be the most active antileishmanial agents, displaying activities with IC50 values 1.78 ± 0.35, 0.44 ± 0.02, 1.91 ± 0.04 and 4.28 ± 0.75 µg/mL, respectively, which were even better than the standard drug, pentamidine (IC50 = 5.09 ± 0.04 µg/mL). This study presents the first example of exhibition of leishmanicidal potential by isatin-thiosemicarbazones and as such furnishes a solid basis for further research on these compounds to develop more potent antileishmanial agents.

(2Z)-N-(2-Chloro-benz-yl)-2-(2-oxo-2,3-dihydro-1H-indol-3-yl-idene)hydrazinecarbothio-amide.

In the title compound, C(16)H(13)ClN(4)OS, the isatin ring system is oriented at dihedral angles of 10.60 (7) and 72.60 (3)° with respect to the thio-semicarbazide and 2-chloro-benzyl groups, respectively. The near planarity of the isatin and thio-semicarbazide groups [r.m.s. deviations of 0.0420 and 0.0163 Å, respectively] is reinforced by intra-molecular N-H⋯O and N-H⋯N hydrogen bonds, which generate S(6) and S(5) rings, respectively. In the crystal, inversion dimers linked by pairs of N-H⋯O hydrogen bonds generate R(2) (2)(8) loops. Aromatic π-π stacking inter-actions between the centroids of heterocyclic five-membered and benzene rings [distance = 3.6866 (11) Å] are also observed.

Synthesis and biological evaluation of some new N4-aryl substituted 5-chloroisatin-3-thiosemicarbazones.

A new series of sixteen N4-aryl substituted 5-chloroisatin-3-thiosemicarbazones 2a-2p has been synthesized, characterized and tested for selected biological activities i.e. cytotoxicity, phytotoxicity and urease inhibition. In the brine shrimp bioassay, all the synthesized compounds gave LD50 values>2.30x10(-4) M-2.79x10(-4) M and were, therefore, found to be almost inactive, whereas in phytotoxicity assay, regardless of the nature of aryl substituents, they displayed weak to moderate (5-40%) phytotoxic activity at the highest tested concentrations (500 or 1000 µg/mL). In urease inhibition bioassay, compounds 2a, 2c, 2e, 2f, 2k and 2m exhibited relatively a higher degree of urease inhibition with IC50 values ranging from 38.91 µM to 76.65 µM and thus proved to be potent inhibitors of the enzyme. Of these, 2f and 2m displayed pronounced inhibition with IC50 values 38.91 µM and 39.50 µM, respectively, and may act as lead compounds for further studies. Structure-activity relationship (SAR) studies revealed that electronic effects of the substituents about the phenyl ring at N4 of the thiosemicarbazone moiety played an important role in enhancing the urease inhibitory potential of some of the synthesized compounds.

(Z)-4-(3-Fluoro-phen-yl)-1-(5-nitro-2-oxo-indolin-3-yl-idene)thio-semicarbazide.

In the title compound, C(15)H(10)FN(5)O(3)S, an intra-molecular N-H⋯N hydrogen bond generates an S(5) ring, whereas N-H⋯O and C-H⋯S inter-actions complete S(6) ring motifs. The dihedral angle between the isatin ring system and the fluoro-benzene ring is 5.96 (6)° and the complete mol-ecule is close to planar (r.m.s. deviation for all the non-H atoms = 0.112 Å). In the crystal, mol-ecules are linked by N-H⋯O hydrogen bonds to form C(8) chains along the [100] direction and C-H⋯O inter-actions are also observed.

Improved physicochemical characteristics of artemisinin-nicotinamide solid dispersions by solvent evaporation and freeze dried methods.

Artemisinin (ARMN) is a drug of choice against drug-resistant malaria especially due to Plasmodium falciparum. Being poorly soluble in water, its solid dispersions with nicotinamide (NA) were prepared at various drug-carrier ratios (1:1, 1:4, 1:6, 1:8, 1:10) by solvent evaporation and freeze drying methods. These solid dispersions were characterized by differential scanning calorimetery (DSC), fourier transform infrared spectroscopy (FTIR), X-ray diffraction patterns (XRD), phase solubility and dissolution studies. Artemisinin and nicotinamide both were found completely crystalline as shown by their XRD patterns. Physical mixtures (PMs) showed decreased intensity in their XRD patterns while solid dispersions by solvent evaporation method (SLVPs) exhibited displaced angles and decreased intensity whereas freeze dried solid dispersions (FDSDs) showed least number of peaks having low intensity and maximum displaced angles. DSC thermograms of drug-carrier ratios at 1:1-1:4 showed lower melting temperature than artemisinin and nicotinamide in all preparations. Endothermic temperature of artemisinin in PMs and SLVPs increased with rise of nicotinamide content upto 1:6 ratio followed by decline. All samples showed crystallization temperature below the artemisinin except drug-carrier ratio 1:6 of PMs while δH value was minimum at this ratio. FDSDs produced lowest endothermic temperature than corresponding PMs and SLVPs. SLVPs exhibited band shifting in both functional and fingerprint region compared to respective PMs as exhibited by their FTIR spectra. FDSDs and SLVPs showed different nature of bonding among artemisinin and nicotinamide. FDSDs produced strongest CONH(2) bonding followed by SLVPs and PMs respectively. PMs produced significantly higher aqueous solubility and rate of dissolution as compared to artemisinin alone. SLVPs exhibited improved solubility and dissolution profile corresponding to PMs. FDSDs showed highest release rate and aqueous solubility followed by SLVPs and PMs at all ratios. PMs and SLVPs showed their highest dissolution profile at 1:6 drug-carrier ratio followed by gradual decrease while FDSDs progressed in dissolution rate with increase of nicotinamide content successively upto maximum at 1:10 ratio.

Synthesis and Toxicity Evaluation of Some N4-Aryl Substituted 5-Trifluoromethoxyisatin-3-thiosemicarbazones.

A series of twenty one N4-aryl substituted 5-trifluoromethoxyisatin-3-thiosemicarbazones 3a-3u was synthesized by the reaction of trifluoromethoxyisatin 1 with different arylthiosemicarbazides 2 in aqueous ethanol (50%), containing a few drops of acetic acid. Their structures were established on the basis of analytical (CHN) and spectral (IR, 1H-NMR, EIMS) data. All the synthesized compounds were evaluated for their toxicity potential by a brine shrimp lethality bioassay. Ten compounds i.e., 3a, 3e, 3i-3l and 3n-3q proved to be active in this assay, displaying promising toxicity (LD50 = 1.11 × 10-5 M - 1.80 × 10-4 M). Amongst these, 3k, 3n and 3o were found to be the most active ones (LD50 = 1.11 × 10-5 M - 1.43 × 10-5 M). Compound 3k showed the highest activity with a LD50 value of 1.11 × 10-5 M and can, therefore, be used as a lead for further studies. Structure-activity relationship (SAR) studies revealed that the presence of strong inductively electron-attracting trifluoromethoxy substituent at position-5 of the isatin moiety played an important role in inducing or enhancing toxic potentiality of some of the synthesized compounds.

4-(2-Fluoro-phen-yl)-1-(2-oxoindolin-3-yl-idene)thio-semicarbazide.

The title compound, C(15)H(11)FN(4)OS, is almost planar, the dihedral angle between the aromatic ring systems being 5.00 (13)°. The conformation is stabilized by intra-molecular N-H⋯N and N-H⋯O hydrogen bonds, which generate S(5) and S(6) rings, respectively. N-H⋯F and C-H⋯S inter-actions also occur. In the crystal, inversion dimers linked by pairs of N-H⋯O hydrogen bonds occur, generating R(2) (2)(8) loops.

4-(5-Chloro-2-methyl-phen-yl)-1-[2-oxo-5-(trifluoro-meth-oxy)indolin-3-yl-idene]thio-semicarbazide.

The asymmetric unit of the title compound, C(17)H(12)ClF(3)N(4)O(2)S, contains two mol-ecules, which differ in their planarity and hydrogen bonding. In one mol-ecule, the 2-oxoindolin (C(8)/N/O A), thio-semicarbazide (N(3)/C/S B) and 5-chloro-2-methyl-phenyl (C(7)/Cl C) units are planar with r.m.s. deviations of 0.0110, 0.0173 and 0.0259 Å, respectively. The dihedral angles A/B, B/C and A/C are 1.74 (15), 40.70 (13) and 41.00 (11)°, respectively. In the other mol-ecule the deviations are 0.0455, 0.0007 and 0.0143 Å, respectively, and the dihedral angles are 5.01 (14), 4.53 (16) and 3.38 (13)°, respectively. In both mol-ecules, intra-molecular N-H⋯N and N-H⋯O hydrogen bonds form S(5) and S(6) ring motifs, respectively and C-H⋯S interactions occur. In one of the molecules, an intramolecular C-H⋯F interaction is also present. In the crystal, the mol-ecules are linked by N-H⋯O, C-H⋯F, C-H⋯O and N-H⋯S hydrogen bonding, forming a polymeric network.

4-(3-Methoxy-phen-yl)-1-(2-oxoindolin-3-yl-idene)thio-semicarbazide.

In the title compound, C(16)H(14)N(4)O(2)S, intra-molecular N-H⋯N hydrogen bonding forms an S(5) ring, whereas N-H⋯O and C-H⋯S inter-actions complete S(6) ring motifs. In the crystal, mol-ecules form inversion dimers due to N-H⋯O inter-actions. The dimers are inter-linked through N-H⋯S hydrogen bonds and π-π inter-actions occur with a centroid-centroid distance of 3.8422 (11) Šbetween the meth-oxy-containing benzene ring and the five-membered heterocyclic ring.

4-(3-Nitro-phen-yl)-1-(2-oxoindolin-3-yl-idene)thio-semicarbazide.

In the title compound, C(15)H(11)N(5)O(3)S, intra-molecular N-H⋯N hydrogen bonding forms an S(5) ring motif, whereas N-H⋯O and C-H⋯S inter-actions type complete S(6) ring motifs. The 2-oxoindoline and 3-methoxy-phenyl rings are almost planar, with r.m.s. deviations of 0.0178 and 0.0149 Å, respectively, and form a dihedral angle of 33.59 (3)°. In the crystal, mol-ecules are inter-linked through the nitro groups in an end-to-end fashion via N-H⋯O and C-H⋯O inter-actions.

4-(3-Fluoro-phen-yl)-1-(2-oxoindolin-3-yl-idene)thio-semicarbazide.

In the title compound, C(15)H(11)FN(4)OS, there are three independent mol-ecules, each with a disordered 3-fluoro-phenyl group [occupancy ratios = 0.547 (17):0.453 (17), 0.645 (5):0.355 (5) and 0.626 (15):0.374 (15)] and displaying dihedral angles of 4.2 (3), 25.2 (6) and 32.4 (5)° between the 2-oxoindoline and fluoro-substituted phenyl rings. Strong intra-molecular N-H⋯N and N-H⋯O and weak intra-molecular C-H⋯S hydrogen bonds complete S(5) and S(6) ring motifs, while strong inter-molecular N-H⋯O hydrogen bonds inter-connect the three independent mol-ecules through R(3) (3)(12) ring motifs. The three-mol-ecule units are in turn linked into polymeric sheets via C-H⋯F and C-H⋯S hydrogen bonds and π-π inter-actions [centroid-centroid distances in the range 3.520 (2)-3.820 (9) Å].

4-(2-Ethyl-phen-yl)-1-(2-oxoindolin-3-yl-idene)thio-semicarbazide.

The title compound, C(17)H(16)N(4)OS, is stabilized in the form of a two-dimensional polymeric network due to inter-molecular N-H⋯S and N-H⋯O hydrogen bonds. An intra-molecular N-H⋯N hydrogen bond forms an S(5) ring, whereas inter-actions of the N-H⋯O and C-H⋯S types complete S(6) ring motifs. π-π inter-actions with a centroid-centroid distance of 3.6514 (10) Šare found between the ethyl-substituted benzene ring and the heterocyclic ring of the isatin derivative.

4-(3-Iodo-phen-yl)-1-(2-oxoindolin-3-yl-idene)thio-semicarbazide.

In the title compound, C(15)H(11)IN(4)OS, intra-molecular N-H⋯N, N-H⋯O and C-H⋯S inter-actions generate one S(5) and two S(6) ring motifs. In the crystal, mol-ecules form centrosymmetric dimers via pairs of N-H⋯O inter-actions, generating R(2) (2)(8) ring motifs. In addition a short inter-molecular I⋯S contact of 3.352 (3) Šis observed.

1-Acetyl-3-[2-(2,3,5,6-tetra-fluoro-phen-yl)hydrazin-1-yl-idene]indolin-2-one.

In the title compound, C(16)H(9)F(4)N(3)O(2), the dihedral angle between the aromatic ring systems is 4.10 (14)° and a bifurcated intra-molecular N-H⋯(O,F) hydrogen bond generates an S(6) ring for the O-atom acceptor and an S(5) ring for the F-atom acceptor. A short C-H⋯O conact also occurs. In the crystal, mol-ecules are linked by C-H⋯O inter-actions.

1-[1-(4-Bromo-phen-yl)ethyl-idene]-4-(2,4-dimeth-oxy-phen-yl)thio-semicarbazide.

In the title compound, C(17)H(18)BrN(3)O(2)S, the dihedral angle between the aromatic rings is 9.15 (17)°. A bifurcated intra-molecular N-H⋯(N,O) hydrogen bond generates two S(5) rings and a weak intra-molecular C-H⋯S inter-action completes an S(6) ring motif. In the crystal, inversion dimers linked by pairs of N-H⋯S hydrogen bonds generate R(2) (2)(8) loops and weak C-H⋯S and C-H⋯π inter-actions are also present.

N-{2-[N-(4-Methyl-phen-yl)oxamo-yl]phen-yl}propanamide.

The title compound, C(18)H(18)N(2)O(3), is the product of the heterocyclic ring cleavage at position 2 of 1-propionylisatin. Two centrosymmetric cyclic motifs, viz. R(2) (2)(14) and R(2) (2)(18), are formed by N-H⋯O hydrogen bonds with the propanamide and amino-phenyl units, respectively, as the N-H donors. These motifs combine into two C(2) (2)(8) chain motifs parallel to the b axis. The chain structure is stabilized by C-H⋯π inter-actions between the benzene rings, where C-H is from the phenyl ring of the cleaved part of 1-pro-pionylisatin.

1-[2-Oxo-5-(trifluoro-meth-oxy)indolin-3-yl-idene]-4-[4-(trifluoro-methyl)-phen-yl]thio-semicarbazide.

In the title compound, C(17)H(10)F(6)N(4)O(2)S, an intra-molecular N-H⋯N hydrogen bonds forms an S(5) ring whereas N-H⋯O and C-H⋯S inter-actions complete S(6) ring motifs. The dihedral angle between the fused ring system and the phenyl ring is 6.68 (8)°. In the crystal, the mol-ecules are dimerized due to N-H⋯O inter-actions. π-π inter-actions are present between the benzene rings [centroid-centroid distance = 3.6913 (15) Å] and between the five membered ring and the trifluoro-meth-yl)phenyl ring [centroids-centroid distance = 3.7827 (16) Å]. One of the trifluoro-meth-oxy F atoms is disordered over two sites with occupancy ratio of 0.76 (3):0.24 (3). The F atoms of the p-trifluoro-methyl substituent are disordered over three sets of sites with an occupancy ratio of 0.70 (2):0.152 (11):0.147 (13).