Novel urea and bis-urea primaquine derivatives with hydroxyphenyl or halogenphenyl substituents: Synthesis and biological evaluation.

A series of novel compounds 3a-j and 6a-j with primaquine and hydroxyl or halogen substituted benzene moieties bridged by urea or bis-urea functionalities were designed, synthesized and evaluated for biological activity. The title compounds were prepared using benzotriazole as the synthon, through several synthetic steps. 3-[3,5-Bis(trifluoromethyl)phenyl]-1-{4-[(6-methoxyquinolin-8-yl)amino]pentyl}urea (3j) was the most active urea and 1-[({4-[(6-methoxyquinolin-8-yl)amino]pentyl}carbamoyl)amino]-3-[3-(trifluoromethyl)phenyl]urea (6h) the most active bis-urea derivative in antiproliferative screening in vitro against eight tested cancer cell lines. Urea derivatives 3a-g with hydroxy group or one halogen atom showed moderate antiproliferative effects against all the tested cell lines, but stronger activity against breast carcinoma MCF-7 cell line, while trifluoromethyl derivatives 3h-j showed antiproliferative effects against all the tested cell lines in low micromolar range. Finally, bis-ureas with hydroxy and fluoro substituents 6a-d showed extreme selectivity and chloro or bromo derivatives 6e-g high selectivity against MCF-7 cells (IC50 0.1-2.6 µM). p-Fluoro derivative 6d, namely 3-(4-fluorophenyl)-1-[({4-[(6-methoxyquinolin-8-yl)amino]pentyl}carbamoyl)amino]urea, is the most promising compound. Further biological experiments showed that 6d affected cell cycle and induced cell death of MCF-7 cell line. Due to its high activity against MCF-7 cell line (IC50 0.31 µM), extreme selectivity and full agreement with the Lipinski's and Gelovani's rules for prospective small molecular drugs, 6d may be considered as a lead compound in development of breast carcinoma drugs. Urea 3b and almost all bis-ureas showed high antioxidant activity in DPPH assay, but urea derivatives were more active in lipid peroxidation test. Only few compounds exhibited weak inhibition of soybean lipoxygenase. Compound 3j exhibited the strongest antimicrobial activity in susceptibility assay in vitro (MIC = 1.6-12.5 µg ml-1).

Preparation and pharmacochemical evaluation of mixed ligand copper(II) complexes with triethanolamine and thiophenyl-2 saturated carboxylic acids.

The dinuclear complex [Cu(2)(L(1))(2)(H(2)tea)(2)] (1) as well as the linear trinuclear complexes [Cu(3)(L(1))(4)(H(2)tea)(2)] (2), [Cu(3)(L(2))(4)(H(2)tea)(2)] (3) and [Cu(3)(L(1))(2)(H(2)tea)(2)(NO(3))(2)] (4) where L(1) = 2-thiophene carboxylato, L(2) = 2-thiophene acetato and H(2)tea = the single deprotonated form of triethanolamine have been prepared and pharmacochemically studied. The crystal structure of 1 is also reported. In vitro antioxidant activity of free ligands and their respective copper complexes includes: a) interaction with 1,1-diphenyl-2-picrylhydrazyl stable free radical, b) the ΗΟ˙ mediated oxidation of DMSO, c) scavenging of superoxide anion radicals, d) inhibition of lipid peroxidation and e) soybean lipoxygenase inhibition. The results indicate selectivity of the complexes to different free radicals as a consequence of their physichochemical features. The majority of the complexes 1, 2, 3, 4 effectively inhibit lipid peroxidation. The trinuclear complex 3 is by far the most active with IC(50)=10 µM, within the set, followed by complexes 1 and 2. The complexes were evaluated for their efficacy as anticancer agents against different cancer and normal human cell lines. Results showed that, these compounds mediate a moderate cytotoxic response to normal and cancer cell lines tested and induce cell cycle arrest in G2/M phase of the cell cycle. Flow cytometric analysis suggested that the tested compounds can induce apoptosis.

Effect of N-acetyl-L-cysteine supplementation in semen extenders on semen quality and reactive oxygen species of chilled canine spermatozoa.

The objective of this study was to evaluate the quality of chilled dog semen processed with extenders containing various concentrations of N-acetyl-L-cysteine (NAC). Ejaculates from five dogs were collected, pooled and evaluated for concentration, motility, rapid steady forward movement (RSF-movement), viability, acrosomal integrity and by the hypo-osmotic swelling test (HOST). In addition, superoxide anion (O(2)(-*)) production, hydroxyl radicals (OH(*)) and total reactive oxygen species (tROS) were determined. The pool was divided into five aliquots, which were diluted to a final concentration of 66.66 x 10(6) spermatozoa/ml with Tris-glucose-egg yolk extender containing one of the following concentrations of NAC (0, 0.5, 1, 2.5 or 5 mm). The semen aliquots were chilled and preserved at 4 degrees C. Semen quality was evaluated after rewarming at 72 h. Sperm motility was significantly higher with the 0.5 mm concentration compared with the control group (p = 0.001). Rapid steady forward movement was higher with the 0.5 and 1 mm concentrations compared with the control and 5 mm group (p < 0.001). Viability and HOST percentages were not significantly altered. Compared with the control, the 5 mm concentration showed significantly reduced percentages of spermatozoa with normal acrosomes (p = 0.049). None of the ROS values at 72 h were significantly affected by the presence of NAC in semen extenders, although all NAC concentrations showed lower O(2)(-*) and OH(*) values compared with the control. Only the concentrations of 1 and 5 mm inhibited the significant increase of tROS values after 72 h, compared with the fresh semen value. In conclusion, NAC supplementation of semen extenders is beneficial to semen motility of canine spermatozoa during chilling with the 0.5 mm concentration being the most effective, although no significant ROS inhibition was observed at 72 h.

Urea and carbamate derivatives of primaquine: synthesis, cytostatic and antioxidant activities.

The novel urea primaquine derivatives 3 were prepared by aminolysis of primaquine benzotriazolide 2 with several hydroxyamines and ethylendiamine, while carbamates 4 were synthesized from the same precursor 2 and alcohols. All compounds are fully chemically characterized and evaluated for their cytostatic and antioxidant activities. The most prominent antiproliferative activity was obtained by compounds 3c, 3d, 3g, and 5b (IC(50)=9-40 microM). 1-(5-Hydroxypentyl)-3-[4-(6-methoxy-quinolin-8-ylamino)-pentyl]urea (3c) showed extreme selectivity toward SW 620 colon cancer cells (IC(50)=0.2 microM) and a bit less toward lung cancer cells H 460. Hydroxyurea 3h showed the highest interaction with DPPH. Primaquine twin drug 3g showed very significant inhibition on LOX soybean (IC(50)=62 microM). Almost all the tested derivatives highly inhibited lipid peroxidation, significantly stronger than primaquine phosphate.

Effect of antioxidant supplementation in semen extenders on semen quality and reactive oxygen species of chilled canine spermatozoa.

The objective of this study was to evaluate quality of chilled dog semen processed with extenders containing various antioxidants. Single ejaculates from five dogs were always pooled and evaluated for concentration, sperm motility, progressive motility (RSF-movement), viability, acrosomal integrity and by the hypo-osmotic swelling (HOS)-test. Also, superoxide (O(2)(-)) production, hydroxyl radicals (OH) and total reactive oxygen species (tROS) were determined. Pooled semen was divided in seven aliquots (for control and test conditions), which were diluted to a final concentration of 67x10(6)spermatozoa/ml with TRIS-glucose-egg yolk extender with or without the following supplements: control (without antioxidants), vitamin C (0.5mM), N-acetyl-l-cysteine (NAC; 0.5mM), taurine (0.2mM), catalase (100u/ml), vitamin E (0.1mM) and 5-(4-dimethylamino-phenyl)-2-phenyl-penta-2,4-dienoic acid (B16; 0.1mM). The semen aliquots were chilled and preserved at 4 degrees C. Portions of chilled semen were removed at 24 and 72h, and semen quality was evaluated after rewarming. At 24h the mean (+/-S.E.M.) sperm motility was higher (p<0.001) when vitamin E, taurine and B16 were added in the extender, whereas more spermatozoa with RSF-movement were observed (p<0.001) in the vitamin E, catalase, B16 and taurine groups. Sperm viability was higher (p=0.040) in B16 and vitamin E groups and the percentage of swollen spermatozoa was higher (p=0.002) only in the B16 group. Acrosomal integrity and OH were not significantly influenced by any of the antioxidants tested. Superoxide production was significantly lower when vitamin C, B16 and vitamin E were added in semen extenders compared with the control (p=0.017). All antioxidant groups, except vitamin C and NAC, contained less tROS compared to the control group, but only the B16 group value differed significantly (p=0.05). At 72h sperm motility was higher (p<0.001) when vitamin E, catalase, B16, taurine and NAC were added in the extender. More spermatozoa with RSF-movement were observed (p<0.001) in the vitamin E, catalase, B16, taurine and NAC treatment groups. Sperm viability was higher (p=0.001) when vitamin E, B16, taurine and vitamin C were added in semen extenders. HOS-test percentages were higher (p=0.016) in the B16, vitamin E, catalase and NAC groups. Acrosomal integrity was not influenced in any case. Production of O(2)(-) was significantly higher using catalase compared to all the other groups (p=0.006), while OH was not significantly influenced by any of the antioxidants tested. The addition of vitamin E, catalase and B16 in semen extenders resulted in significantly lower tROS values compared with the controls (p<0.0005). The results suggest that vitamin E and B16 had the most pronounced effect in preserving semen quality of chilled dog spermatozoa.

Evaluation of anti-inflammatory and antioxidant activities of copper (II) Schiff mono-base and copper(II) Schiff base coordination compounds of dien with heterocyclic aldehydes and 2-amino-5-methyl-thiazole.

Highly reactive radicals are implicated in many pathological conditions. The quest for radical scavengers or antioxidants, spans the previous decades. A new series of complexes of the type [Cu (dien) (2a-2tzn) Y(2)] and [Cu (dienXXY(2)) (2a-5mt)] and of the type [Cu (dptaS) Cl(2)] and [Cu (dptaS) Br(2)] (dptaS = 1, 3-propanediamine) or Schiff mono-base of dipropylenetriamine with 2-thiophene-carboxaldehyde, has been tested for anti-inflammatory and antioxidant activity. The tested compounds inhibit the carrageenin-induced rat paw edema (52.0-82.6%) and present important scavenging activity. Compound 6 is the most potent (82.6%) in the in vivo experiment. Lipophilicity-as R(M) values - has been determined. The results support that in general, adducts of the type [Cu (dienXXY(2)) (2a-5mt)] exhibit increased activity compared to the starting material of type [Cu (dienXXY(2))]. An attempt to correlate the biological results with their structural characteristics and physicochemical parameters has been made.

Quality and reactive oxygen species of extended canine semen after vitamin C supplementation.

The objective of this study was to evaluate the quality of extended dog semen processed with diluents containing various concentrations of vitamin C. Ejaculates from five dogs were collected, pooled and evaluated for concentration, sperm motility, rapid steady forward movement (RSF-movement), viability, acrosomal integrity and by the hypo-osmotic swelling test. Also, superoxide (O(2)(-)*) production, hydroxyl radicals (OH*) and total reactive oxygen species (tROS) were determined. The pool was divided in five aliquots, which were diluted to a final concentration of 66 x 10(6) spermatozoa/ml with a Tris-glucose-egg yolk extender containing one of the following concentrations of vitamin C (0, 0.1, 0.5, 1 or 2.5 mM). The semen aliquots were chilled and preserved at 4 degrees C. Portions of chilled semen were removed at 24 and 72 h, and semen quality was evaluated after rewarming. This process was repeated 10 times in pooled semen of the same origin and data were analysed by one-way analysis of variance. At both times, none of the semen quality parameters were positively influenced (p>0.05) by vitamin C supplementation. At 24 h, none of the reactive oxygen species (O(2)(-)*, OH*, tROS) were significantly altered. At 72 h, significant reductions of O(2)(-)* production were observed by the concentrations of 0.1, 0.5 and 2.5 mM compared with the 0 mM concentration (p=0.049). Also, at 72 h, the 2.5 mM concentration showed significantly lower OH* values in comparison with the control group (p=0.048). In conclusion, addition of vitamin C to semen extenders does not benefit the quality of canine extended spermatozoa.

Synthesis and pharmacochemical evaluation of novel aryl-acetic acid inhibitors of lipoxygenase, antioxidants, and anti-inflammatory agents.

Lipoxygenase catalyzes the first two steps of the transformation of arachidonic acid into leukotrienes which are implicated in host defense reactions. It is well known that many acids possess potent anti-inflammatory activity. Taking into account that compounds bearing a thienyl, naphthyl, pyrollyl, and 2,4-di-tert-butyl-phenol moieties possess anti-inflammatory activity which is related to their capacity to transfer electrons and to scavenge reactive oxygen species, we synthesized some new aryl-acetic acids and we explored their ability to inhibit soybean lipoxygenase, to present antioxidant and anti-inflammatory activities, and to interact with glutathione. The compounds have shown important antioxidant activity, medium anti-inflammatory activity, and very good inhibition of soybean lipoxygenase. Compound 3-(3,5-di-tert-butyl-2-hydroxy-phenyl)-2-phenyl-acrylic acid (1i) showed significant in vitro LO inhibition (IC(50) 65 microM). The results are discussed in terms of structural and physicochemical characteristics of the compounds. The structures of the synthesized compounds were confirmed by spectral and elemental analysis. Their lipophilicity are experimentally determined by RPTLC method.

Effect of antioxidant supplementation on semen quality and reactive oxygen species of frozen-thawed canine spermatozoa.

The objective of this study was to evaluate post-thaw quality of frozen dog semen processed with diluents containing different antioxidants. Ejaculates were collected, pooled and evaluated for concentration, motility, rapid steady forward movement (RSF movement), viability, acrosomal integrity and by the hypo-osmotic swelling test. Also, superoxide production, hydroxyl radicals and total reactive oxygen species (tROS) were determined. The pool was divided in seven aliquots, for control and test conditions, which were processed for cryopreservation. The sperm pellets were diluted to a final concentration of 200x10(6)sperm/ml with TRIS-glucose-egg yolk extender containing one of the following supplements: vitamin C (1.5mM), NAC (N-acetyl-l-cysteine; 1.5mM), taurine (0.6mM), catalase (300U/ml), vitamin E (0.3mM) and B16 [5-(4-dimethylamino-phenyl)-2-phenyl-penta-2,4-dienoic acid; 0.3mM]. Post-thaw semen evaluation showed that mean (+/-S.E.M.) motility was increased (p<0.001) after addition of catalase (49.75+/-3.63 versus 39.00+/-2.90 in controls), whereas more spermatozoa with RSF movement were observed (p<0.001) after the catalase, NAC and vitamin E treatments (31.75+/-3.46, 28.00+/-3.27, 26.75+/-3.15, respectively, versus 17.00+/-2.26 in controls). Viability was increased (p<0.001) after addition of catalase, taurine, NAC and tocopherol (66.00+/-3.03, 61.90+/-2.48, 60.60+/-1.93 and 60.50+/-4.12, respectively, versus 51.70+/-2.81 in controls). The percentage of swollen spermatozoa was increased after addition of catalase and taurine (61.75+/-1.61 and 61.25+/-1.49, respectively, versus 55.65+/-1.64 in controls). Acrosomal integrity was not influenced in any case. B16 addition had adverse effects on all parameters evaluated. None of the reactive oxygen species were significantly reduced post-thaw in antioxidant treated semen. The results suggest that catalase had the most pronounced effect in improving post-thaw quality of canine spermatozoa.

Synthesis of phenyl-substituted amides with antioxidant and anti-inflammatory activity as novel lipoxygenase inhibitors.

Lipoxygenases (LO) have been implicated in several inflammatory diseases such as asthma, immune disorders, and cancers. Lipoxygenases play an essential role in the biosynthesis of the leukotrienes. Leukotrienes have been implicated as mediators in the pathophysiology of inflammatory diseases, host defense reactions and they were found to play important role in the propagation of the diseases states, exacerbating the local events and ultimately leading to tissue damage. As a consequence of these broad biological implications, there is a great interest in synthesising new compounds with this activity. The synthesis of new amides of aryl acetic acid is described. The structures of the synthesized compounds were confirmed by spectral and elemental analysis. Since lipophilicity is a significant physicochemical property determining distribution, bioavailability, metabolic activity and elimination, their lipophilicity is experimentally determined from RPTLC method. Several parameters were theoretically calculated and were used for a QSAR study. The compounds are tested in vitro on: a) soybean lipoxygenase inhibition, b) interaction with 1,1-diphenyl-2-picryl-hydrazyl (DPPH) stable free radical, c) the HO* mediated oxidation of DMSO, d) inhibition of lipid peroxidation, e) scavenging of superoxide anion radicals f) interaction with glutathione and g) in vivo for the inhibition of carrageenin induced rat paw edema. The compounds present significant antioxidant activities, medium anti-inflammatory activity and potent inhibition of soybean lipoxygenase as a result of their physicochemical features.

Synthesis and pharmacochemical study of new Cu(II) complexes with thiophen-2-yl saturated and alpha,beta-unsaturated substituted carboxylic acids.

Copper complexes with thiophen-2-yl saturated and alpha,beta-unsaturated carboxylic acids as ligands were prepared, characterized and pharmacochemically studied. The available evidence supports a dimeric structure for the complexes of the general formula [Cu2(L)4(MeOH)2] where L are the anions of thiophene 2-carboxylic acid (HL1), 2-(thiophen-2-yl)-acetic acid (HL2), 3-thiophen-2-yl-acrylic acid (HL3), 2-phenyl-3-thiophen-2-yl-acrylic acid (HL4) respectively. The crystal structure of [Cu2(L1)4(MeOH)2] (2) was determined while preliminary X-ray analysis of the copper complex with L4 isolated from MeOH/DMSO solution proved to contain three crystallographically independent dimers of the formula [Cu2(L4)4(MeOH)(dmso)][Cu2(L4)4(MeOH)2][Cu2(L4)4(dmso)2].8MeOH (9). Since lipophilicity is a significant physicochemical property determining distribution, bioavailability, metabolic activity and elimination, we tried to measure experimentally their lipophilicity from RPTLC method. The copper complexes and the ligands (thiophen-2-yl saturated and alpha,beta-unsaturated carboxylic acids) were tested in vitro on: (a) soybean lipoxygenase inhibition, (b) interaction with 1,1-diphenyl-2-picryl-hydrazyl (DPPH) stable free radical, (c) the HO* mediated oxidation of DMSO, (d) inhibition of lipid peroxidation, (e) scavenging of superoxide anion radicals and in vivo for the inhibition of carrageenin-induced rat paw edema. The compounds have shown important antioxidant activity, significant anti-inflammatory activity and potent inhibition of soybean lipoxygenase as a result of their physichochemical features. Complex [Cu2(L1)4(MeOH)2] (2) presents the higher in vivo activity (77.4%) followed by complex [Cu2(L2)4(MeOH)2] (4) (51%). Both complexes are more potent anti-inflammatory agents compared to their respective ligands. Moreover we have performed in vitro studies upon their effect on dsDNA, using adsorptive transfer stripping voltammetry and a dsDNA modified carbon paste electrode. Our conclusions were mainly based upon the effect of the studied compounds on the oxidation signal of guanine and adenine. From the given data it seems that complexes show similarities in their behaviour.

Antioxidant and anti-inflammatory activity of aryl-acetic and hydroxamic acids as novel lipoxygenase inhibitors.

Lipoxygenase plays an essential role in the biosynthesis of the leukotrienes. Leukotrienes, as LO metabolites of arachidonic acid (AA), have been implicated as mediators in the pathophysiology of inflammatory diseases, host defense reactions and to play important role in the propagation of the diseases states, exacerbating the local events and ultimately leading to tissue damage. Simple stable molecules containing the hydroxamic acid functionality have been shown to inhibit 5-lipoxygenase. In fact, several hydroxamates are orally active inhibitors of the enzyme as determined by their ability to block the biosynthesis of leukotriene in vivo. In order to establish the inhibitory utility of simple hydroxamates several omega-phenylalkyl and omega-naphthylalkyl hydroxamic acids were synthesized. In an attempt to expand and delineate these results we tried to synthesize some more for a further pharmacochemical study. Since lipophilicity is a significant physicochemical property determining distribution, bioavailability, metabolic activity and elimination, we tried to determine experimentally their lipophilicity from RPTLC method. The compounds are tested in vitro on: a) soybean lipoxygenase inhibition, b) interaction with 1,1-diphenyl-2-picryl-hydrazyl (DPPH) stable free radical, c) the HO* radical mediated oxidation of DMSO, d) inhibition of lipid peroxidation, e) scavenging of superoxide anion radicals f) interaction with glutathione and g) in vivo for the inhibition of carrageenin induced rat paw edema. The compounds have shown important antioxidant activity, medium anti-inflammatory activity and potent inhibition of soybean lipoxygenase as a result of their physichochemical features.

Evaluation of anti-inflammatory and antioxidant activities of mixed-ligand Cu(II) complexes of dien and its Schiff dibases with heterocyclic aldehydes and 2-amino-2-thiazoline.

A new series of complexes of the type [Cu(dien)(2a-2tzn)Y2] and [Cu(dienXX)(2a-2tzn)Y2] has been tested for anti-inflammatory and antioxidant activity. The tested compounds inhibit significantly the carrageenin induced paw edema (36.4-55.8%) and present important scavenging activities. Although their interaction with the free stable radical DPPH is not high they peroxide anions. Compound 7 is the most potent (55.8%) in the in vivo experiment. Lipophilicity--as RM values and theoretically calculated logP values--has been determined. An attempt to correlate the biological results with their structural characteristics and physicochemical parameters has been done.