Anti-inflammatory and antinociceptive effects of the isatin derivative (Z)-2-(5-chloro-2-oxoindolin-3-ylidene)-N-phenyl-hydrazinecarbothioamide in mice

Several isatin derivatives have shown important biological activities, which have attracted interest from researchers. For this reason, the present study aimed to evaluate the anti-inflammatory and antinociceptive effects of the isatin derivative (Z)-2-(5-chloro-2-oxoindolin-3-ylidene)-N-phenyl-hydrazinecarbothioamide (COPHCT) in mice. Three doses of this compound were tested: 1.0, 2.5, and 5.0 mg/kg. The anti-inflammatory activity was assessed using the carrageenan-induced paw edema model and the zymosan-induced air pouch model. The evaluation of the antinociceptive effect was performed through the formalin test and the acetic acid-induced abdominal writhing test. The paw edema assay demonstrated that all doses of the compound showed a significant reduction of the edema in the second hour evaluated, but a better response was observed in the fourth hour. The zymosan-induced air pouch model indicated that the compound, in all doses, significantly reduced leukocyte migration and total protein concentration levels. In the formalin test, the doses 1.0, 2.5, and 5.0 mg/kg of COPHCT showed activity only in the second phase, with reduction in paw pain time of 73.61, 79.46, and 73.85%, respectively. The number of abdominal writhings decreased with the increasing dose, but only 5.0 mg/kg COPHCT exhibited a significant response, with a reduction of 24.88%. These results demonstrated the ability of this compound to interfere in the anti-inflammatory activity of edema, vascular permeability, and cell migration. In addition, its possible antinociceptive effect may be related to the dose used.

Isatin (2,3-dioxoindole): a competitive inhibitor of Na(+)-dependent lysine uptake in rat instestine.

Isatin (2,3-dioxoindole) competitively inhibited (27-40%) Na(+)-dependent L-lysine uptake in rat intestine. The value of Kt was increased from 3.04 mM in control to 5.88 mM in presence of 10 mM isatin. Effect of isatin on the Na(+)-independent amino acid uptake was insignificant (12-18%). The inhibitory constant (Ki) was 2.8 mM under these conditions. The observed inhibition was unaffected by -SH group reacting agents. Isatin (1-10 mM) inhibited Na+, K(+)-ATPase activity in intestine in vitro, the maximum inhibition (66%) being at 10 mM isatin concentration. But the drug had no effect on enzyme activity under in vivo conditions.

In vivo effects of isatin on certain enzymes, lipids & serotonergic system of rat brain.

Isatin (10 microM) strongly inhibited the activity of rat brain monoamine oxidase-B (MAO-B) in vitro. At millimolar concentrations (1-10 mM) it inhibited brain acetylcholinesterase (AChE) and sodium, potassium-adenosine triphosphatase (Na+, K(+)-ATPase) activity also. However, isatin did not affect these enzymes after both acute and chronic treatments in vivo. Administration of isatin to rats at 300 mg/kg body weight for 2 and 6 h significantly raised brain serotonin levels. Chronic treatment for 20 days resulted in enhanced brain glycolipids and plasmalogen levels. There was no change in the levels of 5-hydroxy indole acetic acid (5 HIAA), phospholipids, cholesterol and gangliosides under these conditions.

Studies on peripheral actions of isatin.

The possible peripheral actions of isatin were studied in vivo and in vitro preparations in different experimental models, using conventional techniques. The results showed spasmogenic responses of isatin on guinea pig, rat and rabbit ileum and fundus of rat stomach. Histamine induced broncho-constriction could be antagonised by isatin. Isatin had cardioinhibitory effect on isolated frog heart and had hypotensive and respiratory depressant activities in dog. Isatin had antidiuretic effect. It was devoid of any effect on inflammation and gastric activities. The present results suggest a possible involvement of heterogenic 5-HT3 receptors in gastrointestinal smooth muscle.

Inhibition of brush border sucrase by indoline 2,3-dione (isatin) in rat small intestine.

Isatin (15-25 mM) inhibited rat brush border sucrase by 40% in presence of Na+ and the inhibition was enhanced to over 60% in sodium free medium. Sucrase inhibition by isatin was dependent on pH. Kinetic analysis revealed a pure capacity type (Vmax-effect) inhibition of sucrase activity by isatin in presence of sodium. But it changed to affinity type (K-effect) in sodium free medium. The value of Ki was around 20-25 mM under these conditions. Enzyme inhibition by isatin was alleviated by increasing Na+ or sucrose concentrations. Other monovalent cations like K+, Li+ and Cs+ were also effective in restoring the enzyme activity to control levels. The effectiveness of the metal ions in alleviating the enzyme inhibition was in the order of Na+ > Cs+ > K+ > Li+.

Effect of pH and sodium ions on intestinal uptake of lysine in rats.

Intestinal uptake of lysine in rats progressively decreased with an increase in pH from 5.2 to 8.5, both in the presence and absence of Na+ ions. At pH 5.2 lysine uptake was 30-35% more than that at neutral pH. Na+ activated lysine uptake by 40-50% at pH 5.2 and it was increased to 110-120% at neutral pH. The observed increase in lysine uptake in response to Na+ and H+ gradients was due to enhanced maximal velocity (Vmax), with little change in affinity constant (Kt). Arrhenius analysis revealed a biphasic curve for lysine uptake with transition temperature (Tc) around 20 degrees C (24 degrees C at pH 5.2 in presence of Na+). The energy of activation (Ea) below (16.1-23.4 Kcal/mole) and above (6.7-8.6 Kcal/mole) the Tc was similar at pH 5.2 and 7.0 both in the presence and absence of Na+ ions. The sensitivity of lysine uptake to various inhibitors was also dependent upon pH and Na+ ions.

Effect of administration of isatin on alkaline phosphatase of various organs of rat.

In vivo administration of isatin (200 mg/kg) significantly lowered the activity of rat kidney alkaline phosphatase after 5 hr but enhanced the activity of rat duodenal and jejunal enzyme after 2 and 5 hr (P less than 0.01). The increased activity of the duodenal and jejunal alkaline phosphatase might be due to the induction of the enzyme by isatin.