Synthesis, Characterization and Anti-epileptic Activity of Thiohydantoin Derivatives

Avast number of biological properties have been recognised to Thiohydantoin derivatives, have attracted continuing interest because of their various biological activities, namely anticonvulsant, hypolipidemic activity, anti-thyroidal activity, and anti-microbial actions. In this look upon, multiple numbers of Thiohydantoin derivatives are synthesised besides anti-epileptic mechanisms have been investigated. A series of 3-substituted 2-thiohydantoin Derivatives were synthesised fromBenzil via Benzil-Benzilic AcidRearrangement, characterised and evaluated for their anti-epileptic activity byMaximal Electroshock Seizure (MES) model for the anti-epileptic activity of synthesised compounds was less when compared to standard. Among the activity, synthesised compounds 1 indicated more anti-epileptic activity. With this con irmation, we revised integrated a series of innovative 3-substituted2-thiohydantoin derivatives with a different substituent on the construction and studied the structural activity relationship. The outcomes indicated with the purpose of all the tested compounds have displayed evident anti-epileptic activity. Among the series compounds, 1 showsmoreanti-epileptic activity thanother synthesised compounds.


INTRODUCTION
The chemistry and biological activity of Thiohydantoins and derivatives have been investigated for more than 150 years. The Thiohydantoin moiety, which is present in various pharmacologically active compounds represents a pharmaceutical signi icance.
Thiohydatoins are sulphur compound of hydantoins with carbonyl groups substitute by sulphur group (Wolters Kluwrs, 2006). Among the Thiohydantoins, 2 Thiohydantoins be imperative acknowledged due to various pharmacological application like antihyperlipidemic, anticancer, anti-thyroidal, antiviral, anti-mutagenic, anti-fungal,anti-bacterial, antiulcer, and antiin lammatory. (Wang et al., 2006). Thiohydantoin and its derived compound are furthermore essential intermediates in the production of several amino acids; besides they are the primary material of different generation of long-lasting high temperature established epoxy resins (Asif, 2014). Several Thiohydantoin derivative is made use of rare consumer commodities such as hair spray foundation, some time ago make the most of drug and photographic ilms (Faghihi et al., 2004). In current work, we description about 3-substituted 2-thiohydantoin was synthesised through benzil-benzilic acid reac-tion (Gangadhar et al., 2013), via Benzil and variety of Alkyl/Aryl-Thiourea and in the presence of 10% sodium hydroxide along with ethanol. Synthesised derivatives were assessing for anti-epileptic activity through Maximal Electroshock Seizure induced epileptic in an animal model. (Kulkarni, 2007)

MATERIALS AND METHODS
All the chemicals and reagents, solvents utilise for this effort were obtained from Merck chemicals Pvt limited Bengaluru, India. All the chemicals and reagents were obtained from an industrial source and used further without puri ication.
Melting points were found out by the electrothermal melting point apparatus with an open capillary tube. The inishing point of the reaction and purity of the compounds was observed by Thin Layer Chromatography. Infrared (IR) spectra were con irmation for the derivative on JASCO 4100 FT-IR using Potassium Bromide pellet disc technique. NMR spectra were con irmation on a Bruker advance spectrometer.

Synthesis
Synthesis of Thiohydantoin derivative reaction was carried out resembling scheme-I Step 1: Synthesis of substituted thiourea To dissolve 2 grams appropriate aryl/alkyl amine and 2 ml of concentrated hydrochloric acid the resulting solution was warmed than add slowly saturated solution of Ammonium thiocyanate (30 grams of Ammonium thiocyanate dissolved 30 ml of water) to the above thermal solution. After addition of saturated solution completely re lux mixture for 45 min. in 250 ml round bottom lask then boil the solution got turbid. The resulting turbid solution was poured in cold water. The resulting thiourea derivatives precipitated was iltered. (Gangadhar et al., 2013) Step 2: synthesis of 2-Thiohydantoins The two grams of Benzil was placed in a 250 ml round-bottomed lask with 1gm N-substituted thiourea derivative than dissolve with support of 24 ml of absolute ethanol, and 2.5 ml of 30% aqueous sodium hydroxide were added gradually to this reaction mixture. In the help of Boiling chips were in addition to the reaction mixture and a condenser was attached following packaging the ground glass joint by way of Te lon tape furthermore two hours heated at110-120 • C under a re lux condenser. This reaction solution was allowed to cool at room temperature and poured the reaction solution in 200m1 of ice-cold water. The reaction mixture was unclear, so the suspended materials were eliminated by iltration. Then the apparent solution was carefully acidi ied with concentrated hydrochloric acid, and the product brought together by vacuum iltration and thoroughly washed with water (Liton and Islam, 2006). Various substituted Thiohydantoin synthesised and their physiochemical properties recorded inTable 2.

Animal selection
Either sex of SD albino rats were acquired from the Karpagam University Experimental Animal House in Coimbatore. The animals were provided standard diet water and chow maintained under the standard procedure of temperature (22±2 0 C) light (12-h light/12-h dark cycle) and humidity (55±5%). The rats were randomly allocated to different treatment and control groups. This experiment was agreed by institutional animal ethical committee (KFMSR/M.Pharm/03/2019-20-IAECNO), under the procedures of (CPSCEA) Committee for the Perseverance of Supervision and Control of Experimental procedure for Animals with every attempt made to minimise pain and distress to the animals.

Grouping of animals
SD albino rats weighing between 200-250g were divided into six groups of six animals each.

Assessment of antiepileptic activity
The Corneal electrodes technique is used for the bilateral delivery system of the electrical stimulus. 50 mA for 0.2 sec of Electroconvulsive shock was provided through a corneal electrode to prompt Hind Limb Tonic Extensor phase (HLTE) in rats. There are ive divisions observed in mice after delivering maximal electroshock.
The ive different phases are A. Flexor

D.
Stupor and E. Death or Recovery are noted and correspondingly the time consumed by mice in each division. After mentioned to delivery, the current output was checked by an exhausting multimeter. The electrical inducement was functional using a stimulator apparatus for six groups of six animals, respectively. The positioning for the anticonvulsant effect was closing down of HLTE within 10 sec after delivery of the shock inducement. Kumar et al. (2013) Delivery of using electroconvulsiometer to rat with the assistance of corneal electrode phases occur after giv-    ing electroshock. Figure 1 shows Graphical representation of the anti-epileptic activity of on maximal electroshock (MES) induced seizures in rats Assessments for anti-epileptic activity Control, Standard drug (Phenytoin), synthesised compounds by MES model are given in Table 3 respectively. (Lopez et al., 2016) The test compound and standard drug were administered orally through 0.5% CMC suspension. After one hour of drug administration, an electrical stimulus of 150mA was applied through trans auricular electrodes for 0.2 seconds with the help of an electroconvulsiometer. The maximal seizure was de ined by the tonic extension of the hind limb to an angle close to 180 0 to the plane of the body axis. The duration of Flexion, Extensor, Convulsion, Stupor and Death or Recovery phase of animals protected from seizures were recorded.

Statistical analysis
The outcomes are expressed in mean ± standard error of the mean. (n=6) Statistical analysis was completed by one technique ANOVA, monitored by control. P vs Dunnett multiple comparisons test.

RESULTS AND DISCUSSION
Various substituted Thiohydantoin was synthesised, and their melting point and analytical data are given below Several thioureas were designed by using corresponding aromatic amine with Ammonium Thiocyanate than simple base catalysed condensation of Benzil with Thiourea. The formation of 3-substituted 2-thiohydantoin derivatives these products follows the benzil-benzilic acid rearrangement that is shown in the Scheme I. Purity of all designed compounds was checked by Thin Layer Chromatography and Melting Point. All newly synthesised compounds were investigated by Fourier Transform Infrared, Mass Spectroscopy(EI), 1HNMR. The anti-epileptic activity was carryout by using maximal Electroshock seizure induced epileptic in the rat model. Phenytoin is used as an anti-epileptic activity standard drug. Results outcome are listed in Figure 1 and Table 3 displayed that all newly synthesised compounds of 3-substituted 2-thiohydantoin derivatives possess considerable Anti-Epileptic activity-compounds one more potent anti-epileptic activity than other newly synthesised compounds.

CONCLUSIONS
The simple appropriate technique for the synthesis of 2-thiohydantoins was improved on benzilbenzilic acid rearrangement. Synthesised derivatives were con irmed by Fourier Transform Infrared, Mass Spectroscopy (EI), 1HNMR, all synthesised compounds of 3-substituted 2-thiohydantoin displays noticeable anti-epileptic activity. Lipophilic groups such as the amino group in the para position of 3 phenyl ring displays increase the activity, while hydrophilic groups such as carboxyl on Para position of 3 phenyl ring outcome decrease in an anti-epileptic activity. Further wide-ranging study is required to authorise to the achievement of antiepileptic agents in pharmaceutical ield and studies to enhance the effectiveness of Thiohydantoin derivatives.