Synthesis of Some New Kind of Antipyrine Carbo(thio)amide and 1,2,4-Triazole Derivatives and Comparing Their Anti-tubercular Activities

Bu calismada antipirin (4-Aminoantipirin) baslangic maddesi olarak secilerek karbotiyoamit ve 1,2,4-triazolturevi bilesikler gerceklestirilerek anti-tuberkuler aktiviteleri karsilastirilmistir. 4-aminoantipirin sirasiyla etilbromoasetat, hidrazinhidrat ve cesitli izo(tiyo)siyanatlar ile muamelesi sonucu bazik ortamda reaksiyonu ile halkalasma reaksiyonu gerceklesmesi ile yapiya ikinci bir halka olan 1,2,4-triazol halkasinin ilavesi gerceklestirildi. Sentezlenen yeni antipirin turevi bilesiklerin yapilari IR, NMR ve kutle spektrofotometresi gibi spektroskopik yontemlerle aydinlatildi. Sentezlenen yeni bilesikler, Gram olmayan bakteriye karsi aktivite calismalari gerceklestirilmistir.


Introduction
Antipyrine (2,3-dimethyl-1-phenyl-3-pyrazolin-5-one) is the first pyrazole derivative to be used in pain and inflammation conditions and is active in many studies. Bioactive compounds obtained using this compound, also called 4-aminoantipyrine (1), are known in the literature (Jain et al. 2003, Gürsoy et al. 2000, Turan-Zitouni et al. 2001, Abu Elmaati 2002, Abdel-Latif 2006. Antipyrinederived compounds; analgesics (Filho et al. 1998, Sondhi et al. 1999, anti-inflammatory (İsmail et al. 2007), antimicrobial (Mishra 1999, Raman et al. 2002, Raman et al. 2004) and anticancer (Sondhi et al. 2001) are compounds with bioactive properties (Bondock et al. 2008, Rostom et al. 2009 The synthesis of heterocyclic compounds containing five-membered rings has become increasingly important in recent years due to their pharmacological properties. Examples of such compounds include Vorozol (2), Letrozole (3), Anostrozole (4) and Itraconazole (5), which are currently used in cancer treatment and contain an azole ring (Sun et al. 2004, Verrect et al. 2003. Pathogenic microorganisms have rapidly led to new drug discoveries due to their resistance to existing drugs. And this is of interest to medical chemists. Therefore, synthesis of simple or complex triazole compounds is important for the discovery of new drugs (Rostom et al. 2003, Turan-Zitouni et al. 1999, Kolavi et al. 2006, Holla et al. 2001, Demirbaş et al. 2004, Demirbaş et al. 2002. For this purpose, many working groups have begun to design and synthesize compounds containing triazole rings bearing different functional groups. The 1,2,4-triazole ring is included in the structure of many more therapeutically important drugs. For example; Fluconazole (9) (antifungal), Ribavirin (10) (antiviral), Rizatriptan (11) (antimigren), Alprazolam (12) (anxiolytic) are the best examples. It has been reported in the literature that these compounds have different biological and medicinal properties (Chandra et al. 2006, Dixit et al. 2006, İkizler et al, 1996, İkizler et al. 1999 Novel 1,2,4-triazole analogs were synthesized and antitubercular activity results were evaluated with different pharmacophores (Figure 1 and Figure 2). Compound 5c which contain an allylic group at N-4 atom of 1,2,4-triazole ring showed excellent antitubercular activity compared with Streptomycin standard drug. Compound 5b also showed equivalent antitubercular activity to the standard drug. Carbothioamide derivatives of compound 4b and 4c showed good-moderate activity against Mycobacterium smegmatis test microorganism. As a result, it has seen that the allylic and phenyl group of carbothioamide or 1,2,4-triazole derivatives containing antipyrine group showed very good antitubercular activity.

General Methods for Chemistry
All the chemicals used in this publication were obtained from Sigma-Aldrich and Merck without further purification. Melting points of the synthesized new compounds were obtained by using capillary tube in Stuart Brand SMP apparatus. Reaction times and purities were determined by thin layer chromatography. Infrared spectra were obtained by ATR apparatus on Perkin Elmer brand and 1600 serial IR devices. NMR spectra of the compounds were obtained from BRUKER AVENE

II 400 MHz instrument in Karadeniz Technical University or Giresun University Central Research
Laboratories. Mass spectra of the compounds were also obtained from Agilent Technologies branded 1260 Infinity 6230 TOF LC / MS model device at Giresun University Central Research Laboratory.
The triethylammonium salt formed after this time was removed by filtration. The solvent was evaporated under reduced pressure to precipitate the resulting oily substance from a mixture of nbutylacetate: ether (1: 2) to form a solid. The crude product obtained was purified by crystallization

General Synthesis of Compounds 4a-4e:
To a solution of compound 3 (10 mmol) in absolute dichloromethane, phenyl isothiocyanate (for compound 4a), benzyl isothiocyanate (for compound 4b) or allyl isothiocyanate (for compound 4c) (20 mmol) was added dropwise and the mixture was stirred at room temperature for 24 hours.
After this time, the precipitated solid was filtered off and purified by crystallization in the appropriate solvents.

General Synthesis of Compounds 5a-5e
Compound 4 and NaOH solution (2 %) were refluxed for 4 hours. Then, the crude mixture was acidified to pH 4 with 37% HCl by cooling to room temperature. The resulting solid was filtered and purified by crystallization from the appropriate solvents.

Procedure of Minimal Inhibition Concentration (MIC)
To determine the minimum amount of substance that exhibits antimicrobial activity, the microdilution fluid method is applied in liquid medium and the minimum inhibition concentration (MIC) is determined as micrograms / milliliters (µg / mL) (Woods et al. 2003

Results and Discussion
In this study, antipyrine (1) was chosen as the starting material and the corresponding carbo(thio)amide (4a-4e) derivatives were obtained using different iso(thio)cyanates. Subsequently, the corresponding 1,2,4-triazole derivatives (5a-5e) were synthesized by cyclic reaction in basic medium. Anti-tubercular activities of all compounds obtained were examined. Of these, compound 5c with allyl group in structure has shown better anti-tubercular activity (1.95 µg / mL) even than the standard drug streptomycin (4 µg / mL). In addition, compound 5b with the benzyl group in structure, showed equivalent activity against Mycobacterium smegmatis (Ms) to standard drug streptomycin.
The carbothioamide compound 4c having allyl group in its structure showed also moderate antitubercular activity (Table 1).

Conclusions and Recommendations
This study covers the synthesis of 1,2,4-triazole derivatives with different pharmacophores and investigation of their anti-tubercular activity. Compound 5b and 5c showed excellent activity againts the test microorganism of Ms compared with standard drug Streptomycin.