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Acta Chim. Slov. 2016, 63, 18-25
DOI: 10.17344/acsi.2015.1555
Scientific paper
Synthesis, Characterization and Antimicrobial Activities of Some New Heterocyclic Schiff Bases Derived from Thiocarbohydrazide
Kamelia M. El-Mahdy,1* Azza M. El-Kazak,1 Mohamed Abdel-Megid,12 Magdy Seada1 and Osama Farouk1
1 Department of Chemistry, Faculty of Education, Ain Shams University, Roxy, 11711, Cairo, Egypt
2 Hurymilla College of Science and Humanities, Shaqra University, Kingdom of Saudi Arabia
* Corresponding author: E-mail: kmelmahdy@gmail.com
Received: 24-03-2015
Abstract
The reaction of pyrazolobenzothienopyrimidine-3-carbaldehyde 1 with thiocarbohydrazide afforded the Schiff's base 3. The latter compound reacted with some electrophilic reagents to give 1,2,4-triazoles 4-6 and 1,2,4-triazines 7-9. Treatment of compound 3 with 2-cyano-3,3-bis(methylthio)acrylonitrile gave the corresponding 5-amino-4-cyano-3-methylthiopyrazole derivative 11. The reaction of pyrazole 11 with carbon disulfide afforded dithioxopyrazolopyrimidi-ne 12. Acylation of compound 11 by using acetic anhydride yielded acetamide 13. On the other hand, the cyclocondensation of pyrazole 11 with acetic anhydride in pyridine yielded pyrazolopyrimidine derivative 14. The reactivity of compound 11 towards formamide and phenylisothiocyanate to give the pyrazolopyrimidines 15 and 16 was studied. The newly synthesized compounds were screened for their antimicrobial activity.
Keywords: Schiff bases; triazines, triazoles; carbothiohydrazide; antimicrobial activity.
1. Introduction
Schiff bases1'2 of heterocyclic compounds are interesting class of organic compounds possessing a wide spectrum of biological activities, such as antibacterial, antitubercular, antiinflammatory, anthelmintic, antiviral and antioxidant activities. Study of thiocarbohydrazide and its derivatives is of great interest due to the wide use of these compounds and their valuable reactions. Thiocarbohydra-zides are an important class of compounds which possess applications in many fields. The chemistry of thiocar-bohydrazides has gained increased interest in both synthetic organic chemistry and biological fields and has considerable value in many useful applications, such as the assessment process of the three-dimensional ultra structure examination techniques of interphase nuclei and tissues, besides their therapeutic importance.3 Thiocarbohydra-zide derivatives have attracted much attention in recent years due to their applications in the synthesis of transition metal complexes.4-6 Thiocarbohydrazide7-9 is the clo-
sest structural analogue of thiosemicarbazide, derivatives of which are recommended as effective antitubercular and antiviral preparations. Thiocarbohydrazides of the aromatic series exhibit high antiviral and antimicrobial activity.10-12 In the view of these observations, we planned to synthesis some novel Schiff bases and to evaluate for antimicrobial activity.
2. Results and Discussion
2. 1. Chemistry
Novel Schiff bases were obtained from condensation of thiocarbohydrazide with substituted aldehyde giving the monothiocarbohydrazone. Thus, condensation reaction of 10-oxo-4,6,7,8,9,10-hexahydroprazolo[1,5-a] [1]benzothieno[2,3-d]pyrimidine-3-carbaldehyde (1)13 with thiocarbohydrazide 214 in boiling absolute ethanol containing a catalytic amount of acetic acid gave the corresponding N'-[(10-oxo-4,6,7,8,9,10-hexahydropyrazolo
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o	s
cxiy *
HN	NH EtOH
CHO
NH, NH,
AcOH
O
CXiy
H	H	h
vy
NH,
Scheme 1. Synthesis of thiocarbohydrazone 3.
[ 1,5-a][1 ]benzothieno[2,3-d]pyrimidin-3-yl)methylene] thiocarbohydrazide (3) (Scheme 1).
The structure of the isolated product was confirmed on the basis of its elemental analysis and spectral data. Thus, the IR spectrum of compound 3 showed absorption bands at 1194, 1684, and 3198-3422 cm-1 corresponding to C=S, C=O, NH and NH2 functions, respectively. Its 1H NMR spectrum showed four D2O-exchangeable signals at 5 3.32, 9.50, 11.36 and 12.05 ppm assigned to NH2 and three NH protons, respectively.
Further, it is of interest to note that the thio-carbohydrazide 3 is a convenient intermediate for the synthesis of a novel isolated heterocyclic system. Thus, heating thiocarbohydrazone 3 under reflux with benzoyl chloride in pyridine gave 3-{[(3-phenyl-5-thioxo-1,5-dihydro-4-H-1,2,4-triazol-4-yl)imino]methyl}-6,7,8,9-te-trahydropyrazolo[1,5-a][1]benzothieno[2,3-d]pyrimidin-10(4H)-one (4), while boiling with acetic anhydride afforded 3- {[(3-methyl-5-thioxo-1,5-dihydro-4-H-1,2,4-tria-zol-4-yl)imino]methyl}-6,7,8,9-tetrahydropyrazolo[1,5-a][1]benzothieno[2,3-d]pyrimidin-10(4H)-one (5). Also, reaction of 3 with carbon disulfide in boiling pyridine afforded 3-{[(3,5-dithioxo-1,2,4-triazolidin-4-yl)imino] methyl}-6,7,8,9-tetrahydropyrazolo[1,5-a][1]benzothie-no[2,3-d]pyrimidin-10(4H)-one (6) (Scheme 2). The structures of compounds 4-6 were confirmed by their elemental analysis and spectral data.
Moreover, compound 3 was used for construction of some new Schiff bases bearing 1,2,4-triazine moiety through the reaction with 1,2-bifunctional electrophiles. Thus, heterocyclization of 3 with chloroacetyl chloride, oxalyl chloride and/or sodium pyruvate led to the formation of 3-{[(6-oxo-3-thioxo-1,2,4-triazinan-4-yl)imi-no]methyl}-6,7,8,9-tetrahydropyrazolo[1,5-a][1]benzot-hieno[2,3-d]pyrimidin-10(4H)-one (7), 3-{[(5,6-dioxo-3-thioxo-1,2,4-triazinan-4-yl)imino]methyl}-6,7,8,9-te-trahydropyrazolo[1,5-a][1]benzothieno[2,3-d]pyrimidin-10(4H)-one (8) and 3-{[(6-methyl-5-oxo-3-thioxo-2,5-dihydro-1,2,4-triazin-4(3H)-yl)imino]methyl}-6,7,8,9-te-trahydropyrazolo[1,5-a][1]benzothieno[2,3-d]pyrimidin-10(4H)-one (9), respectively (Scheme 3).
The structures of compounds 7-9 were characterized from their spectroscopic as well as elemental analytical data. Thus, the IR spectrum of compound 7 revealed absorption bands at 1171, 1674 and 3200-3415 cm-1 corresponding to C=S, C=O and NH functions, respectively. Its 1H NMR spectrum showed D2O-exchangeable signals at 5 7.16, 7.37 and 7.50 ppm assigned to three NH protons. The mass spectrum of compound 8 revealed the molecular ion peak at m/z 415 corresponding to the molecular formula C16H13N7O3S2, which agrees well with the molecular weight (415.44). The mass spectrum of compound 9 revealed the molecular ion peak at m/z 413 corresponding to the molecular formula C17H15N7O2S2, which agrees well with
Scheme 2. Formation of 1,2,4-triazole derivatives 4, 5 and 6.
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" Wn-N FH pyridine
yn
CICH-COCI
N-N
H L	NH CICOCOCt
Jj dioxane/prp.
MeCOCOONa
gl.AcOH
I ÏTiï
Me
un
î
9	/
Scheme 3. Formation of 1,2,4-triazine derivatives 7, 8 and 9.
Scheme 4. Synthesis of aminocyanopyrazole 11.
the molecular weight (413.47) and supports the identity of the structure and the base peak at m/z 273 (100%).
Thiocarbohydrazide 3 was treated with 2-cyano-3,3-bis(methylthio)acrylonitrile 10 in dimethylformamide containing a catalytic amount of triethylamine to give 5-amino-4-cyano-3-(methylthio)-N-[(10-oxo-4,6,7,8,9,10-hexahydropyrazolo[1,5-a][1]benzothieno[2,3-d]pyrimi-din-3-yl)methylene] -1H-pyrazole-1-carbothiohydrazide (11) (Scheme 4).
The IR spectrum of compound 11 showed absorption bands at 1182, 1654, 2204 and 3200-3446 cm-1 corresponding to C=S, C=O, C=N, NH and NH2 functions, respectively. Its 1H NMR spectrum revealed signals at 5 2.41 and 9.86 due to SCH3 and CH=N protons, respectively. It showed also three D2O-exchangeable signals at 5 3.34, 8.30 and 8.42 ppm assigned to NH2 and two NH protons, respectively. Also, the structure of 11 was confirmed by its mass spectrum which exhibited the molecular ion peak at m/z 483 with a base peak at m/z 80 (100%).
Heating 11 with an excess of carbon disulfide in pyridine afforded 3-(methylthio)-N-[(10-oxo-4,6,7,8, 9,10-he-xahydropyrazolo[ 1,5-a][ 1 ]benzothieno[2,3-d]pyrimidin-3-yl)methylene]-4,6-dithioxo-4,5,6,7-tetrahydro-1H-pyrazo-lo[3,4-d]pyrimidine-1-carbothiohydrazide (12). Acylation of carbothiohydrazide 11 by using acetic anhydride yielded the amide, N-[4-cyano-3-(methylthio)-1-{[2-(10-oxo-4,6,7,8,9,10-hexahydropyrazolo[1,5-a][1]benzothieno[2,3-d]pyrimidin-3-yl)methylene]hydrazino} carbonothioyl]-1H-pyrazol-5-yl]acetamide (13). Cyclocondensation of
carbothiohydrazide 11 with acetic anhydride in pyridine gave 6-methyl-3-(methylthio)-N-[(10-oxo-4,6,7,8,9,10-he-xahydropyrazolo[1,5-a][1]benzothieno[2,3-d]pyrimidin-3-yl)methylene]-4-oxo-4,5-dihydro-1H-pyrazolo[3,4-d]pyri-midine-1-carbothiohydrazide (14) (Scheme 5).
Scheme 5. Reaction of 11 with carbon disulfide and/or acetic anhydride.
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The structures of compounds 12-14 were determined using spectroscopic as well as elemental analytical data. Thus, no evidence for the presence of an amino (NH2) group was seen in both the IR and NMR spectra. The IR spectrum of compound 13 showed absorption bands at 1254, 1701, 1684, 2205 and 3447 cm-1 corresponding to C=S, C=O, C=N and NH functions, respectively. Its 1H NMR spectrum revealed signals at 5 2.20 and 2.42 due to COCH3 and SCH3 protons, respectively. It also showed three D2O-exchangeable signals at 5 7.82, 8.10 and 8.13 ppm assigned to three NH protons.
Interaction of carbothiohydrazide 11 with formami-de and phenyl isothiocyanate yielded 4-amino-3-(methylt-hio)-N-[(10-oxo-4,6,7,8,9,10-hexahydropyrazolo[1,5-a]
s
Scheme 6. Formation of pyrazolo[3,4-d]pyrimidine derivatives 15 and 16.
[1]benzothieno[2,3-d]pyrimidin-3-yl)methylene]-1H-pyrazolo[3,4-d]pyrimidine-1-carbothiohydrazide (15) and 4-imino-3-(methylthio)-5-phenyl-N-[(10-oxo-4,6,7,8, 9,10-hexahydropyrazolo[1,5-a][1]benzothieno[2,3-d] pyrimidin-3-yl)methylene]-6-thioxo-4,5,6,7-tetrahydro-1H-pyrazolo[3,4-d]pyrimidine-1-carbothiohydrazide (16), respectively (Scheme 6).
The 1H NMR spectrum of compound 15 showed a characteristic singlet at 5 7.62 assigned to pyrimidine-H-6, in addition to three D2O-exchangeable signals at 5 3.38, 7.52 and 7.99 ppm attributed to NH2 and two NH protons, respectively. Also, the structure of 15 was confirmed by its mass spectrum which exhibited the molecular ion peak at m/z 510 (M+, 2%) with a base peak at m/z 336 (100%). On the other hand, the 1H NMR spectrum of compound 16 showed four D2O-exchangeable signals at 5 8.06, 8.20, 8.33 and 8.38 ppm assigned to four NH protons, in addition to an aromatic multiplet in the region 5 6.96-7.60. The structure of 16 was also supported by its mass spectrum which showed the molecular ion peak at m/z 618 (7.5%) corresponding to the molecular formula C26H22N10OS4, which agrees well with the molecular weight (618.77) and supports the identity of the structure and the base peak at m/z 336 (100%).
2. 2. Biological Activities
The standardized disc agar diffusion method15 was followed to determine the activity of the synthesized compounds against the sensitive organisms Staphylococcus
Table 1. The antimicrobial activity of the synthesized compounds
Mean of zone diameter , nearest whole mm. Gram-positive bacteria	Gram-negative bacteria
Comp. No.	S. aureus	B. subtilis	S. typhimurium	E. coli
1	0.5	1	0.5	1	0.5	1	0.5
mg/mL mg/mL mg/mL mg/mL	mg/mL mg/mL mg/mL mg/mL
Fungi C. albicans 1	0.5
mg/mL mg/mL
3	-	-	20	15	-	-	-	-	14	10
4	-	-	-	-	-	-	-	-	24	19
5	-	-	-	-	-	-	-	-	18	11
6	-	-	-	-	-	-	-	-	23	17
7	-	-	-	-	-	-	-	-	23	19
8	21	15	21	16	21	18	17	10	20	14
9	-	-	21	15	13	10	-	-	27	24
11	-	-	8	7	-	-	16	13	-	-
12	-	-	18	15	-	-	-	-	-	-
13	16	10	9	7	-	-	-	-	19	15
14	-	-	-	-	-	-	-	-	16	12
15	16	13	18	16	18	14	-	-	28	24
16	17	13	8	7	-	-	-	-	23	17
Control #	35	26	35	25	36	28	38	27	35	28
= Calculated from 3 values. - = No effect. 7-10 mm: Low activity = Mean of zone diameter < 1/3 of mean zone diameter of control. 11-20: Intermediate activity = Mean of zone diameter < 2/3 of mean zone diameter of control. 21-28: High activity = Mean of zone diameter > 2/3 of mean zone diameter of control. #: Chloramphenicol in the case of Gram-positive bacteria, Cephalothin in the case of Gram-negative bacteria and cyclohe-ximide in the case of fungi.
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aureus and Bacillus subtilis as Gram-positive bacteria, Salmonella typhimurium and Escherichia coli as Gramnegative bacteria and Candida albicans as fungus strain. The compounds were dissolved in DMSO which has no inhibition activity to obtain concentration of 100 pg mL-1. The test was performed on medium potato dextrose agars (PDA) which contains infusion of 200 g potatoes, 6 g dextrose and 15 g agar.16 Uniform size filter paper disks (3 disks per compound) were impregnated by equal volume (10 pL) from the specific concentration of dissolved tested compounds and carefully placed on inoculated agar surface. After incubation for 36 h at 27 °C in the case of bacteria and for 48 h at 24 °C in the case of fungi, inhibition of the organisms was measured and used to calculate mean of inhibition zones.
The results depicted in Table 1 show various activities against all species of microorganisms which suggest that the variations in the structures affect the growth of the microorganisms. Thus, we can conclude from these results that some of the prepared compounds showed a low to high antimicrobial activity towards Gram-positive bacteria, Gram-negative bacteria and the fungal strain (Table 1).
3. Experimental
3. 1. General
Melting points are uncorrected and were recorded in open capillary tubes on a Stuart SMP3 melting point apparatus. Infrared spectra were recorded on FT-IR Jasco 4100 spectrophotometer using a KBr wafer technique. The 1H and 13C NMR spectra were recorded in DMSO-d6 or CDCl3 on Gemini spectrometer (300 MHz for 1H NMR and3 75 MHz for 13C NMR) and the chemical shift ar given as 5 downfield from TMS as an internal standard, measured at the Main Laboratories of the War Chemical. Elemental microanalyses were recorded on a Per-kin Elmer series II CHNS analyzer 2400. Mass spectra were obtained using gas chromatography GCMS qp-2010 and on a Shimadzu instrument mass spectrometer (70 eV) at the Cairo University Microanalytical Center. 10-0xo-4,6,7,8,9,10-hexahydroprazolo[1,5-a][1]benzot-hieno[2,3-d]pyrimidine-3-carbaldehyde (1) has been prepared according to the reported method.13
3. 1. 1. Synthesis of N'-[(10-oxo-4,6,7,8,9,10-
hexahydropyrazolo[1,5-a][1]benzothieno [2,3-d]pyrimidin-3-yl)methylene] thiocarbohydrazide (3)
A mixture of compound 1 (0.273 g, 1 mmol) and
thiocarbohydrazide 2 (0.106 g, 1 mmol) in absolute etha-nol (10 mL) containing a few drops of acetic acid was heated under reflux for 1 h. After cooling, the precipitated solid was filtered off, air dried and recrystallized from ethanol to give 3 as yellow crystals. Yield 81%, mp
200-202 °C. IR (KBr) v 3422-3198 (NH2, NH), 3069 (CHarom), 2933 (CH ), 1684 (C=0), 1611 (C=N), 1194 (C=S) cm-1. 1H NMR (DMS0-d6) 5 1.80-1.82 (m, 4H, 2CH2), 2.72 (t, 2H, CH2), 2.94 (t, 2H, CH2), 3.32 (brs, 2H, D20-exchangeable, NH2), 8.12 (s, 1H, CH=N), 8.26 (s,
IH,	pyrazole-H2), 9.50 (s, 1H, D20-exchangeable, NH),
II.36	(s, 1H, D20-exchangeable, NH), 12.05 (s, 1H, Dp-exchangeable, NH). MS: m/z 361 (M+, 0.4%). Anal. Calcd for C14H15N70S2: C, 46.52; H, 4.18; N, 27.13; S, 17.74. Found: C, 4(5.51; H, 4.16; N, 27.15; S, 17.75.
3. 1. 2. Synthesis of 3-{[(3-phenyl-5-thioxo-1,5-dihydro-4.ff-1,2,4-triazol-4-yl)imino] methyl}-6,7,8,9-tetrahydropyrazolo [1,5-a][1]benzothieno[2,3-d] pyrimidin-10(4.ff)-one (4)
A mixture of compound 3 (0.361 g, 1 mmol) and benzoyl chloride (0.12 mL, 1 mmol) was added dropwise with continuous stirring to dry pyridine (10 mL). The reaction mixture was heated under reflux for 4 h. After cooling, the reaction mixture was poured onto ice/HCl. The solid obtained was filtered off, washed several times with water, and recrystallized from acetic acid to give 4 as pale violet crystals. Yield 81%, mp 229-230 °C. IR (KBr) v 3444, 3125 (2NH), 3020 (CHarom), 2937 (CHaliph), 1682 (C=0), 1627 (C=N), 1217 (C=S) cm-1. 1H NMR (DMS0-d6) 5 1.79-1.82 (m, 4H, 2CH2), 2.66 (t, 2H, CH2), 2.90 (t, 2H, CH2), 6.09 (s, 1H, D20-exchangeable, NH), 7.48-8.41 (m, 6H, ArH's and pyrazole-H2), 8.08 (s, 1H, D20-exchangeable, NH), 8.83 (s, 1H, CH=N). MS: m/z 4428 (M++1, 4.2%). Anal. Calcd for C21H17N70S2: C, 56.36; H, 3.83; N, 21.91; S, 14.33. Found: C, 56.35; H, 3.82; N, 21.92; S, 14.32.
3. 1. 3. Synthesis of 3-{[(3-methyl-5-thioxo-
1,5-dihydro-4.ff-1,2,4-triazol-4-yl)imino] methyl}-6,7,8,9-tetrahydropyrazolo [1,5-a][1]benzothieno[2,3-rf] pyrimidin-10(4.ff)-one (5)
A mixture of compound 3 (0.361 g, 1 mmol) and acetic anhydride (10 mL) in dry pyridine (5 mL) was heated under reflux for 2 h. After cooling, the reaction mixture was poured onto ice. The solid obtained was filtered off, washed several times with water, and recrystallized from DMF to give 5 as pale brown crystals. Yield 63%, mp >300 °C. IR (KBr) v 3421, 3200 (2NH), 3069 (CHarom), 2925 (CH^), 1653 (C=0), 1609 (C=N), 1183 (C=S) cm-1. 1H NMR (DMS0-d6) 5 1.76-1.91 (m, 4H, 2CH2), 2.03 (s, 3H, CH3), 2.763 (t, 2H, CH2), 2.88 (t, 2H, CH2), 8.30 (s, 1H, pyrazole-H2), 8.53 (s, 1H, D20-exchangeable, NH), 8.80 (s, 1H, CH=N), 9.11 (s, 1H, D20-exchangeable, NH). MS: m/z 385 (M+, 0.2%). Anal. Calcd for C16H15N70S2: C, 49.85; H, 3.92; N, 25.44; S, 16.64. Found: C, 49.86; H, 3.91; N, 25.43; S, 16.65.
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3. 1. 4. Synthesis of 3-{[(3,5-dithioxo-1,2,4-
triazolidin-4-yl)-imino]methyl}-6,7,8,9-tetrahydropyrazolo[1,5-a][1]benzothieno [2,3-d]pyrimidin-10(4#)-one (6)
A mixture of compound 3 (0.361 g, 1 mmol) and carbon disulfide (0.06 mL, 1 mmol) in dry pyridine (10 mL) was heated under reflux for 8 h or until evolution of H2S ceased. After cooling, the reaction mixture was poured onto ice/HCl. The solid obtained was filtered off, washed several times with water, and recrystallized from DMF/ethanol to give 6 as pale brown crystals. Yield 58%, mp >300 °C. IR (KBr) v 3440-3280 (3NH), 3050 (CH^), 2929 (CH^), 1660 (C=O), 1606 (C=N), 1181 (2C=S) cm-1. 1H NMR (DMSO-d6) 5 1.76-1.80 (m, 4H, 2CH2), 2.72 (t, 2H, CH2), 2.88 (t, 2H, CH2), 6.96 (s, 1H, D2O-exchangeable, NH), 7.13 (s, 1H, D2O-exchangeable, NH), 8.38 (s, 1H, D2O-exchangeable, N2H), 8.90 (s, 1H, pyrazole-H2), 9.89 (s, 1H, CH=N). MS: m/z 403 (M+, 0.5%). Anal. Calcd for C15H13N7OS3: C, 44.65; H, 3.25; N, 24.30; S, 23.84. Found: C, 44.66; H, 3.24; N, 24.31; S, 23.83.
3. 1. 5. Synthesis of 3-{[(6-oxo-3-thioxo-1,2,4-triazinan-4-yl)imino]methyl}-6,7,8,9-tetrahydropyrazolo[1,5-a][1]benzothieno [2,3-d]pyrimidin-10(4#)-one (7)
A mixture of compound 3 (0.361 g, 1 mmol) and chloroacetyl chloride (0.113 mL, 1 mmol) in dry pyridine (10 mL) was heated under reflux for 6 h. After cooling, the reaction mixture was poured onto ice/HCl. The solid obtained was filtered off and recrystallized from dioxane to give 7 as pale brown crystals. Yield 63%, mp >300 °C. (KBr) v 3415-3200 (3NH), 3055 (CHarom), 2932 (CHaliph), 1674 (2C=O), 1619 (C=N), 1171 (C=S) cm-1. 1H Nlipl^ (DMSO-d6) 5 1.77-1.90 (m, 4H, 2CH2), 2.65 (t, 2H, CH2), 2.88 (t, 2H, CH2), 4.37 (s, 2H, CH2), 7.16 (s, 1H, D20-exchangeable, NH), 7.37 (s, 1H, D2O-exchan-geable, NH), 7.50 (s, 1H, D2O-exchangeable, NH), 8.51 (s, 1H, pyrazole-H2), 8.88 (s, 1H, CH=N). MS: m/z 401 (M+, 0.1%). Anal. Calcd for C16H15N7O2S2: C, 47.87; H, 3.77; N, 24.42; S, 15.97. Found: C, 47.86; H, 3.78; N, 24.43; S, 15.96.
3. 1. 6. Synthesis of 3-{[(5,6-dioxo-3-thioxo-1,2,4-triazinan-4-yl)imino]methyl}-6,7,8,9-tetrahydropyrazolo[1,5-a][1]benzothieno [2,3-d]pyrimidin-10(4#)-one (8)
A mixture of compound 3 (0.361 g, 1 mmol) and ox-alyl chloride (0.129 mL, 1 mmol) was added dropwise in dry dioxane (20 mL) containing catalytic amount of pipe-ridine was heated under reflux for 6 h. After cooling, the reaction mixture was poured onto ice. The solid obtained was filtered off, washed several times with water, and recrystallized from DMF/H2O to give 8 as brown crystals. Yield 61%, mp >300 °C. IR (KBr) v 3420-3200 (3NH),
3050 (CHarom), 2931 (CHaliph), 1640 (3C=O), 1616 (C=N), 1185 (C=S) cm-1. H NMR (DMSO-d6) 5 1.79-1.90 (m, 4H, 2CH2), 2.65 (t, 2H, CH2), 2.93 (t, 2H, CH2), 6.09 (s, 1H, D2O-exchangeable, NH), 7.86 (s, 1H, pyrazole-H2), 8.73 (s, 1H, D2O-exchangeable, NH), 9.93 (s, 1H, CH=N), 11.21 (s, 1H, D2O-exchangeable, NH). MS: m/z 415 (M+, 0.3%). Anal. Calcd for C16H13N7O3S2: C, 46.26; H, 3.15; N, 23.60; S, 15.44. Found: C, 436.25; H, 3.14; N, 23.61; S, 15.45.
3. 1. 7. Synthesis of 3-{[(6-methyl-5-oxo-3-thioxo-2,5-dihydro-1,2,4-triazin-4(3#)-yl)imino] methyl}-6,7,8,9-tetrahydropyrazolo[1,5-a] [1]benzothieno [2,3-d]pyrimidin-10(4.ff)-
one (9)
A mixture of compound 3 (0.361 g, 1 mmol) and sodium pyruvate (0.11 g, 1 mmol) in glacial acetic acid (10 mL) was heated under reflux for 4 h. After cooling, the reaction mixture was poured onto crushed ice. The solid obtained was filtered off, washed several times with water, and recrystallized from acetic acid to give 9 as brown crystals. Yield 71%, mp >300 °C. IR (KBr) v 3453, 3243 (2NH), 3055 (CHarom), 2934 (CH^), 1689, 1642 (2C=O), 1607 (C=N), 1199 (C=S) cm-1. 1H NMR (DMSO-d6) 5 1.78-1.90 (m, 4H, 2CH2), 2.16 (s, 3H, CH3), 2.67 (t, 2H, CH2), 2.88 (t, 2H, CH2), 8.37 (s, 1H, D2O-exchangeable, NH), 8.56 (s, 1H, pyrazole-H2), 9.80 (s, 1H, CH=N), 13.71 (s, 1H, D2O-exchangeable, NH). MS: m/z 413 (M+, 1.2%). Anal. Calcd for C17H15N7O2S2: C, 49.38; H, 3.66; N, 23.71; S, 15.51. Found: C, 49.37; H, 3.65; N, 23.72; S, 15.52.
3. 1. 8. Synthesis of 5-amino-4-cyano-3-
(methylthio)-N-[(10-oxo-4,6,7,8,9,10-hexahydropyrazolo[1,5-a][1]benzothieno [2,3-d]pyrimidin-3-yl)methylene]-1.ff-pyrazole-1-carbothiohydrazide (11)
A mixture of compound 3 (0.361 g, 1 mmol) and 2-cyano-3,3-bis(methylthio)acrylonitrile 10 (0.17 g, 1 mmol) in DMF (10 mL) containing two drops of triethyla-mine was heated under reflux for 4 h. After cooling, the reaction mixture was poured onto ice. The solid obtained after cooling was filtered, washed with ethanol, and recry-stallized from DMF/ethanol to give 11 as violet crystals. Yield 75%, mp 145-147 oC. IR (KBr) v 3446-3200 (NH2, 2NH), 3053 (CHarom), 2931 (CH^), 2204 (C=N), 1654 (C=O), 1630 (C=N), 1182 (C=S) cm-1. 1H NMR (DMSO-d6) 5 1.79-1.82 (m, 4H, 2CH2), 2.41 (s, 3H, SCH3), 2.73 (t, 2H, CH2), 2.92 (t, 2H, CH2), 3.34 (brs, 2H, D2O-exc-hangeable, NH2), 7.95 (s, 1H, pyrazole-H2), 8.30 (s, 1H, D2O-exchangeable, NH), 8.42 (s, 1H, D2O-exchangeable, NH), 9.86 (s, 1H, CH=N). MS: m/z 483 (M+, 0.1%). Anal. Calcd for C19H17N9OS3: C, 47.19; H, 3.54; N, 26.07; S, 19.89. Found: C, 47.18; H, 3.55; N, 26.08; S, 19.88.
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3. 1. 9. Synthesis of 3-(methylthio)-N-[(10-oxo-4,6,7,8,9,10-hexahydropyrazolo[1,5-a][1] benzothieno[2,3-d]pyrimidin-3-yl) methylene]-4,6-dithioxo-4,5,6,7-tetrahydro-1.ff-pyrazolo[3,4-d] pyrimidine-1-carbothiohydrazide (12)
A mixture of compound 11 (0.483 g, 1 mmol) and carbon disulfide (0.06 mL, 1 mmol) in dry pyridine (10 mL) was heated under reflux for 8 h or until evolution of H2S ceased. After cooling, the reaction mixture was poured onto ice/HCl. The solid obtained was filtered off, washed several times with water, and recrystallized from DMF/et-hanol to give 12 as brown crystals. Yield 62%, mp >300 °C. IR (KBr) v 3421 (3NH), 3032 (CH^), 2926 (CH ), 1653 (C=O), 1636 (C=N), 1184 (3C=S) cm-1. 1H NMR (DMSO-d6) 5 1.79-1.82 (m, 4H, 2CH2), 2.41 (s, 3H, SCH3), 2.669 (t, 2H, CH2), 2.93 (t, 2H, CH2), 7.69 (s, 1H, pyraz3ole-H2), 8.05 (s, 2H2 , D2O-exchangeab2le, 2NH), 8.15 (s, 1H, D2O-exchangeable, NH), 8.74 (s, 1H, CH=N). MS: m/z 558 (M+-1, 8.5%). Anal. Calcd for C20H17N9OS5: C, 42.92; H, 3.06; N, 22.52; S, 28.64. Found: C, 42.91; H, 3.05; N, 22.53; S, 28.65.
3. 1. 10. Synthesis of N-[4-cyano-3-(methylthio)-1-{[2-(10-oxo-4,6,7,8,9,10-hexahydro pyrazolo[1,5-a][1]benzothieno [2,3-d]pyrimidin-3-yl)methylene] hydrazino} carbonothioyl]-1.ff-pyrazol-5-yl] acetamide (13)
A mixture of compound 11 (0.483 g, 1 mmol) and acetic anhydride (10 mL) was heated under reflux for 1 h. After cooling, the reaction mixture was poured onto crushed ice. The solid obtained was filtered off, washed several times with water, and recrystallized from DMF to give 13 as pale brown crystals. Yield 74%, mp 198-200 °C. IR (KBr) v 3447 (3NH), 3061 (CHar0m), 2934 (CHa]iph), 2205 (C=N), 1701 (C=OaCetamldo), 1684 (C=O), 1636 (C=N), 1254 (C=S) cm-1. 1H NMR (DMSO-d6) 5 1.79-1.83 (m, 4H, 2CH2), 2.20 (s, 3H, COCH3), 2.42 (s, 3H, SCH3), 2.81 (t, 2H, CH2), 2.91 (t, 2H, CH2), 7.82 (s, 1H, D2O-exchan-geable, NH), 7.92 (s, 1H, pyrazole-H2), 8.10 (s, 1H, D2O-exchangeable, NH), 8.13 (s, 1H, D2O-exchangeable, NH), 8.42 (s, 1H, CH=N). Anal. Calcd for C21H19N9O2S3: C, 47.99; H, 3.64; N, 23.98; S, 18.30. Found: C, 497.98; H, 3.65; N, 23.97; S, 18.31.
3. 1. 11. Synthesis of 6-methyl-3-(methylthio)-N-[(10-oxo-4,6,7,8,9,10-hexahydropyrazolo [1,5-a][1]benzothieno[2,3-rf]pyrimidin-3-yl)methylene]-4-oxo-4,5-dihydro-1.ff-pyrazolo [3,4-d]pyrimidine-1-carbothiohydrazide (14)
A mixture of compound 11 (0.483 g, 1 mmol) and acetic anhydride (10 mL) in dry pyridine (10 mL) was
heated under reflux for 6 h. After cooling, the reaction mixture was poured onto crushed ice. The solid obtained was filtered off, washed several times with water, and recrystallized from dioxane to give 14 as brown crystals. Yield 65%, mp >300 °C. IR (KBr) v 3421-3200 (3NH), 3048 (CHarom), 2933 (CHaliph), 1685, 1654 (2C=O), 1617 (C=N), 1182 (C=S) cm-1. 1H NMR (DMSO-d() 5 1.23 (s, 3H, CH3), 1.78-1.80 (m, 4H, 2CH2), 2.40 (s, 3H, SCH3), 2.67 (t, 2H, CH2), 2.90 (t, 2H, CH2), 7.50 (s, 1H, D2O-exc-hangeable, NH), 8.01 (s, 1H, pyrazole-H2), 8.42 (s, 1H, CH=N), 9.80 (s, 2H, D2O-exchangeable, 2NH). MS: m/z 525 (M+, 12.4%). Anal. Calcd for C21H19N9O2S3: C, 47.99; H, 3.64; N, 23.98; S, 18.30. Found: C, 47.99; H, 3.64; N, 23.98; S, 18.30.
3. 1. 12. Synthesis of 4-amino-3-(methylthio)-.N-[(10-oxo-4,6,7,8,9,10-hexahydropyrazolo [1,5-a][1]benzothieno[2,3-d]pyrimidin-3-yl)methylene]-1.ff-pyrazolo[3,4-d] pyrimidine-1-carbothiohydrazide (15)
A mixture of compound 11 (0.483 g, 1 mmol) and formamide (10 mL) was heated under reflux for 6 h. After cooling, the reaction mixture was poured onto crushed ice. The solid obtained was filtered off, washed several times with water, and recrystallized from ethanol to give 15 as pale brown crystals. Yield 81%, mp 194-196 °C. IR (KBr) v 3449-3411 (NH2, 2NH), 3058 (CHarom), 2931 (CHaliph), 1685 (C=O), 1631 (C=N), 1148 (C=S) cm-1. 1H NMR (DMSO-d6) 5 1.77-1.82 (m, 4H, 2CH2), 2.39 (s, 3H, SCH3), 2.65 (t, 2H, CH2), 2.89 (t, 2H, CH2), 3.38 (brs, 2H, D2O3exchangeable, NH2), 7.41 (s, 1H, Dp-exchangeable, NH), 7.63 (s, 1H, pyrimidine-H6), 7.95 (s, 1H, pyrazole-H2), 7.99 (s, 1H, D2O-exchangeable, NH), 8.13 (s, 1H, CH=N). MS: m/z 510 (M+, 2%). Anal. Calcd for C20H18N10OS3: C, 47.04; H, 3.55; N, 27.43; S, 18.84. Found: C, 47.05; H, 3.54; N, 27.42; S, 18.85.
3. 1. 13. Synthesis of 4-imino-3-(methylthio)-5-phenyl-N-[(10-oxo-4,6,7,8,9,10-hexahy-dropyrazolo[1,5-a][1]benzothieno[2,3-d] pyrimidin-3-yl)methylene]-6-thioxo-4,5,6,7-tetrahydro-1.ff-pyrazolo[3,4-d] pyrimidine-1-carbothiohydrazide (16)
A mixture of compound 11 (0.483 g, 1 mmol) and phenyl isothiocynate (0.135 mL, 1 mmol) in DMF (10 mL) containing a few drops of piperidine was heated under reflux for 4 h. After cooling, the reaction mixture was poured onto crushed ice. The solid obtained was filtered off, washed several times with water, and recrystallized from DMF/H2O to give 16 as brown crystals. Yield 66%, mp 235-237 °2C. IR (KBr) v 3446 (4NH), 3043 (CHarom.), 2927 (CHaliph), 1647 (C=O), 1558 (C=N), 1181 (C=S) cm-1. 1H NMR (DMSO-d() 5 1.79-1.82 (m, 4H, 2CH2), 2.41 (s, 3H, SCH3), 2.73 ((t, 2H, CH2), 2.92 (t, 2H, CH22),
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6.96-7.60 (m, 5H, ArH), 7.95 (s, 1H, pyrazole-H2), 8.06 (s, 1H, D2O-exchangeable, NH), 8.20 (s, 1H, Dp-exchangeable, NH), 8.33 (s, 1H, D2O-exchangeable, NH), 8.38 (s, 1H, D2O-exchangeable, NH), 9.90 (s, 1H, CH=N). MS: m/z 618 (M+, 7.5%). Anal. Calcd for C26H22N10OS4: C, 50.47; H, 3.58; N, 22.64; S, 20.73. Found: C, 50.46; H, 3.59; N, 22.63; S, 20.74.
4.	Conclusions
In conclusion N'-[(10-oxo-4,6,7,8,9,10-hexahydrop-yrazolo[1,5-a][1]benzothieno[2,3-d]pyrimidin-3-yl) methylene] thiocarbohydrazide (3) has proved to be a versatile precursor for the synthesis of some new Schiff bases containing the pyrazolobenzothieno-pyrimidine ring. The synthesized compounds were screened for their antimicrobial activity.
5.	References
1.	H. Schiff, Justus Liebigs Ann. Chem. 1864, 131, 118-119. http://dx.doi.org/10.1542/peds.101A583
2.	G. M. Loudon (Ed.4th): Organic Chemistry, Addison-Wesley, California, 2002, p. 874. http://dx.doi.org/10.1056/NEJMra021561
3.	M. A. Metwally, M. E. Khalifa, M. Koketsu, Am. J. Chem. 2012, 2, 38-51. http://dx.doi.org/10.1210/jcem.85.3.6441
4.	M. S. Chande, M. A. Pankhi, S. B. Ambhaikar, Indian J. Chem. 2000, 39B, 603-609.
http://dx.doi.org/10.1172/JCI115897
5.	O. V. Mikhailov, M. A. Kazymova, T. A. Shumilova, G. A. Chmutova, S. E. Solovieva, Transition Met. Chem. 2005, 30, 299-304.
6.	M. R. Islam, K. Khayer, G. Shaha, M. S. K. Chowd-hury, Indian. J Chem. 1992, 31B, 547-551. http://dx.doi.org/10.1530/eje.L01944
7.	A. A Aly, A. B. Brown, T. I. El-Emary, A. M. M. Ewas, M. Ramadan, Arkivoc 2009, i, 150-197.
8.	X. H. Sun, Y. F. Liu, Huaxue Tongbao 1999, 62, 46-48 (in Chinese). http://dx.doi.org/10.1210/jc.2008-1174
9.	M. P. Sathisha, U. N. Shetti, V. K. Revankar, K. S. R. Pai, Eur. J. Med. Chem. 2008, 43, 2338-2346. http://dx.doi.org/10.1006/abbi.1997.0302
10.	M. Kritsanida, A. Mouroutsou, P. Marakos, N. Pouli, S. P. Garoufalias, C. Pannecouque, M. Witvrouw, E. De Clercq, Il Farmaco 2002, 57, 253-257.
11.	B. S. Holla, P. M. Akberali, M. K. Shivananda, Il Farmaco 2001, 56, 919-927.
12.	S. Sutradhar, A. M. Maliha, A. Humera, Res. J. Pharm., Biol. Chem. Sci. 2013, 4, 878-887. http://dx.doi.org/10.1016/jxlpt.2005.10.002
13.	K. M. EL-mahdy, A. M. El-Kazak, M. Abdel-Megid, M. Seada, O. Farouk, J. Adv. Chem. 2013, 5, 581-591. http://dx.doi.org/10.1210/jcem-57-2-320
14.	N. Petri, Z. Naturforsch. 1961, 16B, 769. http://dx.doi.org/10.1530/eje.0T410132
15.	D. C. Gross, J. E. DeVay, Physiol. Plant Pathol. 1977, 11, 13-28. http://dx.doi.org/10.1530/eje.0T490137
16.	A. W. Bauer, W. M. Kirby, J. C. Sherris, M. Turck, Am. J. Clin. Pathol. 1966, 45, 493-496. http://dx.doi.org/10.1007/s00228-008-0498-2
Povzetek
Pri reakciji med pirazolobenzotienopirimidin-3-karbaldehidom 1 in tiokarbohidrazidom nastane Schiffova baza 3, ki lahko dalje reagira z različnimi elektrofilnimi reagenti pri čemer nastanejo 1,2,4-triazoli 4-6 ali 1,2,4-triazini 7-9. Obdelava spojine 3 z 2-ciano-3,3-bis(metiltio)akrilonitrilom daje ustrezen 5-amino-4-ciano-3-metiltiopirazolski derivat 11. Reakcija pirazola 11 z ogljikovim disulfidom daje ditioksopirazolopirimidin 12. Pri aciliranju spojine 11 z acetanhi-dridom nastane acetamid 13. Po drugi strani pa ciklokondenzacija pirazola 11 z acetanhidridom v piridinu daje pirazo-lopirimidinske derivate 14. Raziskali smo tudi reakcijo spojine 11 s formamidom in fenilizotiocianatom ter ugotovili, da nastaneta pirazolopirimidina 15 in 16. Določili smo tudi antimikrobno aktivnost novih spojin.
El-Mahdy et al.: Synthesis, Characterization and Antimicrobial Activities