Studies on pyrazine derivatives, XLV: Synthesis, reactions, and tuberculostatic activity of N-methyl-N′-(pyrazine-2-carbonyl)- hydrazinecarbodithioic acid methyl ester

Article abstract of DOI:10.1080/10426500500272319

Methyldithiocarbonyl derivative 2 of pyrazine-2-carboxylic acid N′-methylhydrazide 1 was synthesized by methylation of CS₂ adduct. Benzylamine caused the decomposition of compound 2 to pyrazine-2-carboxylic acid benzylamide 5 and 1,3-dibenzylthiourea, 6. N-methyl-N'′-(pyrazine-2-carbonyl)-hydrazinecarbodithioic acid methyl ester 2 were evidenced to cyclize to 3-methyl-5-pyrazin-2-yl-3H-[1,3,4] oxadiazole-2-thione 8 in the presence of triethylamine. In the reactions with secondary amines such as morpholine, pyrrolidine and phenylpiperazine pyrazinoyl derivatives (9-11) of thiosemicarbazide were obtained. Hydrazine, methylhydrazine, aminoalcohols, and N-alkylamino-substituted cyclic amines reacted with cyclization to 4-substituted 1,2,4-triazole-3-thiones 12, 13, and 18-22. Synthesized compounds exhibited low tuberculostatic activity in vitro (MIC 50-100 μg/mL). Copyright Taylor & Francis Group, LLC.

Full text of DOI:10.1080/10426500500272319

Phosphorus, Sulfur, and Silicon, 181:965–975, 2006
Copyright © Taylor & Francis Group, LLC
ISSN: 1042-6507 print / 1563-5325 online
DOI: 10.1080/10426500500272319
Studies on Pyrazine Derivatives, XLV: Synthesis,
Reactions, and Tuberculostatic Activity of N-Methyl-Nꢀ-
(pyrazine-2-carbonyl)-hydrazinecarbodithioic Acid
Methyl Ester
Katarzyna Gobis
Henryk Foks
Department of Organic Chemistry, Medical University of Gdan´sk,
Gdan´sk, Poland
Zofia Zwolska
Ewa Augustynowicz-Kopec
´
Department of Microbiology, Institute of Tuberculosis and Pulmonary
Diseases, Warsaw, Poland
Methyldithiocarbonyl derivative
2
of pyrazine-2-carboxylic acid Nꢀ-methyl-
hydrazide 1 was synthesized by methylation of CS2 adduct. Benzylamine caused
the decomposition of compound 2 to pyrazine-2-carboxylic acid benzylamide 5 and
1,3-dibenzylthiourea 6. N-methyl-Nꢀ-(pyrazine-2-carbonyl)-hydrazinecarbodithioic
acid methyl ester 2 were evidenced to cyclize to 3-methyl-5-pyrazin-2-yl-3H-
[1,3,4]oxadiazole-2-thione 8 in the presence of triethylamine. In the reactions with
secondary amines such as morpholine, pyrrolidine and phenylpiperazine pyrazi-
noyl derivatives (9–11) of thiosemicarbazide were obtained. Hydrazine, methylhy-
drazine, aminoalcohols, and N-alkylamino-substituted cyclic amines reacted with
cyclization to 4-substituted 1,2,4-triazole-3-thiones 12, 13, and 18–22. Synthesized
compounds exhibited low tuberculostatic activity in vitro (MIC 50–100 µg/mL).
Keywords
2-methyl-[1,2,4]triazole-3-thione;
3-methyl-[1,3,4]oxadiazole-2-thione;
methyldithiocarbazate esters; pyrazine-2-carboxylic acid Nꢀ-methyl-hydrazide;
thiosemicarbazides; tuberculostatic activity
INTRODUCTION
Our previous research has shown that mono- and dithioesters of pyrazi-
noylcarbodithioic acid undergo reactions with amines, and the struc-
ture of products depends on the amine type and reaction conditions.
Received April 12, 2005; accepted April 26, 2005.
Address correspondence to Katarzyna Gobis, Medical University of Gdan´sk, Depart-
ment of Chemistry, Al. Fen. Hallera 107, Gdan´sk 80-416, Poland. E-mail: katarzyna.
gobis@wp.pl
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K. Gobis et al.
Derivatives of thiosemicarbazide as well as heterocyclic compounds
such as the 1,3,4-oxadiazole,1,2,4-triazole, and triazolepyrimidine con-
densed structure1−3 can be the products of reactions.
CHEMISTRY
In this work, we tried to investigate the possibilities of the us-
age of the N-Methyl-Nꢀ-(pyrazine-2-carbonyl)-hydrazinecarbodithioic
acid methyl ester 2 obtained from pyrazine-2-carboxylic acid Nꢀ-
methyl-hydrazide 1 in organic synthesis. Hydrazide was synthesized
from pyrazinamide in a result of typical reaction described in the
literature.4,5 Pyrazine-2-carboxylic acid obtained by hydrolysis was
estrificated with methanol in the presence of thionyl chloride. The
originated ester changed into an expected pyrazine-2-carboxylic acid
Nꢀ-methyl-hydrazide 1 under influence of hydrazine. N-methyl-Nꢀ-
(pyrazine-2-carbonyl)-hydrazinecarbodithioic acid methyl ester 2 was
synthesized as a product of CS2 and methyl iodide treatment on com-
pound 1 in a presence of triethylamine (Scheme 1). The reaction ran
fast and with a good yield.
Next, we examined an influence of various amines on com-
pound 2. In our research, we used benzylamine, cyclohexy-
lamine, triethylamine, morpholine, pyrrolidine, 1-phenylpiperazine,
hydrazine and methylhydrazine, ethanolamine, 1,3-diaminopropanol,
SCHEME 1

Studies on Pyrazine Derivatives, XLV:
967
1-(2-aminoethyl)pyrrolidine, 2-piperidinoethylamine, and N-(3-amino-
propyl)imidazole. The products of the reactions were different and de-
pended on the amine structure.
In a reaction with benzylamine we expected two products: thiosemi-
carbazide derivative 3 and a product of its cyclization 4. That possi-
bility was excluded after the synthesis of compound 3 from hydrazide
1 and benzyl isothiocyanate and compound 4 in the cyclization reac-
tion of 3. IR spectra exhibited a different structure of obtained prod-
ucts. On the basis of IR and 1H NMR spectra, we maintained that
pyrazine-2-carboxylic acid benzylamide 5 and 1,3-dibenzylthiourea
6 were the products of a reaction between N-methyl-Nꢀ-(pyrazine-
2-carbonyl)-hydrazinecarbodithioic acid methyl ester 2 and benzy-
lamine. The structures of the obtained compounds were confirmed
by control syntheses: 5 in reaction of pyrazine-2-carboxylic acid
methyl ester with benzylamine and 6 from benzyl isothiocyanate and
benzylamine.
In the reaction of compound 2 with cyclohexylamine, only 1,3-
dicyclohexylthiourea 7 was isolated as crystalline product. That product
can create as a result of methyl mercaptan elimination from compound 2
under the influence of cyclohexylamine while the thiocarbonyl carbon is
attacked by another molecule of amine. Thiourea forms as a result of
the C N bond breaking. Dithiocarboxylic acid monoester without the
methyl group in the hydrazine moiety demonstrated the same course
of reaction with cyclohexylamine as we have described in a previous
article.5 Dicyclohexylthiourea and pyrazin-2-carboxylic acid hydrazide
are the products of that reaction.
The heating of N-methyl-Nꢀ-(pyrazine-2-carbonyl)-hydrazinecarbo-
dithioic acid methyl ester 2 in ethanol in the presence of triethy-
lamine led to 3-methyl-5-pyrazin-2-yl-3H-[1,3,4]oxadiazole-2-thione 8
(Scheme 2).
In the reactions with pyrrolidine, morpholine and 1-phenylpi-
perazine compound 2 gave the products of thiosemicarbazide structures
9–11. IR spectra with a strong band at 1680 cm−1 characteristic for the
carbonyl group confirmed that kind of structure.
The reaction of compound 2 with hydrazine and methylhydrazine
resulted 4-amino-[1,2,4]triazole-3-thione 12 and 4-methylamino-
[1,2,4]triazole-3-thione 13. For derivative 12 possessing a free NH2
group in 4-position, condensation reactions with p-fluoro- and p-
chlorobenzaldehyde were performed giving halobenzylidene com-
pounds 14 and 15. Both processes proceeded with very good yields
when the substrates were heated in ethanol. Derivative 12 also reacted
with allyl isothiocyanate and gave 1,3-disubstituted thiourea 16. 4-
methylamino-[1,2,4]triazole-3-thione 13 did not undergo that reaction,

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K. Gobis et al.
SCHEME 2
and the presence of the NH group in 4-position was evidenced by the
reaction with acetic anhydride, which formed acetamide 17.
In the reactions with ethanolamine, 1,3-diaminopropanol,
1-(2-aminoethyl)pyrrolidine, 2-piperidinoethylamine, and 1-(3-
aminopropyl)imidazole compound 2 gave appropriate 4-aminoalkyl-
[1,2,4]triazole-3-thiones 18–22. All products formed with good yields,
especially when the reactants were liquid and the reactions could be
performed with their large excess towards 2 without a solvent.
MICROBIOLOGY
The newly synthesized derivatives were examined for their tubercu-
lostatic activity towards the Mycobacterium tuberculosis H37Rv strain
and two “wild” strains isolated from tuberculotic patients: one (Spec.
210) resistant to p-aminosalicic acid (PAS), isonicotinic acid hydrazide
(INH), etambutol (ETB), and rifampicine (RFP); another (Spec. 192)
fully sensitive to the administrated drugs. In vitro investigations
were performed by a classical test-tube method of successive dilution
with Youman’s liquid medium containing 10% of bovine serum.5 The

Studies on Pyrazine Derivatives, XLV:
969
determined minimum concentrations (MIC) inhibiting the growth of
tuberculous strains for all the tested compounds were within the limits
50–100 µg/mL, which indicates low antituberculosis activity. The de-
fences in compound activity on particular stains have not been observed.
EXPERIMENTAL
All materials and solvents were of analytical reagent grade. Thin-layer
chromatography was performed on Merck Kieselgel 60F254 plates and
visualized with UV. Silica gel 60 (70–230 mesh, Fluka) was used for
column chromatography. The results of elemental analyses (% C, H,
N) for all the compounds obtained were in good agreement with the
1
data calculated. H NMR spectra in CDCl3 were recorded on Varian
Gemini (200 MHz) and Varian Unity Plus (500 MHz) instruments.
IR Spectra (KBr) were determined as KBr pellets of the solids on a
satellite FT-IR spectrophotometer. Mass spectrum for compound 18
was taken on Finingan MAT 95 (70 eV). Melting points were deter-
mined on BOETIUS apparatus and were uncorrected. Reaction yields,
physical constants, and spectral data of the compounds are given in
Table I.
Pyrazine-2-carboxylic Acid Nꢀ-Methyl-hydrazide (1)
Pyrazine-2-carcoxylic acid methyl ester (15.3 g, 0.1 mol) was dis-
solved in 30 mL of ethanol, and 7.2 g (0.15 mol) of methylhydrazine
were added. The reaction mixture was heated under reflux for 1.5 h.
Then ethanol was evaporated in vacuum, and oily residue was treated
with 20 mL of diethyl ether and 5 mL of ethanol. The precipitate
(15.2 g, 68%) was filtered after cooling and recrystallized from benzene
(95–96◦C).
N-Methyl-Nꢀ-(pyrazine-2-carbonyl)-hydrazinecarbodithioic
Acid Methyl Ester (2)
A quantity of 3 g (20 mmol) of 1 was dissolved in 20 mL of methanol.
Next, 3 mL (21 mmol) of triethylamine and 1.5 mL (25 mmol) of carbon
disulfide were added. After 5 min of stirring, 1.5 mL (25 mmol) of methyl
iodide were added. Reaction mixture was stirred for another 30 min
and diluted with 100 mL of water. The crystals were filtered (2.6 g) and
recrystallized.

970

971

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K. Gobis et al.
Pyrazine-2-carboxylic Acid Nꢀ-Methyl-
Nꢀ-(benzylamine-4-carbothioyl)-hydrazide (3)
Compound 1 (1.5 g, 10 mmol) and benzyl isothiocyanate (1.3 mL, 10
mmol) were refluxed in 10 mL of ethanol for 1 h. The resulting solid
was filtered after cooling and recrystallized. Yield: 2.8 g.
4-Benzyl-2-methyl-5-pyrazin-2-yl-2,4-dihydro-[1,2,4]triazole-
3-Thione (4)
A quantity of 0.3 g (1 mmol) of 3 was suspended in a solution of potas-
sium carbonate (1 g) in water (10 mL). The mixture was heated under
reflux for 30 min and then cooled, yielding crystals (0.24 g), which were
filtered and recrystallized.
Pyrazine-2-carboxylic Acid Benzylamide (5)
A.
Pyrazine-2-carboxylic acid methyl ester (1.38 g, 10 mmol) was re-
fluxed with 2 mL (18 mmol) of benzylamine for 2 h. Then the mixture
was cooled and treated with 5 mL of diethyl ether and 20 mL of cyclo-
hexane. The precipitate (1.5 g) was filtered and recrystallized.
B.
The synthesis was carried out as described in Method A from 0.5 g
1
(3 mmol) of 2 and 1.5 mL (14 mmol) of benzylamine. H NMR and IR
spectra and m.p. determined for the product obtained by both methods
were the same as measured for the compound synthesized by heating
2 with benzylamine.
1,3-Dicyclohexyl-thiourea (7)
Compound 2 (0.5 g, 2 mmol) was refluxed with 2 mL (17 mmol) of cy-
clohexylamine for 4.5 h. Then the mixture was cooled and treated with
diethyl ether and then with petroleum ether until the product precip-
itated completely. Yield: 0.42 g. IR spectrum determined for the prod-
uct was identical as for dicyclohexylthiourea obtained from cyclohexy-
lamine and benzyl isothiocyanate.
3-Methyl-5-pyrazin-2-yl-3H-[1,3,4]oxadiazole-2-thione (8)
A quantity of 0.56 g (2.5 mmol) of 2, 10 mL of ethanol, and 0.5 mL
(3.6 mmol) of triethylamine was refluxed for 30 min, and then the

Studies on Pyrazine Derivatives, XLV:
973
reaction mixture was concentrated. Next, 2 mL of ethanol was added,
and 0.45 g of precipitate was filtered.
Pyrazine-2-carboxylic Acid Nꢀ-Methyl-Nꢀ-(pyrrolidine-4-
carbothioyl)-hydrazide (9)
Compound 2 (1 g, 4 mmol) was suspended in 3.5 mL of pyrrolidine. The
mixture was refluxed for 4 h and then poured on 50 g of ice, the the
water solution was extracted with chloroform and dried with magne-
sium sulfate. Chloroform was evaporated and the residue was extracted
with hot cyclohexane. The solid (0.31 g) was filtered after cooling and
recrystallized.
Pyrazine-2-carboxylic Acid Nꢀ-Methyl-Nꢀ-(morpholine-4-
carbothioyl)-hydrazide (10)
The synthesis was performed as described for 9 from compound 2 (1 g,
4 mmol) and 3 mL of morpholine. Yield: 0.37 g.
Pyrazine-2-carboxylic Acid Nꢀ-Methyl-Nꢀ-[(4-phenyl)-
piperazine-4-carbothioyl]-hydrazide (11)
A quantity of 1 g (4 mmol) of 2 and 3 mL of 1-phenyl-piperazine was
heated at 150◦C with stirring for 3 h. Then, the mixture was cooled,
and the product was isolated by column chromatography with ben-
zene/acetone system 50:1 as a liquid phase. Yield: 0.70 g.
4-Amino-2-methyl-5-pyrazin-2-yl-2,4-dihydro-
[1,2,4]triazole-3-thione (12)
Compound 2 (1.13 g, 5 mmol) was added to the solution consisting of
3 mL of dioxane, 2 mL of ethanol, and 0.5 mL of 100% hydrazine hy-
drate. The mixture was heated under reflux for 2 h. Then, the solution
was cooled, and the precipitate in the amount of 0.6 g was filtered and
recrystallized.
2-Methyl-4-methylamino-5-pyrazin-2-yl-2,4-dihydro-
[1,2,4]triazole-3-thione (13)
A quantity of 2 (4.52 g, 20 mmol), 15 mL of dioxane, and 1.6 mL
(30 mmol) of methylhydrazine were refluxed for 2 h, and then the sol-
vent was evaporated. Next, 10 mL of water was added to the residue.
The solid (2.82 g) was filtered and recrystallized.

974
K. Gobis et al.
4-[(4-Fluoro-benzylidene)-amino]-2-methyl-5-pyrazin-2-yl-2,4-
dihydro-[1,2,4]triazole-3-thione (14)
Equimolecular amounts (2.5 mmol) of compound
8
and p-
fluorobenzaldehyde were heated under reflux in 10 ml of ethanol
for 30 min. The yellow crystals (0.35 g) were filtered after cooling.
4-[(4-Chloro-benzylidene)-amino]-2-methyl-5-pyrazin-2-yl-2,4-
dihydro-[1,2,4]triazole-3-thione (15)
The synthesis was performed as described for compound 14 from
2.5 mmol of 8 and p-chlorobenzaldehyde, yielding 0.6 g of the product.
1-Allyl-3-(1-methyl-3-pyrazin-2-yl-thixo-1,5-dihydro-
[1,2,4]triazol-4-yl)-thiourea (16)
Compound 12 (0.5 g, 2.6 mmol) was dissolved in 5 mL of ethanol, and
0.53 mL (5.2 mmol) of allyl isothiocyanate was added. The mixture was
heated under reflux for 4 h, and then the solvent was evaporated. The
oily residue was extracted with hot cyclohexane, yielding the solid after
cooling. Yield: 0.47 g.
N-Methyl-N-(1-methyl-3-pyrazin-2-yl-5-thioxo-1,5-dihydro-
[1,2,4]triazol-4-yl)-acetamide (17)
Compound 13 (0.54 g, 2.6 mmol) was dissolved in 2.5 mL of acetic an-
hydride. The solution was refluxed for 2 h and then poured into 10 mL
of water. After 15 min of stirring, the solution was neutralized with
solid potassium carbonate. The precipitate (0.41 g) was filtered and
recrystallized.
4-(2-Hydroxy-ethyl)-2-methyl-5-pyrazin-2-yl-2,4-
dihydro-[1,2,4]triazole-3-thione (18)
A quantity of 3.4 g (15 mmol) of 2 in 2 mL of ethanolamine was refluxed
for 3 h. Then, 50 mL of water was added, and the solution was extracted
with chloroform and dried with magnesium sulfate. The solvent was
evaporated and 2.08 g of the residual was recrystallized. MS, m/e (%):
72.3 (20.27); 79.1 (33.82); 106 (40.51), 194 (100), M+ 237 (7.28).
4-(3-Amino-2-hydroxy-propyl)-2-methyl-5-pyrazin-2-yl-2,4-
dihydro-[1,2,4]triazole-3-thione (19)
Compound 2 (1.13 g, 5 mmol) and 1,3-diamino-2-propanol (0.6 g,
6.5 mmol) were heated under reflux in combination of 10 mL of dioxane

Studies on Pyrazine Derivatives, XLV:
975
and 3 mL of ethanol for 5 h. The mixture was cooled, and the solu-
tion was separated from the lower oily layer and treated with 80 mL of
petroleum ether. The precipitate (1.2 g) was filtered and recrystallized.
2-Methyl-5-pyrazin-2-yl-4-(2-pyrrolidin-1-yl-ethyl)-2,4-dihydro-
[1,2,4]triazole-3-thione (20)
Quantities of 0.75 g (3 mmol) of 2 and 1.5 mL of 1-(2-aminoethyl)pyrro-
lidine were heated at 120◦C with stirring for 1.5 h. Then, the reaction
mixture was cooled and diluted with 25 mL of water. The precipitate
(0.65 g) was filtered and recrystallized.
2-Methyl-5-pyrazin-2-yl-4-(2-pyridin-1-yl-ethyl)-2,4-dihydro-
[1,2,4]triazole-3-thione (21)
The synthesis was carried out as described for compound 20 from 1 g
(4 mmol) of 2 and 2 mL of 2-piperidinoethylamine. Yield: 0.98 g.
4-(2-Imidazol-1-yl-ethyl)-2-methyl-5-pyrazin-2-yl-2,4-dihydro-
[1,2,4]triazole-3-thione (22)
A
quantity of 0.75 g (3 mmol) of 2 in 1.4 mL of N-(3-
aminopropyl)imidazole was heated at 120◦C with stirring for 1.5 h.
Then, the reaction mixture was cooled and diluted with 25 mL of water.
The solution was extracted with chloroform and dried with magnesium
sulfate. The solvent was evaporated, and the residue was treated with
diethyl ether, yielding 0.66 g of the solid.
REFERENCES
[1] H. Foks, J. Mieczkowska, and M. Sitarz, Phosphorus, Sulfur, and Silicon, 158, 107
(2000).
[2] H. Foks, J. Trapkowska, M. Janowiec, Z. Zwolska, and E. Augustynowicz-Kopec´,
Chim. Geterotsikl. Soedin., 1368, 9 (2004).
[3] C. Orlewska, H. Foks, M. Janowiec, and Z. Zwolska-Kwiek, Pharmazie, 50, 565 (1995).
[4] H. Foks and J. Sawlewicz, Acta Polon. Pharm., 21, 429 (1964).
[5] D. Pancechowska-Ksepko, H. Foks, M. Janowiec, and Z. Zwolska-Kwiek, Acta Polon.
Pharm., 45, 193 (1988).