Synthesis and antibacterial activity of substituted thiosemicarbazides and of 1,3,4-thiadiazole or 1,2,4-triazole derivatives

Article abstract of DOI:10.1080/10426500500269851

The synthesized series of new thiosemicarbazide derivatives (1, 6-10) in reactions with carbon disulphide produced, according to the reaction conditions, the dithioacids (4, 30) or the 5-substituted l,3,4-thiadiazolo-2-thiol derivatives (2, 27). The dithi

Full text of DOI:10.1080/10426500500269851

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Phosphorus, Sulfur, and Silicon
and the Related Elements
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Synthesis and Antibacterial
Activity of Substituted
Thiosemicarbazides and of
1,3,4-Thiadiazole or 1,2,4-
Triazole Derivatives
Katarzyna Spalińska a , Henryk Foks a , Anna Kedzia
b , Marta Wierzbowska b , Ewa Kwapisz b , Alina
Gebska b & Marta Zilkółwska-Klinkosz b
a Department of Organic Chemistry, Medical
University of Gdańsk, Gdańsk, Poland
b Department of Oral Microbiology, Medical
University of Gdańsk, Gdańsk, Poland
Published online: 01 Feb 2007.
To cite this article: Katarzyna Spaliska , Henryk Foks , Anna Kedzia , Marta
Wierzbowska , Ewa Kwapisz , Alina Gebska & Marta Zilkwska-Klinkosz (2006) Synthesis
and Antibacterial Activity of Substituted Thiosemicarbazides and of 1,3,4-Thiadiazole
or 1,2,4-Triazole Derivatives, Phosphorus, Sulfur, and Silicon and the Related
Elements, 181:3, 609-625, DOI: 10.1080/10426500500269851
To link to this article: http://dx.doi.org/10.1080/10426500500269851
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Phosphorus, Sulfur, and Silicon, 181:609–625, 2006
Copyright © Taylor & Francis Group, LLC
ISSN: 1042-6507 print / 1563-5325 online
DOI: 10.1080/10426500500269851
Synthesis and Antibacterial Activity of Substituted
Thiosemicarbazides and of 1,3,4-Thiadiazole or
1,2,4-Triazole Derivatives
´
Katarzyna Spalinska
Henryk Foks
Department of Organic Chemistry, Medical University of Gdan´sk,
Gdan´sk, Poland
Anna Ke¸dzia
Marta Wierzbowska
Ewa Kwapisz
Alina Ge¸bska
´
Marta Ziolkowska-Klinkosz
ꢀ
Department of Oral Microbiology, Medical University of Gdan´sk,
Gdan´sk, Poland
The synthesized series of new thiosemicarbazide derivatives (1, 6–10) in reac-
tions with carbon disulphide produced, according to the reaction conditions, the
dithioacids (4, 30) or the 5-substituted 1,3,4-thiadiazolo-2-thiol derivatives (2,
27). The dithioacids were cyclized, in the reaction with hydrazine, into the 4-
ami-no-1,2,4-triazolo-2-thiol derivatives (5, 31). One of these compounds (31) was
transformed into the 1,2,4-triazolo-1,3,4-thiadiazine derivative (33). The compo-
unds 6–9 were also exposed to the condensation with aldehydes. 4-phenylpipera-
zinocarbothiohydrazide (6) was exposed to the action of isothiocyanates, which gave
the compounds 16–20, and these cyclized to the 1,3,4-thiadiazoloamino derivatives
(21–23).
The susceptibility of aerobic and anaerobic bacteria to some of the new derivatives
were tested. The anaerobes were the most susceptible at concentrations in ranges
less than 6.2 to 100 µg/mL to derivative: 9 (64% were susceptible), 1, 13 (for 60%),
and 7 (for 56%).
Keywords 1,3,4-thiadiazole; 1,2,4-triazole; 1,2,4-triazolo-1,3,4-thiadiazine; thiose-
micarbazides; antibacterial activity
Received April 26, 2005; accepted May 12, 2005.
Address correspondence to Henryk Foks Department of Organic Chemistry, Med-
ical University of Gdan´sk, Al. Gen. J. Hallera 107, Gdan´sk 80416, Poland. E-mail:
hfoks@amg.gda.pl
609

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K. Spalin´ska et al.
INTRODUCTION
Many thiosemicarbazide derivatives described in the chemical
literature produced evidence of the antibacterial, antimalarial,1
antituberculitic,3 as well as anticancer2 activity. The compounds ob-
tained from these heterocyclic systems appeared to be of varied
biological activity as well. The 1,3,4-thiadiazole system-containing
derivatives showed antitubercu-litic,4,5 antibacterial, antimycotic,6,7
and analgesic8 actions. The 1,2,4-triazolo system-con-taining com-
pounds appeared to be antimycotic,9 Alzheimer’s-disease retarding,10
and icterus-Type C-virus-inhibiting11 agents. Their influence over the
immunology of HIV-1 virus infections12 was proved as well. How-
ever, the 1,2,4-triazolo-1,3,4-thiadiazine deri-vatives were recognized
as antiparasitic13 substances. Thus, further syntheses and investiga-
tion of this group of compounds seems to be well founded. Syntheses of
the new thiosemicarbazide derivatives, as well as of amino-substituted
azol derivatives, are reported in this article.
RESULTS AND DISCUSSION
In continuation of our search for potentially antibacterial substances,
the new thiosemi-carbazides and the products of their cyclization re-
actions with carbon disulphide to 1,3,4-thiadiazole, 1,2,4-triazole, and
1,2,4-triazolo-1,3,4-thiadiazine derivatives were obtained.
The substrate for further syntheses was 4-methyl-4-phenyl-3-
thiosemicarbazide 1, obtained by carbon disulphide addition to N-
methylaniline, followed by methylphenylthio-carbaminoylsulphonyl-
acetic acid and thioacetic acid elimination by a hydrazine hydrate
action.14
The products of the carbon disulphide addition to compound 1 de-
pended upon the reaction conditions. At the elevated temperature, 5-
N-methyl-N-phenyl-1,3,4-thiadiazolo-2-thiol 2 was obtained, which on
a methyl iodide action gave its S-methyl derivative 3. At r.t., however,
dithioacid 4 was obtained. The reaction of compound 4 with hydrazine
resulted in a 4-amino-5-N-methyl-N-phenyl-1,2,4-triazolo-2-thiol 5 for-
mation. From compound 1, the dithiocarbazoic acid methyl ester 15 was
obtained as well, but an attempt at obtaining from this compound the
1,3,4-thiadiazole substituted with amine groups and then the morpho-
line derivative of this thiadiazole failed, only compound 2 was obtained
instead.
The action of corresponding amines on compound 1 produced 4-N-
substituted thiosemicarbazide derivatives (6–10). Three of them were
condensed with 5-nitrofuryl aldehyde, which gave the condensation
products (12–14).

Synthesis and Antibacterial Activity
611
The thiosemicarbazide derivative 6 was also obtained by a hydrazine
hydrate action upon 4-phenylpiperazin-1-carbothiomethyl ester 11,
which was produced in the reaction of 1-phenylpiperazine with carbon
disulphide and methyl iodide in the presence of triethylamine. This
method, however, was less effective (Scheme 1).
At the next stage of work, the derivatives of 4-phenylpiperazin-1-
carbothiohydrazide 6 were obtained. As in the 4-phenyl-4-methyl-3-
thiosemicarbazide case, the products of the carbon disulphide addition
were dependent upon the conditions. The corresponding dithio-acid 30,
or 1,3,4-thiadiazolo-2-thiol 27, was produced. From compound 27, the
S-methyl derivative 28 was obtained, and this one, when heated with
morpholine, gave derivative 29. The same compound was obtained as
well, when the dithioacid 30 was refluxed in an excess of morpholine.
The potassium salt of compound 30, necessary for further syntheses,
was obtained by a carbon disulphide addition to 6 in the presence of
potassium hydroxide. From compound 30, potassium salt the deriva-
tive 27 was obtained by microwave irradiation. The same potassium
salt was transformed, under the hydrazine hydrate influence, into the
4-N-amino derivative of 1,2,4-triazolo-2-thiol 31. This last compound
was produced by two methods: either by several hours of heating of
30b salt in ethanol or by several minutes of action of microwave in
radiation. Both methods gave good yields. From compound 31 the con-
densate 32 with p-chlorobenzoic aldehyde was obtained. With regard
to the communication,13 1,2,4-triazolo-1,3,4-thiazine derivatives have
been recognized as antiparasitic agents; the phenylpiperazine deriva-
tive of this condensed system (33) was synthesised by phenacyl bromide
action upon the compound 31.
4-phenylpiperazin-1-carbothiohydrazine 6 was also exposed to the
action of several isothiocyanates, which gave the corresponding thio-
carbamino derivatives (16–20). Some of them were cyclized in boiling
ethanol to the corresponding 2,5-diamino-1,3,4-thiadiazole derivatives
(21–23).
In the condensation of 6 with salicylaldehyde, p-methoxybenzalde-
hyde, and 3,4,5-tri-methoxybenzaldehyde, the products 24–26 were ob-
tained. The choice of aldehydes was dictated by the good results ob-
tained earlier of the antibacterial investigations (Scheme 2). The phys-
ical data are summarized in Table I.
MICROBILOGICAL ACTIVITY
The investigations included 25 strains of anaerobic bacteria (see
Table II) and 25 strains of aerobic bacteria (Staphylococcus aureus,

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614

615

616

617

618

619

620
K. Spalin´ska et al.
Corynebacterium spp., Klebsiella pneumoniae, Acinetobacter baumanii,
Escherichia coli, Pseudomonas aeruginosa, and Pseudomonas stutzeri).
The aerobic bacteria and anaerobic bacteria were isolated from patients
with infections of the oral cavity, respiratory tract, and abdominal cav-
ity.
The susceptibility (MIC) anaerobes were determined by means of
a plate dilution technique in Brucella agar supplemented with 5% of
sheep blood,15,16 whereas aerobes were determined in Mueller-Hinton
agar. The incubation was performed at 37◦C for 48 h in anaero-
bic or aerobic conditions for 24 h. MIC was regarded as the lowest
thiosemicarbazide derivative concentration inhibiting the growth of
bacteria.
The derivatives tested exhibited differential activity against anaero-
bic bacteria. The anaerobes were the most susceptible at concentrations
in ranges for ≤6.2 to 100 µg/mL to derivative: 9 (64% were susceptible),
1, 13 (for 60%) and 7 (56%).
The anaerobes were much less susceptible to the following deriva-
tives: 4 (for 96% strains MIC ≥200 µg/mL); 29, 15 (for 92% MIC
≥200 µg/mL); 16 (for 88% MIC ≥200 µg/mL).
The aerobic bacteria were generally not susceptible to concentrations
of the derivatives studied, with the exeption of derivative 10, which
was active against 52% of strains at concentrations in ranges of 50–100
µg/mL.
The derivatives had activities comparable to those of metronidazole
against anaerobic bacteria and comparable to those of amikacin against
aerobes (Table II).
EXPERIMENTAL
Melting points were determined with the Mettler FP-2 apparatus and
are uncorrected. The IR spectra were taken with Satellite FTIR spec-
1
trophotometer. The H NMR spectra were taken with a Varian Gem-
ini 200 BS-487c spectrometer. The microwave Plazmotronika (100 W,
500 MHz wave frequency) reactor was used for the syntheses of two
compounds.
5-N-Methyl-N-Phenyl-1,3,4-Thiadiazolo-2-Thiol (2)
4-phenyl-4-methyl-3-thiosemicarbazide (0.91 g, 5 mmole), DMF (3 mL),
and CS2 (1.5 mL) were refluxed for 4 h. Then ice water was added until
the product precipitated, which was collected and recrystallized.

Synthesis and Antibacterial Activity
621
2-Methylsulphonyl-5-N-Methyl-N-Phenyl-1,3,4-Thiadiazole (3)
To KOH (0.085 g, 1.5 mmole) dissolved in ethanol (5 mL), compound
2 (0.22 g, 1 mmole) was added and, when dissolved, was treated with
methyl iodide (0.1 mL, 1.5 mmole). The whole was refluxed for 0.5 h.
On cooling the precipitated product was collected and washed on the
filter with cold ethanol.
4-Phenyl-4-Methyl-1-Carbodithiosemicarbazide Acid (4)
To KOH (1.1 g, 20 mmole) dissolved in water (5 mL) and ethanol (20 mL),
4-phenyl-4-methyl-3-thiosemicarbazide (1.81 g, 10 mmole) was added
and, when dissolved, was treated with CS2 (2 mL). The solution was
stirred for 1.5 h. On acidifying the mixture with acetic acid, compound
4 precipitated.
4-Amino-5-N-Methyl-N-Phenyl-1,2,4-Triazolo-2-Thiol (5)
The mixture of compound 4 (0.26 g, 1 mmole), ethanol (5 mL), and 98%
hydrazine hydrate (0.15 mL, 3 mmole) was refluxed for 2 h. On cooling,
compound 5 precipitated and was filtered.
4-N-Substituted-1-Carbothioic Acid Hydrazide (6–10)
A. Compound 1 (1.81 g, 10 mmole) dissolved in hot acetonitrile (10 mL)
was treated with the appropriated amine (10 mmole). The mixture
was refluxed for 0.5 h. The mixture was cooled then the precipitate
was filtered and recrystallized.
B. Compound 11 (2 g, 8 mmole) dissolved in ethanol (20 ml) was treated
with 98%-hydrazine hydrate and refluxed for 16 h. On cooling the
precipitated compound 6 was filtered, washed thoroughly with wa-
ter and recrystallized.
4-Phenyl-Piperazin-1-Dithiocarbazoic Acid Methyl Ester (11)
4-phenylpiperazine (1.5 mL, 10 mmole) dissolved in ethanol (10 mL)
was treated with triethylamine (1.4 mL, 10 mmole) and carbon disul-
phide (0.75 mL, 12.5 mmole). The precipitate formed was dissolved with
water, then methyl iodide (0.63 mL, 10 mmole) was added. The mixture
was stirred for 1 h. The precipitate of compound 11 was filtered and
recrystallized.

622
K. Spalin´ska et al.
The Condensates of 4-N-Substituted Thiosemicarbazides
With 5-Nitrofuryl Aldehyde (12–14)
The compound 7, 8, or 9 (10 mmole), respectively, was dissolved in hot
ethanol (10 mL) and treated with a stoichiometric amount of 5 nitrofuryl
aldehyde. The mixture was refluxed for 5 min. and then allowed to stand
at an ambient temperature for 12 h. The precipitated compounds 12–14,
respectively, were collected and washed on the filter with ethanol.
Dithiocarbazoic Acid Methyl Ester (15)
To KOH (0.56 g, 10 mmole) dissolved in water (3 mL) and ethanol
(10 mL), 4-phenyl-4-methyl-3-thiosemicarbazide (1.8 g, 10 mmole) was
added, and, when dissolved, it was treated with carbon disulphide
(0.64 mL, 10 mmole). The mixture was stirred for 15 min., then methyl
iodide (0.63 mL, 10 mmole) was added and stirred for another 15 min.
On cooling the product precipitated.
Thiocarbamino Derivatives (16–20)
Compound 6 (0.6 g, 2.5 mmole) dissolved in hot ethanol (10 mL) was
treated with a stoichiometric amount of the appropriate isothiocyanate
and refluxed for 10 min. The products precipitated either while hot or
on cooling.
1,3,4-Thiadiazolo-2-Ylo-Amines (21–23)
Compounds 18–20 (1 mmole), respectively, dissolved in ethanol (15 mL)
were refluxed for 6 h. On cooling, the precipitate of compounds (21–23),
respectively, were filtered and recrystallized.
The Condensates of 4-N-Phenyl-Piperazinothiosemicarbazide
With Aldehydes (24–26)
Compound 6 (2.4 g, 10 mmole) dissolved in ethanol (10 mL) was treated
with the corresponding benzaldehyde (10 mmole). The mixture was re-
fluxed for 10 min. and allowed to stand at r.t. for 12 h. The precipitated
compounds (24–26) were filtered and recrystallized.
5-(4-Phenyl-Piperazin-1-Ylo)-[1,3,4]-Thiadiazolo-2-Thiol (27)
A. The mixture of compound 6 (1.2 g, 5 mmole), dimethylformamide
(3.0 mL), and carbon disulphide (1.5 mL) was refluxed for 4 h. Ice

Synthesis and Antibacterial Activity
623
water was added, and then the precipitated product was filtered and
recrystallized.
B. Compound 30b (1.75 g, 5 mmole) dissolved in dimethylsulphoxide
(3 mL) was irradiated with microwaves (3 min., temp. 170–180◦C).
When the reaction was completed, the mixture was treated with
ice (20 g) and acidified with acetic acid to pH 7. The precipitate of
compound 27 was filtered and recrystallized.
2-Methylsulphanyl-5-(4-Phenyl-Piperazin-1-Ylo)-[1,3,4]-
Thiadiazole (28)
To KOH (0.085 g, 1.5 mmole) dissolved in ethanol (5 mL), compound
27 (0.28 g, 1 mmole) was added and, when dissolved, was treated with
methyl iodide (0.7 mL, 1.5 mmole). The mixture was stirred for 0.5 h.
On cooling, the product precipitated.
5-(4-Phenylpiperazin)-2-Morpholino-1,3,4-Thiadiazole (29)
A. Compound 28 (0.3 g, 1 mmole) was refluxed with morpholine (3 mL)
and dimethylformamide (2 mL) for 30 h. On cooling the mixture, the
precipitated compound 29 was filtered.
B. Compound 30a (0.78 g, 2.5 mmole) was refluxed with morpholine
(2 mL) for 2 h. On cooling the mixture, the precipitated compound
29 was filtered.
N’-4-Phenyl-Piperazin-1-Carbothioyl)-
Hydrazinodithiocarbazoic Acid (30a)
To KOH (1.1 g, 20 mmole) dissolved in water (5 mL) and ethanol (20 mL),
compound 6 (2.4 g, 10 mmole) was added and, when dissolved, was
treated with carbon disulphide (2 mL). The mixture was stirred for 1.5 h,
and then acidified with acetic acid and cooled with ice; the precipitated
compound 30a was filtered.
N’-(4-Phenyl-Piperazin-1-Carbothioyl)-
Hydrazinodithiocarbazoic Acid potassium Salt (30b)
To KOH (0.56 g, 10 mmole) dissolved in ethanol (20 mL), compound 6
(2.36 g, 10 mmole) was added and stirred for 5 min. Then carbon disul-
phide (1.2 mL, 20 mmole) was added and stirred for another 45 min.
The reaction mixture was treated with anhydrous diethyl ether (20 mL),
and then the precipitate of salt 30b was filtered.

624
K. Spalin´ska et al.
4-Amino-5-(4-Phenyl-Piperazin-1-Ylo)-4H-[1,2,4]-Triazolo-3-
Thiol (31)
A. The mixture of compound 30b (0.35 g, 1 mmole), ethanol (5 mL),
and 98% hydrazine hydrate (0.1 mL, 2 mmole) was refluxed for 4 h.
Water (20 mL) was added, and then neutralized with acetic acid.
The precipitated compound 31 was filtered and recrystallized.
B. Compound 30b (1.75 g, 5 mmole) was dissolved in dimethylsulphox-
ide (3 mL) and 98% hydrazine hydrate (0.24 mL, 5 mmole) was
added. The mixture was irradiated with microwaves (3 min., temp.
170–180◦C). Ice (20 g) was added, and then the mixture neutralized
with acetic acid. The precipitate of compound 31 was collected and
recrystallized.
The Condensate of 4-Amino-5-(4-Phenylpiperazin)-1,2,4-
Triazolo-2-Thiol With P-Chloro-Benzoic Aldehyde (32)
Compound 31 (0.41 g, 1.5 mmole) was dissolved in hot ethanol (10 mL)
and treated with p-chlorobenzoic aldehyde (0.21 g, 1.5 mmole). The
mixture was refluxed for 15 min. Compound 31 precipitated while hot.
On cooling with ice, the product was filtered.
6-Phenyl-3-(4-Phenyl-Piperazin-1-Ylo)-7H-[1,2,4]-
Triazolo[3,4-b][1,3,4]thiazidine (33)
Compound 31 (0.69 g, 2.5 mmole) dissolved in hot absolute ethanol
(10 mL) was treated with phenacyl bromide (0.5 g, 2.5 mmole). The
whole was stirred under refux for 6 h. The reaction mixture was cooled to
r.t. and neutralized with an aqueous solution of Na2CO3. The precipitate
of compound 33 was filtered and recrystallized.
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