Synthesis and Pharmacological Activity of Tris(2-hydroxyethyl)ammonium 4-Chlorophenylsulfonylacetate

Article abstract of DOI:10.1134/S1070427218040249

Methods for the synthesis of 4-chlorophenylsulfonylacetic acid and its tris(2-hydroxyethyl)ammonium salt (“sulfacetamine”) were studied. The results of in vitro and in vivo experiments showed that sulfacetamine possesses high antithrombotic, antioxidant, hypocholesterolemic, immunostimulating, and protective-adaptive activities.

Full text of DOI:10.1134/S1070427218040249

ISSN 1070-4272, Russian Journal of Applied Chemistry, 2018, Vol. 91, No. 4, pp. 701−705. © Pleiades Publishing, Ltd., 2018.
Original Russian Text © S.N. Adamovich, E.N. Oborina, A.N. Mirskova, 2018, published in Zhurnal Prikladnoi Khimii, 2018, Vol. 91, No. 4, pp. 602−606.
BRIEF
COMMUNICATIONS
Synthesis and Pharmacological Activity
of Tris(2-hydroxyethyl)ammonium
4-Chlorophenylsulfonylacetate
S. N. Adamovicha,b, E. N. Oborinaa,b, and A. N. Mirskovaa
a Favorsky State Institute of Chemistry, Siberian Branch, Russian Academy of Sciences,
ul. Favorskogo 1a, Irkutsk, 664033 Russia
b Irkutsk Scientific Center, Siberian Branch, Russian Academy of Sciences,
ul. Lermontova 134, Irkutsk, 664033 Russia
e-mail: mir@irioch.irk.ru
Received April 12, 2018
Abstract—Methods for the synthesis of 4-chlorophenylsulfonylacetic acid and its tris(2-hydroxyethyl)ammonium
salt (“sulfacetamine”) were studied. The results of in vitro and in vivo experiments showed that sulfacetamine
possesses high antithrombotic, antioxidant, hypocholesterolemic, immunostimulating, and protective-adaptive
activities.
DOI: 10.1134/S1070427218040249
Ethanolammonium
fanyl)(sulfonyl)acetic
salts
acids
of
aryl(oxy)(sul-
It had been demonstrated that SA in microconcen-
trations (10–5 to 10–8 mol L–1), while being low-toxic
(LD50 = 6000 mg kg–1), exhibits high physiological
(specifically, growth-promoting) activity in the cultiva-
tion of beneficial bacteria, baker’s yeast, edible citric
acid, fungi, as well as in the technology of barley germi-
nation to produce brewing malt [9].
ArХCH2COO–·
HN+RR1(CH2CH2OH)n, where Ar = Ph etc.; X = O, S,
SO2; R, R1 = H, Alk; n = 1–3, are distinguished with high
biological activity. Among them, compounds exhibiting
adaptogenic, hemopoiesis-modulating, immunomodulat-
ing, anticancer, antimetastatic, and/or antiinflammatory
activities, as well as compounds increasing animals’ tol-
erance to hypoxia, hyper- and hypothermia, alcohol and
heavy metal intoxication, and/or to microwave radiation
had been discovered [1–8].
The purpose of this work was to develop an optimal
method for synthesis of SA, as well as to study its phar-
macological activity.
4-Chlorophenylsulfonylacetic acid (1), as the basic
starting compound for SA preparation, was synthesized
in 90% yield by selective oxidation of 4-chlorophenyl-
sulfanylacetic acid (2) with an excess of 30% hydrogen
peroxide in glacial acetic acid:
tris(2-Hydroxyethyl)ammonium4-chlorophenylsulfo-
nylacetate, 4-ClC6H4SO2CH2COO–·HN+(CH2CH2OH)3,
which we refer to as “sulfacetamine” (SA), represents
new biologically active substances of practical interest.
4-ClC6H4SCH2COOH + 2H2O2 → 4-ClC6H4SO2CH2COOH + 2H2O
2
1
As a more convenient and environmentally friendly
method for synthesis of 1 based on commercially avail-
able products, the reaction of sodium 4-chlorophenyl-
The weakness of this method is the need to initially
prepare acid 2 by interaction of toxic 4-chlorophenyl-
thiol with monochloroacetic acid (MCAA).
701

702
ADAMOVICH et al.
sulfinate with MCAA was investigated:
4-ClC6H4SO2Na + ClCH2COOH → 1 + NaCl
In a series of in vitro experiments, SA was found to
inhibit by 80–85% the platelet aggregation induced by
adenosine diphosphate (ADP) and almost completely
(up to 99%) inhibit the platelet aggregation induced by
serotonin, adrenaline, histamine, and thrombin.
The yield of acid 1 depends on the basicity of the
medium, the temperature, and the reaction time.
In in vivo experiments, with intravenous administra-
tion of SA to rabbits, the ADP-induced platelet aggrega-
tion was reduced by 57% compared to the control.
When the reaction was conducted in an aqueous
medium at pH 10 and 100°C for 1 hour, the desired
product was obtained in a high yield (92%). However, as
1 undergoes decarboxylation, the reaction is complicated
by the formation of sulfone 3 as a by-product:
SA slows down (from 50 to 234 s) the initiation of
blood clotting in as soon as 15 min.
In the “thromboplastin time” test, the effect of SA
also manifested itself after 15 min and lasted 24 h.
1 → 4-ClC6H4SO2CH3 + CO2
3
The study of SA’s antithrombotic effect in an acute
thrombosismodelrevealeditsdistinctprotectiveactivity.
For example, administration of SA to rats at a dose of
2 mg kg–1 followed by administration of thrombin (at
a dose lethal to 70% of animals) increased the survival
rate of animals by 50–65%.
Formation of 3 was avoided by replacing MCAA
with its methyl ester 4. The latter, like MCAA, is a
commercially available product for industrial organic
synthesis. Ester 5 was synthesized in quantitative yield.
Hydrolysis of ester 5 gives rise to acid 1 in high yield
and purity:
The processes of lipid peroxidation (LPO) of
membranes can activate blood clotting. In order to
find drugs that combine antiaggregant and membrane-
stabilizing properties, the effect of SA on the LPO of
platelet membranes was examined.
4-ClC6H4SO2Na + ClCH2COOCH3
4
H2O/Na2CO3
→ 4-ClC6H4SO2CH2COOCH3
1
5
SA’s effect on LPO was assessed from the
accumulation of malonic dialdehyde (MDA) in platelets
as a result of their interaction with thiobarbituric acid
(TBA).
Thus, the environmentally friendly and technologi-
cally sound method for synthesizing acid 1 is the reac-
tion of sodium 4-chlorophenylsulfinate with the methyl
ester of monochloroacetic acid followed by hydrolysis
of the ester formed.
Incubation of platelets with disodium salt of ADP
leads to a significant increase in their MDA level, up
to 8-fold of the original quantity. On the contrary, the
incubation of platelets with SA reduces the content of
TBA products by a factor of 3 compared to the control.
Then tris(2-hydroxyethyl)ammonium 4-chlorophe-
nylsulfonylacetate (SA) was synthesized in 90% yield
through the reaction of acid (1) with triethanolamine in
an alcohol medium:
One promising pathway to influence the thrombotic
process is the use of drugs that combine antiaggregant
activity and antioxidant activity (AOA). SA’s AOA
was examined in comparison with a known antioxidant
such as ionol. SA in a concentration of 10–6 mol L–1
demonstrated a high AOA (57%), thus indicating a
much higher activity than ionol (15% at 10–3 mol L–1,
i.e. even at a dose three orders of magnitude higher).
4-ClC6H4SO2CH2COOH + N(CH2CH2OH)3
→ 4-ClC6H4SO2CH2COO–·HN+(CH2CH2OH)3 (SА)
The composition and structure of the synthesized
compoundswere confirmedbyIR, 1Н, 13С, and15NNMR
spectroscopy, elemental analysis and potentiometric
titration.
The pathogenesis of atherosclerosis is closely
associated with thrombosis and hypercholesterolemia,
which promote the development of cardiovascular
lesions. The most promising pathway to treat these
diseases is the use of drugs that combine antithrombotic,
Together with the Research Institute of Fundamen-
tal and Clinical Immunology, Novosibirsk, Russia, the
antithrombotic, antioxidant, hypocholesterolemic, im-
munostimulating, and protective-adaptive activities of
sulfacetamine (SA) were studied.
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SYNTHESIS AND PHARMACOLOGICAL ACTIVITY
703
4-Chlorophenylsulfonylaceticacid(1).(a)10mLof
30%H2O2 wasaddeddropwisewithstirringtoasolution
of 10.13 g (0.05 mol) of 4-chlorophenylsulfonylacetic
acid (2) in 10 mL of acetic acid. The mixture was
stirred at 20°C for 3 h. Then it was heated in a boiling
water bath for 15 min. The solvent was distilled off
in vacuo before a precipitate appeared, dissolved in
the 10-fold amount of water, cooled to 5°C, and held
for 12 h. The precipitated crystals were filtered off
and dried in vacuo over P2O5. 10.67 g (91%) of acid 1
antioxidant, and hypocholesterolemic activities with
low toxicity.
The hypocholesterolemic activity of SA was
investigated in outbred white rats.
SA caused a decrease in cholesterol by 53% at a dose
of 25 mg kg–1 and by 63% at a dose of 50 mg kg–1, as
compared to the control. At the same time, a known
hypocholesterolemic drug such as “clofibrate” at a dose
of 25 and 50 mg kg–1 lowered the cholesterol level by
only 25% and 27%, respectively. The effect of SA at a
dose of 25 mg kg–1 not only reduces the total cholesterol,
but, most importantly, decreases the atherogenic
lipoproteins in the blood by 30%.
1
was obtained, mp 120–122°C. Н NMR spectrum, δ,
ppm (СD3OD): 7.36–7.30 m (4Н, С6Н4), 4.06 s (2Н,
СН2СОО). 13С NMR spectrum, δ, ppm (CD3OD):
169.99 (С=О), 146.81 (С6Н4SО2), 135.78–111.55
(С6Н4), 62.89 (СН2СОО). IR spectrum, ν, cm–1:
1720 (СОО), 1150, 1330 (SO2), 3380 (ОН). Found,
%: C 40.79, H 2.92, Cl 15.02, S 13.50. C8H7ClO4S.
Calculated (%): C 40.94, H 3.01, Cl 15.11, S 13.66.
The immunotropic activity of SA was studied in
hybrid lines of mice. SA’s ability to increase the content
of white blood cells and immunoglobulin-M-antibody-
forming cells (IgM AFC) is established, which indicates
its immunostimulating activity.
The protective properties of SA against hypoxia and
physical exercise were studied by its intraperitoneal
administration to white outbred mice. The drug was
used for 1 month at doses of 4 and 60 mg kg–1.
(b) Sodium 4-chlorophenylsulfinate [9.93
g
(0.05 mol)] and monochloroacetic acid [4.72 g
(0.05 mol)] were dissolved in 20 mL of water, adjusted
to pH 10 with a NaOH solution, and boiled in a water
bath for 1 h. The reaction mixture was cooled, and ex-
tracted with benzene (2 × 30 mL). From the benzene
solution, after distilling off the solvent, 0.67 g (7%)
of sulfone (3), i.e., 4-СlC6H4SO2CH3 was obtained,
mp 93–95°C. Found, %: С 44.30, Н 3.58, Cl 18.41,
S 16.67. С7Н7ClО2S. Calculated (%): С 44.10, Н 3.71,
Cl 18.59, S 16.82
The hypoxia tolerance was measured in a Ku-7 pres-
sure chamber, by determining the maximum “climb” al-
titude and the maximum residence time of animals at an
altitude of 12000 m until they assume a lateral position.
With SAadministered, the animals’stay at an altitude
of 12000 m was found to be 3 times longer than in the
control. The “climb” ceiling was 3000 m higher for the
tested animals in comparison with the control.
The aqueous layer was acidified with HCl until
neutral and held at 5°C for 12 h. The precipitate was
recrystallized (chloroform–hexane, 1 : 2) and dried in
vacuo over P2O5. 10.78 g (92%) of acid (1) was obtained,
mp 120–122°C.
The tolerance of animals to physical exercise was
assessed from the duration of their swimming in water
at 18°C until the first immersion.
The administration of SA was demonstrated to
increase the tolerance to physical exercise. The mice’s
swimming duration more than doubled as compared to
intact animals.
Methyl ester of 4-chlorophenylsulfonylacetic acid
(5). A solution of 108.53 g (1 mol) of methyl ester of
monochloroacetic acid (4) in 200 mL of DMF was add-
ed to a suspension of 198.6 g (1 mol) of sodium 4-chlo-
rophenylsulfinate (prepared by reacting 4-chlorophenyl-
sulfochloride with sodium sulfite in 500 mL of DMF)
heated to 45°C. The reaction mixture was heated with
stirring to 115°C for 3 h. After cooling, it was poured
into 3 L of cold water and held in a refrigerator for 12 h.
The precipitate was filtered off, recrystallized from chlo-
roform–hexane, and dried over P2O5. 233.7 g (98%) of
ester 5 was obtained, mp 81–82°C. 1Н NMR spectrum,
EXPERIMENTAL
The IR spectra of suspensions of test substances in
vaseline oil or in KBr pellets were recorded on a Var-
ian 3100 FTIR75 spectrophotometer, while the 1H, 13C,
15
and N NMR spectra in D2O or methanol D4 were re-
corded on a DPX 400 spectrometer (400.13 MHz for
1H, 101.62 MHz for 13C, and 40.18 MHz for 15N) at
T = 25°C.
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ADAMOVICH et al.
δ, ppm (СD3OD): 7.40–7.31 m (4Н, С6Н4), 4.05 s (2Н,
СН2СОО), 3.29 s (3Н, ОСН3). IR spectrum, ν, cm–1:
1735 (СОО), 1150, 1330 (SO2). Found, %: C 43.80,
H 3.45, Cl 14.11, S 13.12. C9H9ClO4S. Calculated, %:
C 43.46, H 3.66, Cl 14.25, S 12.89.
age), outbred white rats, and rabbits according to the
methods described in [10, 11].
CONCLUSIONS
(1) An environmentally friendly and technological-
ly sound method for synthesis of high purity 4-chloro-
phenylsulfonylacetic acid and its tris(2-hydroxyethyl)
ammonium salt (sulfacetamine) has been developed.
Hydrolysis of methyl ester of 4-chlorophenylsul-
fonylacetic acid (5). 24.8 g (0.1 mol) of ester 5 was
added to a solution of 10.6 g (0.1 mol) of Na2CO3 in
60 mL of water. The reaction mixture was stirred at
45°C for 12 h and filtered. The filtrate was acidified
with HCl to pH 2 and held at 5°C for 12 h. The precipi-
tate was separated, washed several times with water,
and dried in air, then recrystallized from chloroform–
hexane and dried in vacuo over P2O5. 20.06 g (90%)
of acid 1 was obtained, mp 120–122°C. The content
of the main substance, determined by potentiometric
titration, is 99.8%.
(2) In both in vitro and in vivo experiments, sulfac-
etamine exhibits antithrombotic, membrane-stabilizing,
and antioxidant activities and lowers cholesterol in the
blood. When applied in low doses, sulfacetamine ex-
hibits pronounced immunostimulating and protective-
adaptive activities.
ACKNOWLEDGMENTS
tris(2-Hydroxyethyl)ammonium 4-chlorophenyl-
sulfonylacetate (SA). A solution of 14.9 g (0.1 mol)
of freshly distilled triethanolamine in 20 mL of meth-
anol was added to a solution of 23.4 g (0.1 mol) of
acid 1 in 50 mL of methanol. The reaction mixture was
boiled for 15 min, cooled, and poured into 150 mL of
ester. The precipitate was filtered off, washed repeat-
edly with ester, and dried in vacuo. 34.6 g (90%) of
SA was obtained, mp 94°C. The content of the main
substance is 99.7%. 1Н NMR spectrum, δ, ppm (D2O):
7.76–7.51 m (4Н, С6Н4), 4.07 s (2Н, СН2СОО), 3.82 t
(6Н, ОСН2), 3.34 t (6Н, NСН2). 13С NMR spectrum,
δ, ppm (D2O): 167.27 (С=О), 140.27 (С6Н4SО2),
135.75–114.45 (С6Н4), 62.45 (СН2СОО), 54.91
(ОСН2), 54.63 (NСН2). 15N NMR spectrum, δ, ppm
(CD3OD): –340.43 (with respect to МеNО2). IR spec-
trum, ν, cm–1: 164 0 (СОО–), 1180, 1320 (SO2), 3380
(ОН). Found, %: C 43.91, H 5.69, Cl 9.20, N 3.72,
S 8.29. C14H22ClNO7S. Calculated (%): C 43.80,
H 5.79, Cl 9.23, N 3.65, S 8.35.
The authors are grateful to O. P. Kolesnikova
(Research Institute of Fundamental and Clinical
Immunology, Novosibirsk, Russia) for her participation
in biological research.
The major results were obtained using the equipment
of the Baikal Analytical Center for Collective Use
(Siberian Branch, Russian Academy of Sciences),
Irkutsk, Russia.
The work was performed within the framework of
the Integration Program of the Irkutsk Scientific Center
(Siberian Branch, Russian Academy of Sciences)
“Fundamental research and breakthrough technology as
the basis for the advanced development of the Baikal
region and its interregional relations.”
The study was carried out with the financial support
of the Russian Foundation for Basic Research (RFBR)
and the Government of the Irkutsk Region within the
framework of the research project No. 17-43-380006.
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hypocholesterolemic, immunotropic, and protective-
adaptive activities of SA were studied in hybrid
(CBAxC5BL/6)F1 mice of both sexes (8–10 weeks of
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