Synthesis and antibacterial activity of furo[3,2-b]pyrrole derivatives

Article abstract of DOI:10.24820/ark.5550190.p010.240

8-Ethoxyfuro[2′,3′:4,5]pyrrolo[1,2-d][1,2,4]triazine was synthesized by reaction of appropriate triazinone with POCl3 and subsequent treating of 8-chloro derivative with sodium ethoxide in ethanol. Furo[3,2-b]pyrrole-5- carboxylates were hydrolysed to for

Full text of DOI:10.24820/ark.5550190.p010.240

The Free Internet Journal
for Organic Chemistry
Paper
Archive for
Organic Chemistry
Arkivoc 2017, part v, 204-215
Synthesis and antibacterial activity of furo[3,2-b]pyrrole derivatives
Ivana Zemanová,a Renata Gašparová,a* Andrej Boháč,b Tibor Maliar,c Filip Kraic,d and Gabriela Addová e
a Department of Chemistry and c Department of Biotechnology, Faculty of Natural Sciences,
University of Ss. Cyril and Methodius, Námestie J. Herdu 2, 917 01 Trnava, Slovak Republic
b Department of Organic Chemistry and e Institute of Chemistry, Faculty of Natural Sciences,
Comenius University, Ilkovičova 6, 842 15 Bratislava, Slovak Republic
d Saneca Pharmaceuticals, Nitrianska 100, 920 01 Hlohovec, Slovak Republic
Email: renata.gasparova@ucm.sk
Received 06-28-2017
Accepted 09-22-2017
Published on line 10-16-2017
Abstract
8-Ethoxyfuro[2',3':4,5]pyrrolo[1,2-d][1,2,4]triazine was synthesized by reaction of appropriate triazinone with
POCl3 and subsequent treating of 8-chloro derivative with sodium ethoxide in ethanol. Furo[3,2-b]pyrrole-5-
carboxylates were hydrolysed to form acids, which underwent one-pot decarboxylation with TFA and
formylation of the in situ formed furo[3,2-b]pyrrole with triethyl orthoformate to give 5-carbaldehydes.
Hydrazinolysis of bis-esters led to bis-carbohydrazides which subsequently cyclized in acetic acid under
microwave irradiation to form either pyrazine or acetamide derivatives with unusual chirality. Prepared
compounds were evaluated for their antibacterial activity on Escherichia coli and Micrococcus luteus.
Keywords: Furo[3,2-b]pyrrole, cyclisation, formylation, antibacterial activity, microwave irradiation, chirality
DOI: https://doi.org/10.24820/ark.5550190.p010.240
Page 204
©ARKAT USA, Inc

Arkivoc 2017, v, 204-215
Zemanova, I. et al.
Introduction
Furo[3,2-b]pyrroles are isosteres of the indole ring system in which the benzene ring is replaced by a furan ring.
Interest in pyrrolo-fused heteroaromatic compounds such as furo-, thieno- and seleno-pyrroles stems mainly
from the array of interesting biological activities1-3 or their use as fluorescent dyes.4
More complex furo- and thieno-pyrroles, such as 6,7-dihydrofuro- and 6,7-dihydrothieno[2’,3’:4,5]pyrrolo-
[1,2-a]pyrazin-8(5H)-ones,5 furo[2,3:4,5]pyrrolo[2,1-c][1,4]oxazines,6 furo[2,3:4,5]pyrrolo[1,2-d][1,2,4]-triaz-
olo[3,4-f][1,2,4]triazines7 or thieno[3,2-b]pyrrolo[3,2-d]pyridazinones were synthesized and evaluated for their
anticancer activity.8 Moreover 2,3,5,7-tetrabromobenzofuro[3,2-b]pyrrole has been isolated from Pseudo-
alteromonas species and showed antimicrobial activity against methicillin-resistant Staphylococcus aureus.9
Among fused furo[3,2-b]pyrrole derived heterocycles, the synthesis of furo[2,3:4,5]pyrrolo[1,2-d][1,2,4]triazin-
8(7H)-ones has been studied extensively.7,10,11 However the preparation of appropriate 8-substituted derivatives
(in the triazine ring) is limited to 8-hydrazino7,10 or 8-methylsulfanyl12 compounds.
As part of our current studies on the development of new methods in heterocyclic synthesis,13,14 we report
an efficient synthesis of furo[3,2-b]pyrrole derivatives fused with six-membered rings and evaluation of the
synthesized compounds as to their antibacterial activity.
Results and Discussion
We have recently reported15 the synthesis of furo[2,3:4,5]pyrrolo[1,2-d][1,2,4]triazin-8(7H)-ones 2 from
substituted carboxylates 1. Herein we report the synthesis of furo[[2,3:4,5]pyrrolo[1,2-d][1,2,4]triazine
derivatives 3 and 4, 7-amino-2-methylfuro[[2,3:4,5]pyrrolo[1,2-a]pyrazine-6,8(5H,7H)-diones 9 and acet-
amides 10, as well as the synthesis of furo[3,2-b]pyrrole derived acids and aldehydes, in order to study their
antibacterial activity.
Scheme 1. Synthesis of furo[3,2-b]pyrrole derivatives 3-6.
Page 205
©ARKAT USA, Inc

Arkivoc 2017, v, 204-215
Zemanova, I. et al.
A convenient synthetic route to transform a triazinone into a substituted triazine ring consists in treating
the triazinone 2 with phosphorus oxychloride for 4h to give the 8-chloro substituted triazine 3, which was used
for the next step without further purification. Finally, use of sodium ethoxide resulted in the substitution of
1
compound 3 and the title 8-ethoxytriazine 4 was obtained in 57% yield after 48h of reflux (Scheme 1). The H
NMR spectrum of 4 shows the singlets of H-5, H-3 and H-9 at 7.93, 7.57, and 7.32 ppm, respectively. Protons of
the ethoxy group resonate as a quartet at 4.16 ppm and triplet at 1.97 ppm. The 13C NMR spectra of 4 display
the C-5 and C-8 carbons at 155.9 and 148.3 ppm, respectively.
The synthetic availability of the starting carboxylates 1 was also investigated with the intention to synthesize
furo[3,2-b]pyrrole derivatives for screening of their antibacterial activity. Thus, 2-triphenylmethyl-4H-furo[3,2-
b]pyrrole-5-carboxylate 1d was synthesized in high yield (89%) by reaction of 1a with triphenylmethyl chloride
in dimethylformamide and sodium hydride, according to the method used for the [2,3-b]-isomer16 (Scheme 1).
Compound 1d displays in its 1H NMR spectrum a singlet due to the NH group at 11.66 ppm, and singlets of H-6
and H-3 at 6.74 and 6.06 ppm, respectively. The 13C NMR spectrum of 1d shows the signal of ester carboxyl
carbon at 165.2 ppm. The proposed mechanism involves formation of pyrrole N-anion in the initial step of
reaction, but the direct bond interaction between bulky tritylium ion and pyrrole N-anion would be
thermodynamicaly unstable due to steric hindrance caused by the adjacent -COOMe group and the furane ring.
The ion pair can be formed initially, but thanks to the suitable mesomerism the relatively stable tritylium ion
can move towards the most electron-rich C-2 carbon, which is also stericaly more favorable (Scheme 2).
Scheme 2. Mechanism explaining the synthesis of methyl 2-trityl-4H-furo[3,2-b]pyrrole-5-carboxylate 1d.
Carboxylates 1 were converted into the appropriate 4H-furo[3,2-b]pyrrole-5-carboxylic acids 5a-5c in 66-
80% yields by hydrolysis of 1b-1d in aqueous NaOH for 1.5-4h (Scheme 1). Acids 5a-5c display in their 1H NMR
spectra a broad singlet at 12.8-12.13 ppm of carboxylic hydrogen and a singlet at 11.29-11.44 ppm due to the
NH group. The H-6 protons of 5a and 5c resonate as singlets at 6.52-6.65 ppm or, in case of 5b, as doublet at
6.58 ppm (J 1.8Hz). The IR spectra of 5a-5c exhibit absorption bands of the NH group at 3373-3190 cm-1 and the
carbonyl group at 1630-1640 cm-1. The 13C NMR spectra of 5a-5c display the carboxyl carbons at 163.1 ppm.
Furo[3,2-b]pyrrole derivatives can be formylated under Vilsmeier-Haack reaction conditions17 and 2-
formylated products are obtained preferably. When the C-2 position is occupied, the formylation at C-5 or N-4
can take place, while the C-6 position is the least reactive. The synthesis of 4H-furo[3,2-b]pyrrole-5-
carbaldehydes 6 was taken place by method of Umezawa,4 which consists in the decarboxylation of acids 5 with
trifluoroacetic acid and formylation of the in situ formed 4H-furo[3,2-b]pyrrole with triethyl orthoformate. The
resulting aldehydes 6a and 6b were obtained in 55 and 58% yields, respectively. Compounds 6a, 6b display in
1
their H NMR spectra the singlets at 11.68-11.75 ppm due to NH group and at 9.32-9.31 ppm of the formyl
group. The H-6 protons resonate as singlets at 6.76-6.82 ppm. IR spectra of 6 exhibit a formyl absorption band
at 1618-1626 cm-1. The 13C NMR spectra of 6a,b show the formyl carbon at 179.2 and 179.1 ppm, respectively.
Alkylation of methyl 4H-furo[3,2-b]pyrrole-5-carboxylates 1a, 1b and 1e with methyl chloroacetate in
dimethylformamide in the presence of sodium hydride at room temperature overnight provided methyl 4-(2-
methoxy-2-oxoethyl)-4H-furo[3,2-b]pyrrole-5-carboxylates 7a-7c in 61-67% yields (Scheme 3). The structures of
Page 206
©ARKAT USA, Inc

Arkivoc 2017, v, 204-215
Zemanova, I. et al.
compounds 7 were established by 1H NMR, 13C NMR and IR spectroscopy. 1H NMR spectra show doublets of H-
6 protons at 7.11- 6.78 ppm region (J 0.6 Hz), H-3 protons resonate either as doublets of doublets (7a) at 6.83
ppm (J 2.1, 0.9 Hz) or as singlets (7b, 7c) at 6.89 and 6.43 ppm, respectively. The CH2 protons appear as singlets
at 5.15-5.21 ppm. 13C NMR spectra show ester carboxyl carbon signals of at 168.6-169.6 ppm and 161.5-162.1
ppm. The characteristic bands observed at 1747-1754 and 1685 cm-1 in IR spectra of 7b and 7c correspond to
the ester carbonyl groups.
The presence of 2-methoxy-2-oxoethyl and methyl carboxylate groups in α-position enables the cyclisation
with hydrazine to afford ring with either N-aminoimide or N,N'-diacylhydrazine structural units. Monge and co-
workers18 have reported the reaction of methyl 2-(2-methoxy-2-oxoethyl)-1H-indole-3-carboxylate with 40%
hydrazine hydrate without any solvent for 36h to obtain 2,3,5,6-tetrahydro[1,2]diazepino[5,4-b]indole-1,4-
dione, while higher concentration of hydrazine led to the bis-hydrazide. On the other hand Voieduvskyi and co-
workers19 have synthesized a 2-amino-1,3-dioxo-1,2,3,4-tetrahydropyrrolo[1,2-a]pyrazine derivative by
reaction of an appropriate 1-(2-methoxy-2-oxoethyl)pyrrole-2-carboxylate with 95% hydrazine hydrate in
ethanol at 60 oC for 1h.
In order to synthesize new tricyclic 5-5-6 fused furo[3,2-b]pyrrole derivatives, we have realized the
cyclisation of 7 either by the method of Monge18 with 40% hydrazine overnight or by heating in ethanol19 for
48h. In both cases the cyclisation has not taken place and only bis-hydrazides 8a-8c were formed in 65-70%
yields. Compounds 8a-8c show in their 1H NMR spectra two singlets at 9.23-9.34 and 9.09-9.15 ppm regions due
to NH groups. Broad singlets of NH2 groups appear at 4.28-4.29 and 4.18-4.21 ppm, respectively and singlets of
CH2 group at 4.98-5.02 ppm. IR spectra of 8a-8c exhibit absorption bands of ester carbonyl groups at 1675-1644
cm-1. The 13C NMR spectra of 8a-8c display the hydrazide carbonyl carbons at 167.2 and 162.5 ppm.
Scheme 3. Synthesis of bis-hydrazides 8 and fused pyrazinediones 9 and 10.
The cyclisation of bis-hydrazides to pyridazine ring can be achieved by heating in acid media.20 Conventional
heating at 80-90o C was not effective, because the starting derivative 7 was observed on TLC after 3 days of
Page 207
©ARKAT USA, Inc

Arkivoc 2017, v, 204-215
Zemanova, I. et al.
heating, therefore we heated the bis-hydrazides 8 in acetic acid by microwave irradiation. When compounds 8a
or 8c were irradiated in microwave oven at 180W and 80o C for 35 min, acetamides 10a, 10b were synthesized
in 85 and 86% yields, respectively; irradiation of 8b for a shorter period (12 min) at 90 W and 80o C led to the
pyrazine 9 in 87 % yield (Scheme 3).
The structures of compounds 9 and 10 were established by 1H NMR, 13C NMR and IR spectroscopy. While
1H NMR spectrum of 9 show no signal of NH group and one singlet signal of NH2 group at 5.31 ppm, in the
spectra of 10a, 10b there is a singlet for one NH group at 10.41 ppm and no signals of NH2 groups were observed.
The most distinct signals of 10 in the 13C NMR spectra were the three carbonyl group signals at 168.6, 164.8 and
156.1 ppm. That of C-5 appears at 48.7 ppm, and the methyl carbon at 20.8 ppm. The characteristic bands
observed at 1729-1669 cm-1 in the IR spectra correspond to the C=O groups. By an analysis of 1H-NMR spectrum
of product 10b we observed an unexpected chemical shift doubling (5.43 and 5.32 δ) and splitting (19.1 Hz) of
the methylene hydrogens at C-5 (Figure 1).
Figure 1. Graphical diagram of 1H- and 13C-NMR spectral characteristics for particular H and C atoms. The exact
assignments are based on analysis of 1D and 2D NMR (HSQC and HMBC) spectra. The numbers in the
bottom picture represent observed HMBC interactions between hydrogens and carbons up to a distance
of three bonds.
The observed AB type of multiplicity for methylene group in 1H-NMR spectrum of 10a, 10b is a consequence
of diastereotopicity of methylene hydrogens in the chiral molecules 10. The chirality of 10 is either due to the
presence of a stereochemically unstable amidic nitrogen stereogenic unit and / or by an axial chirality based on
the electrostatic repulsion between two oxygens of dihydropyrazinedione and oxygen from a partially enolised
amidic carbonyl group that cannot freely rotate around the hydrazidic N-N bond in 10. Considering the above
effects two enantiomers of 10b can be drawn (Figure 2). According to our knowledge, this is the first time to
explain this kind of the chirality issue on unsymmetrical triacylhydrazide compounds, although some similar
structures have been described.21-23 Unfortunately no X-ray structure analysis or chirality study was performed
on these type of compounds.
Page 208
©ARKAT USA, Inc

Arkivoc 2017, v, 204-215
Zemanova, I. et al.
Figure 2. The structures of the two enantiomers of 10b are shown as an object and its non-superimposable
mirror image. The asterisk means either the central (A) or the axial (B) stereogenic unit in 10b responsible
for its chirality. The same is valid for a compound 10a (not shown here).
Antibacterial activity
Compounds 1e, 5a-5c, 6a, 7b, 7c and 8a-8c were screened for their antibacterial activity against G- bacterial
taxon Escherichia coli, CCM 7929 and G+ bacterial taxon Micrococcus luteus, CCM 732. The antibacterial activity
of all tested structures were compared with standard 6-aminopenicillanic acid (6-APA). The results are
presented in Table 1.
Table 1. Antibacterial activity of standard 6-APA and furo[3,2-b]pyrroles 1e-8c on a G- bacterium Escherichia
coli CCM 7929 and a G+ bacterium Micrococcus luteus CCM 732
MIC (mM)
Compound
Escherichia coli
3.84
Micrococcus luteus
5.12
6-APA
1e
3.84
5.12
5a
5b
5c
5.12
20.48
0.16
>20.48
>20.48
>0.1
6a
7b
7c
20.48
16.38
10.24
5.12
>20.48
>20.48
15.36
8a
5.12
8b
8c
10.24
2.56
>20.48
1.92
The antibacterial activity of the tested compounds, expressed as MIC parameter, is over a very wide range
(0.1-20.48 mM). Two compounds, 1e and 8a, are comparable with the activity of selected standard 6-APA
Page 209
©ARKAT USA, Inc

Arkivoc 2017, v, 204-215
Zemanova, I. et al.
against both bacterial species. Two compounds expressed a higher antibacterial activity than the standard,
particularly compound 8c with MIC value 2.56 mM on Escherichia coli and 1.92 mM on Micrococcus luteus and
mainly compound 5c with activity in micromolar range (MIC = 0.16 mM) on Escherichia coli and MIC value under
0.1 mM on Micrococcus luteus. Compounds 5a, 5b, 6a and 7b expressed antibacterial activity over the testing
range with MIC value >20.48 mM on Micrococcus luteus.
Because oxygen heteroatom of furane ring in furo[3,2-b]pyrrole core can serve as an acceptor of hydrogen
bond (AHb), there is necessary for better activity to offer functional groups - hydrogen bond donors (DHb),
because antibacterial effect is the result of either the receptor occupation or enzyme inhibition mechanism. This
is the probable reason for the lower MIC value for compounds 6a and 8b.
The promising antibacterial activity of compound 5c on both bacterial species could be explained by the
presence of bulky lipophilic triphenylmethyl substituent, representing the lipophilic tail of the molecule,
opposite to the carboxy group on the other side of the molecule. This construction leads to compounds with
"detergent" properties and contributes to the passage of the bacterial membrane barrier of such compounds.
Especially compound 5c should be a lead structure for the next generation of compounds with increased
antibacterial potency.
Conclusions
New furo[3,2-b]pyrrole derivatives and furo[3,2-b]pyrrolo-fused triazines, their carboxylic acids and aldehydes,
were synthesized. Hydrazinolysis of bis-esters 7 led to bis-carbohydrazides 8 which subsequently cyclized in
acetic acid under microwave irradiation to form either pyrazine 9 or the acetamides 10. The synthesized
compounds were evaluated for their antibacterial activity on Escherichia coli and Micrococcus luteus.
Micromolar range of activity on Escherichia coli was achieved in the case of 2-tritylfuro[3,2-b]pyrrole-5-
carboxylic acid.
Experimental Section
General. Melting points of products were determined on a Kofler hot plate apparatus. 1H NMR/13C NMR spectra
were obtained on a 300 MHz/75 MHz spectrometer Varian Gemini 2000 in DMSO-d6 with tetramethylsilane as
the internal standard. The infrared spectra were taken on Agilent Cary 630 FTIR spectrometer with diamond
ATR. Elemental analyses were performed on a Flash EA 2000 CHNS/O-OEA analyser. MS spectra were measured
at Agilent Technologies 1200 Series apparatus. All solvents were distilled and dried appropriately prior to use.
Microwave-assisted reactions were performed in an Initiator Biotage microwave synthesizer in a sealed vessel
and the reaction mixture was continuously stirred by magnetic stirring. The course of reactions was monitored
by TLC in ethyl acetate –hexane. Methyl 4H-furo[3,2-b]pyrrole-5-carboxylates 1a-1c were synthesized following
the published procedures.10,24 Other chemicals were purchased from the suppliers as the highest purity grade.
Bacteriological thermostat BT 120 (Czech Republic) was used for the cultivation of samples. All bacterial species
were purchased from the Czech Collection of Microorganisms - CCM (Brno, Czech Republic). Microplates were
purchased from VWR, Inc. (Vienna, Austria).
Methyl 2-(triphenylmethyl)-4H-furo[3,2-b]pyrrole-5-carboxylate (1d). A solution of ester 1a (0.83 g, 5 mmol)
in DMF (10 mL) was added slowly to NaH (0.24 g, 6 mmol) in DMF (15 mL). The mixture was stirred at 20 oC until
evolution of hydrogen ceased, then triphenylmethylchloride (1.39 g, 5 mmol) was added and stirring was
o
continued at 25 C overnight. The solution was poured into ice water (100 mL) and the precipitate was
Page 210
©ARKAT USA, Inc

Arkivoc 2017, v, 204-215
Zemanova, I. et al.
crystallized from methanol to give 1d as a white solid, yield 89%, mp 262-265 oC. 1H NMR (DMSO-d6) δ 11.66 (s,
1H, NH); 7.36-7.28 (m, 9H, H-3', H-4', H-5'); 7.07 (dd, J 8.4, 1.8 Hz, 6H, H-2', H-6'); 6.74 (s, 1H, H-6); 6.06 (d, 1H,
J 0.6 Hz, H-3); 3.77 (s, 3H, CH3). 13C NMR (75 MHz, DMSO-d6) δ 165.2, 161.6, 146.9, 144.1, 129.7, 129.2, 127.9,
126.9, 122.6, 100.5, 96.2, 61.5, 51.2. Anal. Calcd. for C27H21NO3 (407.47) C 79.59, H 5.19, N 3.44. Found: C 79.32,
H 5.16, N 3.42 %.
8-Chloro-2-methylfuro[2,3:4,5]pyrrolo[1,2-d][1,2,4]triazine (3). A mixture of triazinone 215 (1 g, 5.7 mmol) and
POCl3 (5 mL) was refluxed and stirred for 4h. After cooling, the mixture was poured into ice and neutralized with
ammonia (5 mL). The solid precipitate was washed with water and dried. The crude product was used without
isolation for the next reaction step.
8-Ethoxy-2-methylfuro[2,3:4,5]pyrrolo[1,2-d][1,2,4]triazine (4). A solution of chlorotriazine 3 (0.58g, 3.1
mmol) in absolute ethanol (5 mL) was slowly added to sodium ethoxide (0.02g, 8 mmol of Na) in ethanol (5 mL)
at 0°C. The reaction mixture was stirred at room temperature for 3h and then heated to reflux for 48h. After
cooling the solvent was evaporated, the crude solid product was filtered off, washed with water (15 mL) and
crystallized from ethanol to give 4 as yellow solid. Yield 57 %, mp 217-220 oC. 1H NMR (DMSO-d6) δ 7.93 (s, 1H,
H-5); 7.57 (s, 1H, H-6); 7.33 (s, 1H, H-3); 4.16 (q, 2H, CH2); 2.40 (s, 3H, CH3); 1.19 (t, 3H, J 6.9Hz, CH3). 13C NMR
(75 MHz, DMSO-d6) δ 155.9, 148.3, 147.2, 145.6, 127.7, 118.8, 96.5, 92.2, 56.5, 18.9, 15.1. Anal. Calcd. for
C11H11N3O2 (217.23) C 60.82, H 5.10, N 19.34. Found: C 61.16, H 5.18, N 19.78 %.
2-Substituted 4H-furo[3,2-b]pyrrole-5-carboxylic acids (5a-5c).
To a solution of 1 (2.8 g, 15 mmol) in ethanol (10 mL) was added NaOH (0.9 g, 22 mmol) in water (10mL) and
the mixture was refluxed for 1.5-4h. After cooling, the mixture was acidified with concentrated hydrochloric
acid. The resulting solid product was filtered off, washed with water and recrystallized from ethanol to give 5a-
5c as grey solids.
o
2-Methyl-4H-furo[3,2-b]pyrrole-5-carboxylic acid (5a). Yield 66%, mp 192-194 C. IR (KBr) /cm-1 3373 (NH),
1640 (C=O). 1H NMR (DMSO-d6) δ 12.13 (brs, 1H, OH); 11.29 (s, 1H, NH); 6.58 (s, 1H, H-6); 6.21(s, 1H, H-3); 2.34
(s, 3H, CH3). 13C NMR (75 MHz, DMSO-d6) δ 163.1, 158.8, 146.3, 130.4, 123.0, 96.2, 96.0, 15.1. Anal. Calcd. for
C8H7NO3 (165.15) C 58.18, H 4.27, N 8.48. Found: C 57.91, H 4.29, N 8.27 %.
2,3-Dimethyl-4H-furo[3,2-b]pyrrole-5-carboxylic acid (5b). Yield 70%, mp 162-164 oC. IR (KBr) /cm-1 3200 (NH),
1630 (C=O). 1H NMR (DMSO-d6) δ 12.08 (brs, 1H, OH); 11.38 (s, 1H, NH); 6.52 (d, J 1.8 Hz, 1H, H-6); 2.26 (s, 3H,
CH3); 2.01(s, 3H, CH3). 13C NMR (75 MHz, DMSO-d6) δ 163.2, 154.1, 144.9, 132.0, 122.8, 104.3, 95.9, 12.9, 8.3.
Anal. Calcd. for C9H9NO3 (179.18) C 60.33, H 5.06, N 7.82. Found C 69.88, H 5.11, N 7.15%.
2-Triphenylmethyl-4H-furo[3,2-b]pyrrole-5-carboxylic acid (5c). Yield 80%, mp 138-140 oC. IR (KBr) /cm-1 3190
(NH), 1640 (C=O). 1H NMR (DMSO-d6) δ 11.44 (s, 1H, NH); 7.33 (t, 3H, J 7.5, 3.6 Hz, H-4'); 7.27 (t, J 7.2, 3.6 Hz,
6H, H-3', H-5'); 7.04 (d, J 3.9 Hz, 6H, H-2', H-6'); 6.65 (s, 1H, H-6); 6.01(s, 1H, H-3). 13C NMR (75 MHz, DMSO-d6)
δ 165.1, 163.1, 147.4, 144.7, 130.2, 129.0, 128.4, 127.3, 124.3, 101.1, 96.4, 61.9. Anal. Calcd. for C26H19NO3
(393.44) C 79.37, H 4.87, N 3.56. Found: C 78.96, H 4.94, N 3.48%.
Substituted 4H-furo[3,2-b]pyrrole-5-carbaldehydes (6a, 6b).
Appropriate 4H-furo[3,2-b]pyrrole-5-carboxylic acid 5 (6 mmol) was dissolved in trifluoroacetic acid (8 mL) and
stirred at 50oC for 10 min. Triethyl orthoformate (1 mL) was added into the reaction mixture, and stirring
continued for further 10 min. After cooling, the reaction mixture was poured into saturated aqueous NaHCO3.
The formed precipitate was filtered off, washed with water and purified by column chromatography on SiO2
with n-hexane/ethyl acetate (75:25 to 60:40) to obtain compounds 6 as yellow solids.
o
2-Methyl-4H-furo[3,2-b]pyrrole-5-carbaldehyde (6a). Yield 55%, mp 162-164 C. IR (KBr) /cm-1 3198 (NH),
1618 (C=O). 1H NMR (DMSO-d6) δ 11.68 (s, 1H, NH); 9.32 (s, 1H, H-C=O); 6.82 (s, 1H, H-6); 6.29 (s, 1H, H-3); 2.37
Page 211
©ARKAT USA, Inc

Arkivoc 2017, v, 204-215
Zemanova, I. et al.
(s, 3H, CH3). 13C NMR (75 MHz, DMSO-d6) δ 179.3, 161.9, 147.1, 134.2, 133.6, 101.8, 96.2, 15.3. Anal. Calcd. for
C8H7NO2 (149.15) C 64.42, H 4.73, N 9.39. Found: C 64.81, H 4.69, N 8.94 %.
2,3-Dimethyl-4H-furo[3.2-b]pyrrole-5-carbaldehyde (6b). Yield 58%, mp 132-134 oC. IR (KBr) /cm-1 3205 (NH),
1626 (C=O). 1H NMR (DMSO-d6) δ 11.75 (s, 1H, NH); 9.31 (s, 1H, H-C=O); 6.77 (s, 1H, H-6); 2.29 (s, 3H, CH3); 2.02
(s, 3H, CH3). 13C NMR (75 MHz, DMSO-d6) δ 179.1, 157.4, 145.7, 135.8, 133.5, 104.3, 101.7, 13.1, 8.1. Anal. Calcd.
for C9H9NO2 (163.18) C 66.25, H 5.56, N 8.58. Found: C 66.03, H 5.59, N 8.21%.
Methyl 4-(2-methoxy-2-oxoethyl)-4H-furo[3,2-b]pyrrole-5-carboxylates (7a-7c).
A solution of 2-substituted methyl 4H-furo[3,2-b]pyrrolo-5-carboxylate 1 (0.66g, 4.0 mmol) in dimethyl
formamide (15 mL) was added to a suspension of NaH (0.5 g, 21 mmol) in dimethyl formamide (10 mL). After
stirring at room temperature for 5 min, methyl chloroacetate 0.8 mL (9 mmol) was added dropwise. The mixture
was stirred at room temperature overnight, then poured into ice water (20 mL) and HCl was added to reach pH
2. The mixture was extracted in diethylether (2 x 20 mL), the organic layer was dried with Na2SO4 and the solvent
was evaporated and and the solid products were crystallized from ethanol to obtain 7a-7c as yellow solids.
o
1
Methyl 4-(2-methoxy-2-oxoethyl)-4H-furo[3,2-b]pyrrole-5-carboxylate (7a). Yield 61%, mp 72-74 C. H NMR
(DMSO-d6) δ 7.83 (d, J 2.4 Hz, 1H, H-2); 6.89 (d, J 0.9 Hz, 1H, H-6); 6.83 (dd, J 2.4, 0.9, 1H, H-3); 5.20 (s, 2H, CH2);
3.73 (s, 6H, 2 x CH3). 13C NMR (75 MHz, DMSO-d6) δ 168.6, 161.5, 149.5, 144.7, 133.7, 122.8, 99.2, 98.5, 60.8,
51.1, 48.7. Anal. Calcd. for C11H11NO5 (237.21) C 55.70, H 4.67, N 5.90. Found: C 55.31, H 4.56, N 5.48 %.
Methyl 4-(2-methoxy-2-oxoethyl)-2-methyl-4H-furo[3,2-b]pyrrole-5-carboxylate (7b). Yield 55%, mp 95-97 oC.
1
IR (KBr) /cm-1 1754, 1685 (C=O). H NMR (300 MHz, DMSO-d6) δ 6.78 (d, 1H, J 0.6 Hz, H-6); 6.43 (s, 1H, H-3);
5.15 (s, 2H, CH2); 3.69 (s, 3H, CH3); 3.65 (s, 3H, CH3); 2.36 (s, 3H, CH3). 13C NMR (75 MHz, DMSO-d6) δ 169.6,
162.1, 159.7, 144.0, 135.3, 121.4, 98.9, 95.9, 52.4, 51.4, 48.9, 15.2. Anal. Calcd. for C12H13NO5 (251.24) C 57.37,
H 5.22, N 5.58. Found: C 56.99, H 5.29, N 5.69 %.
Methyl 4-(2-methoxy-2-oxoethyl)-2-(4-methoxyphenyl)-4H-furo[3,2-b]pyrrole-5-carboxylate (7c). Yield 67 %,
mp 173-175 oC. IR (KBr) /cm-1 1747, 1685 (C=O). 1H NMR (300 MHz, DMSO-d6) δ 7.67(d, 2H, J 4.5 Hz, H-2', H-
6'); 7.11 (s, 1H, H-6); 7.02 (d, 2H, J 4.2 Hz, H-3', H-5'); 6.89 (s, 1H, H-3); 5.21 (s, 2H, CH2); 3.78 (s, 3H, CH3); 3.72
(s, 3H, CH3); 3.67(s, 3H, CH3). 13C NMR (75 MHz, DMSO-d6) δ 169.6, 161.9, 159.9, 159.8, 144.5, 135.8, 125.8,
123.6, 122.6, 115.0, 99.0, 93.2, 55.7, 52.5, 51.6, 49.0. Anal. Calcd. for C18H17NO6 (343.34) C 62.97, H 4.99, N 4.08.
Found: C 63.22, H 5.08, N 4.53 %.
4-(2-Hydrazino-2-oxoethyl)-4H-furo[3,2-b]pyrrole-5-carbohydrazides (8a-8c).
Compounds 7a-c (10 mmol) were refluxed in ethanol (30 mL) with hydrazine hydrate (20 mmol) for 48h. After
cooling the solid compounds were filtered off and crystallized from ethanol to give 8a-8c as yellow solids.
o
1
4-(2-Hydrazino-2-oxoethyl)-4H-furo[3,2-b]pyrrole-5-carbohydrazide (8a). Yield 70%, mp 194-196 C; H NMR
(300 MHz, DMSO-d6) δ 9.34 (s, 1H, NH); 9.12 (s, 1H, NH); 7.65 (d, 1H, J 2.4 Hz, H-2); 6.78 (s, 1H, H-6); 6.64 (dd,
1H, J 2.1, 0.6 Hz, H-3); 5.02 (s, 2H, CH2); 4.28 (brs, 2H, NH2); 4.18 (brs, 2H, NH2). 13C NMR (75 MHz, DMSO-d6) δ
167.2, 162.1, 147.2, 144.9, 131.2, 125.9, 99.4, 93.6, 48.0. Anal. Calcd. for C9H11N5O3 (237.22) C 45.57, H 4.67, N
29.52. Found: C 45.98, H 4.66, N 28.87 %.
o
4-(2-Hydrazino-2-oxoethyl)-2-methyl-4H-furo[3,2-b]pyrrole-5-carbohydrazide (8b). Yield 70%, mp 188-190 C;
1
IR (KBr) /cm-1 3296, 3195, 2919, 2851 (NH), 1655, 1602 (C=O). H NMR (300 MHz, DMSO-d6) δ 9.23 (s, 1H, NH);
9.09 (s, 1H, NH); 6.72 (s, 1H, H-6); 6.26 (s, 1H, H-3); 4.98 (s, 2H, CH2); 4.21(brs, 4H, 2 x NH2); 2.33 (s, 3H, CH3). 13
C
NMR (75 MHz, DMSO-d6) δ 167.7, 162.8, 157.1, 144.2, 132.7, 124.5, 96.1, 94.1, 48.5, 15.1. Anal. Calcd. for
C10H13N5O3 (251.25) C 47.81, H 5.22, N 27.88. Found: C 47.59, H 5.19, N 28.04 %
4-(2-Hydrazino-2-oxoethyl)-2-(4-methoxyphenyl)-4H-furo[3,2-b]pyrrole-5-carbohydrazide (8c). Yield 65%, mp
263-266 oC; IR (KBr) /cm-1 3275, 3159, 2918, 2850 (NH), 1675, 1644 (C=O). 1H NMR (300 MHz, DMSO-d6) δ 9.34
(s, 1H, NH); 9.15 (s, 1H, NH); 7.65 (d, 2H, J 4.2 Hz, H-2', H-6'); 6.99 (d, 2H, J 4.4 Hz, H-3', H-5'); 6.96 (s, 1H, H-6);
Page 212
©ARKAT USA, Inc

Arkivoc 2017, v, 204-215
Zemanova, I. et al.
6.81 (s, 1H, H-3); 5.05 (s, 2H, CH2); 4.28 (brs, 2H, NH2); 4.20 (brs, 2H, NH2); 3.77(s, 3H, CH3). 13C NMR (75 MHz,
DMSO-d6) δ 167.7, 162.5, 159.3, 157.6, 144.9, 133.4, 125.8, 125.3, 124.3, 114.9, 94.2, 93.6, 55.7, 48.5. MS (ES):
m/z 342.0 (MH-). Anal. Calcd. for C16H17N5O4 (343.34) C 55.97, H 4.99, N 20.40. Found: C 56.28, H 4.94, N 19.97
%.
7-Amino-2-methylfuro[2',3':4,5]pyrrolo[1,2-a]pyrazine-6,8(5H,7H)-dione (9). Compound 8b (0.5g, 2 mmol)
was dissolved in acetic acid (5 mL) and irradiated in a microwave oven at 80 oC and 90W for 12 min. After cooling,
the solid product was filtered off, washed with water (20 mL) and crystallized from ethanol to give 9 as orange
solid. Yield 87%, mp 221-224 oC; IR (KBr) /cm-1 3337 (NH), 1665 (C=O). 1H NMR (300 MHz, DMSO-d6) δ 6.83 (s,
1H, H-6); 6.41 (s, 1H, H-3); 5.31 (s, 2H, NH); 5.13 (s, 2H, CH2); 2.39 (s, 3H, CH3). 13C NMR (75 MHz, DMSO-d6) δ
163.9, 160.4, 156.3, 146.5, 132.1, 120.6, 96.0, 95.0, 48.3, 15.2. Anal. Calcd. for C10H9N3O3 (219.20) C 54.79, H
4.14, N 19.17. Found: C 54.38, H 4.17, N 19.65 %.
N-(6,8-Dioxo-5,6-dihydrofuro[2',3':4,5]pyrrolo[1,2-a]pyrazin-7(8H)-yl)acetamides (10a, 10b). Compounds 8 (2
o
mmol) were dissolved in acetic acid (5 mL) and irradiated in a microwave oven at 80 C and 180W for 35 min.
After cooling the solid products were filtered off, washed with water (20 mL) and crystallized from ethanol to
give 10a, 10b as red solids.
N-(6,8-Dioxo-5,6-dihydrofuro[2',3':4,5]pyrrolo[1,2-a]pyrazin-7(8H)-yl)acetamide (10a). Yield 86%, mp 278-280
oC; IR (KBr) /cm-1 3191 (NH), 1729, 1669 (C=O). 1H NMR (300 MHz, DMSO-d6) δ 10.41 (s, 1H, NH); 7.91(d, 1H, J
2.4 Hz, H-2); 6.99 (d, 1H, J 0.9 Hz, H-9); 6.79 (dd, 1H, J 2.1, 0.6 Hz, H-3); 5.44 (d, 1H, J 19.6 Hz, H-5α); 5.34 (d, 1H,
J 19.6 Hz, H-5β); 1.99 (s, 3H, CH3). 13C NMR (75 MHz, DMSO-d6) δ 168.6, 164.9, 156.1, 151.2, 147.7, 131.2, 122.0,
99.9, 96.2, 48.7, 20.8. Anal. Calcd. for C11H9N3O4 (247.21) C 53.44, H 3.67, N 17.00. Found: C 52.98, H 3.71, N
17.32 %.
N-[2-(4-methoxyphenyl)-6,8-dioxo-5,6-dihydrofuro[2',3':4,5]pyrrolo[1,2-a]pyrazin-7(8H)-yl]acetamide (10b).
Yield 85%, mp 286-288 oC; IR (KBr) /cm-1 3180 (NH), 1729, 1670, 1607 (C=O). 1H NMR (300 MHz, DMSO-d6) δ
10.41 (s, 1H, NH); 7.75 (d, 2H, J 8.9 Hz, H-2', H-6'); 7.12 (d, 1H, J 0.8 Hz, H-3); 7.03 (d, 3H, J 8.7, Hz, H-9, H-3', H-
5'); 5.43 (d, 1H, J 19.1 Hz, H-5α); 5.32 (d, 1H, J 19.1 Hz, H-5β); 3.79 (s, 3H, CH3); 2.00 (s, 3H, CH3). 13C NMR (75
MHz, DMSO-d6) δ 168.6, 164.8, 160.9, 160.1, 155.8, 147.0, 133.4, 126.1, 123.3, 121.3, 115.1, 96.3, 93.3, 55.8,
48.7, 20.8. Anal. Calcd. for C18H15N3O5 (353.33) C 61.19, H 4.28, N 11.89. Found: C 60.94, H 4.16, N 11.26 %.
Antibacterial determination of Minimal Inhibition Concentration (MIC) parameters. On determination of MIC
parameters there were used sterile microplates (type P), where the suspension of bacterial species in nutrient
broth medium with dissolved tested compound has been achieved by convenient dilution method using
automatic multichannel pipets. The concentration from the column 1 to column 12 was in the decreasing order:
20.48 mM; 10.24 mM; 5.12 mM; 2.56 mM; 1.28 mM; 0.64 mM; 0.32 mM; 0.16 mM; 0.08 mM; 0.04 mM; 0.02
mM and 0.01 mM. The inoculum concentration of bacterial species suspension in nutrient broth medium was
before filling set by McFarland Densitometer DEN-1 (UK) on the value 0.1. The first two rows A and B were
occupied by the standard 6-aminopenicillanic acid (6-APA) on each microplate and the tested compounds were
in the rows C-G. After 24h of cultivation at 37 °C in the bacteriological thermostat 40 µl of 0.03% solution of
Thiazolyl Blue (MTT) in water was added to each well and incubated again for 1h under the same conditions.
Bacterial proliferation led to the production of bacterial mitochondrial dehydrogenase, which turned yellow
colored solution of MTT to intensely blue colored formazan product. MIC parameter was identified visually as
the last not colored well in the row. All experiments were carried out in triplicate.
Page 213
©ARKAT USA, Inc

Arkivoc 2017, v, 204-215
Zemanova, I. et al.
Acknowledgements
This work was supported by the Slovak Research Agency under the contract No. VEGA 1/0534/16. The authors
are indebted to Dr. Juraj Filo for 1D and 2D NMR spectra measurements.
Supplementary Material
1H and 13C NMR, and IR spectra of compounds 1e-10b.
References
1.
Sparey, T.; Abeywickrema, P.; Almond, S.; Brandon, N.; Byrne, N.; Campbell, A.; Hutson, P. H.; Jacobson,
M.; Jones, B.; Munshi, S.; Pascarella, D.; Pike, A.; Prasad, G. S.; Sachs, N.; Sakatis, M.; Sardana, V.;
Venkatraman, S.; Young, M. B. Bioorg. Med. Chem. Lett. 2008, 18, 3386.
https://doi.org/10.1016/j.bmcl.2008.04.020
2.
3.
Kawasima, Y.; Amanuma, F.; Sato, M.; Okuyama, S.; Nakashima, Y.; Sota, K.; Moriguchi, I. J. Med. Chem.
1986, 29, 2284.
https://doi.org/10.1021/jm00161a026
Zhuang, H. S.; Lin, Ch. Y.; Chou, Ch. L.; Hsu, H. M.; Lin, Y. H.; Huang, H. Ch.; Lien, Ch. J.; Kuo, CH. S; Huang,
J. L. Eur. J. Med. Chem. 2013, 66, 466.
https://doi.org/10.1016/j.ejmech.2013.06.012
4.
5.
Umezawa, K.; Nakamura, Y.; Makino, H.; Citterio, D.; Suzuki, K. J. Am. Chem. Soc. 2008, 130, 1550.
https://doi.org/10.1021/ja077756j
Ilyin, A.P.; Kobak, V. V.; Dmitrieva, I. G.; Peregudova, Y. N.; Kustova, V. A.; Mishunina, Y. S.; Tkachenko, S.
E.; Ivachtchenko, A. V. Eur. J. Org. Chem. 2005, 21, 4670.
https://doi.org/10.1002/ejoc.200500522
6.
7.
8.
Krutošíková, A.; Kryštofová-Labudová, L.; Dandárová, M. Chem. Heterocycl. Compd. 2001, 37, 1511.
https://doi.org/10.1023/A:1014557226260
Gajdoš, P.; Pavlíková, S.; Bureš, F.; Krutošíková, A. Cent. Eur. J. Chem. 2005, 3, 311.
https://doi.org/10.2478/BF02475999
Jiang, J.-k.; Boxer, M. B.; Vander Heiden, M. G.; Shen, M.; Skoumbourdis, A. P.; Southall, N.; Veith, H.;
Leister, W.; Austin, C. P.; Park, H. W.; Inglese, J.; Cantley, L. C.; Auld, D. S.; Thomas, C. J. Bioorg. Med. Chem.
Lett. 2010, 20, 3387.
https://doi.org/10.1016/j.bmcl.2010.04.015
9.
Fehér, D.; Barlow, R.; McAtee, J.; Hemscheidt, T. K. J. Nat. Prod. 2010, 73, 1963.
https://doi.org/10.1021/np100506z
10. Krutošíková, A.; Kováč, J.; Dandárová, M. Collect. Czech. Chem. Commun. 1984, 49, 65.
http://doi.org/10.1135/cccc19840065
11. Bobošík, V.; Krutošíková, A.; Dandárová, M. Collect. Czech. Chem. Commun. 1995, 60, 709.
http://doi.org/10.1135/cccc19950709
12. Krutošíková, A.; Mastik, S.; Dandárová, M.; Lyčka, A. Collect. Czech. Chem. Commun. 1997, 62, 1612.
https://doi.org/10.1135/cccc19971612
Page 214
©ARKAT USA, Inc

Arkivoc 2017, v, 204-215
Zemanova, I. et al.
13. Zemanová, I.; Gašparová, R.; Nova Biotech. Chem. 2013, 12, 100.
http://doi.org/10.2478/nbec-2013-0012
14. Gašparová, R.; Moncman, M.; Horváth, B. Cent. Eur. J. Chem. 2008, 6, 180.
https://doi.org/10.2478/s11532-008-0009-4
15. Zemanová, I; Gašparová, R.; Kraic, F.; Kružlicová, D.; Maliar, T.; Boháč, A.; Addová, G. Arkivoc 2017, (iv),
184.
http://doi.org/10.3998/ark.5550190.p009.788
16. Sleziak R., Krutošíková; A., Cyrański M.K., Krygowski T.M. Polish. J. Chem. 2000, 74, 207.
17. Kráľovičová, E.; Krutošíková, A.; Kováč, J.; Dandárová, M. Collect. Czech. Chem. Commun. 1986, 51, 106.
http://doi.org/10.1135/cccc19860106
18. Monge, A.; Palop, J. A.; Goni, T.; Martinez, A.; Fernandez-Alvarez, E. J. Heterocycl. Chem. 1984, 21, 381.
https://doi.org/10.1002/jhet.5570210221
19. Voievudskyi, M.; Astakhina, V.; Kryshchyk, O.; Petruhova, O.; Shyshkina, S. Monatsh. Chem. 2016, 147,
783.
https://doi.org/10.1007/s00706-015-1619-0
20. Biagi, G.; Ciambrone, F.; Giorgi, I.; Livi, O.; Scartoni, V.; Barili, P. L. J. Heterocyclic Chem. 2002, 39, 889.
https://doi.org/10.1002/jhet.5570390506
21. Haggag, B. M.; Allam, Y. A. Egypt. J. Chem.1999, 42, 199.
22. Takahashi, M.; Shinoda, T.; Osada, H.; Nakajima, T. Bull. Chem. Soc. Japan 1975, 48, 2915.
https://doi.org/10.1246/bcsj.48.2915
23. Okubo, T.; Iida, K.; Nitta, H.; Kitao, A. U.S. Patent 20150232678 A1, 2015.
24. Krutošíková, A.; Dandárová, M.; Chylová, J.; Vegh, D. Monatsh. Chem. 1992, 123, 807.
https://doi.org/10.1007/BF00812330
Page 215
©ARKAT USA, Inc