A new approach to the synthesis of rare thiazino[6,5-b]indol-4-one derivatives. First total synthesis of the indole phytoalexin cyclobrassinon

Article abstract of DOI:10.1016/S0040-4020(02)01124-9

The first synthesis of the indole phytoalexin cyclobrassinon and some of its analogues, possessing a thiazino[6,5-b]indol-4-one tricyclic ring system was performed starting from 1-substitued 2-chloroindole-3-carboxaldehydes. The route employed the intramolecular Et₃N-mediated or photochemical nucleophilic substitution of a chlorine atom in the 2-position of the indole ring with a sulfur atom as a key step. Examination of biological activity against the selected tumor cell lines, bacteria and fungi revealed no expressive activity of synthesized compounds.

Full text of DOI:10.1016/S0040-4020(02)01124-9

Tetrahedron 58 (2002) 9029–9039
A new approach to the synthesis of rare thiazino[6,5-b]indol-4-one
derivatives. First total synthesis of the indole
phytoalexin cyclobrassinon
a
b
a
c
Peter Kutschy,a Mojmır Suchy, Aldo Andreani, Milan Dzurilla, Vladimır Kovacik,
,
*
´
Juraj Alfoldi, Maddalena Rossi and Maria Gramatova
´
´
´ˇ
c
b
a
´
´
¨
a
ˇ
´
ˇ
Faculty of Science, Institute of Chemical Sciences, P.J. Safarik University, Moyzesova 11, 041 67 Kosice, Slovak Republic
bDepartment of Pharmaceutical Sciences, University of Bologna, Via Belmeloro 6, I-40126 Bologna, Italy
´ ´
Institute of Chemistry, Slovak Academy of Sciences, Dubravska cesta 9, 842 38 Bratislava, Slovak Republic
c
Received 27 June 2002; revised 13 August 2002; accepted 5 September 2002
Abstract—The first synthesis of the indole phytoalexin cyclobrassinon and some of its analogues, possessing a thiazino[6,5-b]indol-4-one
tricyclic ring system was performed starting from 1-substitued 2-chloroindole-3-carboxaldehydes. The route employed the intramolecular
Et3N-mediated or photochemical nucleophilic substitution of a chlorine atom in the 2-position of the indole ring with a sulfur atom as a key
step. Examination of biological activity against the selected tumor cell lines, bacteria and fungi revealed no expressive activity of synthesized
compounds. q 2002 Elsevier Science Ltd. All rights reserved.
1. Introduction
indolyl)]thioureas were heated with polyphosphoric acid,
1,3-thiazino[5,4-b]indol-4-one derivatives with interesting
serine protease inhibitory activity were obtained.2 Com-
pounds of type B, 1,3-thiazino[6,5-b]indoles, 4–10 are the
indole phytoalexins (Fig. 2).
1,3-Thiazinoindoles represent an interesting group of
heterocyclic compounds, because of their unusual chemical
structure and interesting biological properties. There are
four possible types (A–D) of ring fusion of the indole at the
bond b and the 1,3-thiazine ring or its hydrogenated forms
(Fig. 1).
Phytoalexins are low molecular weight secondary
metabolites, produced by plants after their exposure to
physical, biological or chemical stress.3 A specific group of
these natural products represent the indole phytoalexins,
produced by economically important plants of the family
Cruciferae, cultivated worldwide.4 With respect to the well
known anticarcinogenic properties of brassica vegetables5 it
The compounds of type A, possessing a 1,3-thiazino[5,4-
b]indole ring system, were prepared for the first time by
treatment of 3-aminoindole with carbon disulfide in
ethanol.1 When N0-alkyl or N0-aryl-N-3[(2-ethoxycarbonyl-
Figure 1.
Keywords: indoles; phytoalexins; natural products; substitution;
photochemical; antitumor activity; antimicrobial activity.
*
Corresponding author. Tel.: þ421-55-62-22610x192; fax: þ421-55-62-
22124; e-mail: kutschy@kosice.upjs.sk
Figure 2.
0040–4020/02/$ - see front matter q 2002 Elsevier Science Ltd. All rights reserved.
PII: S0040-4020(02)01124-9

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P. Kutschy et al. / Tetrahedron 58 (2002) 9029–9039
Scheme 1.
is important to study the biological activity of the indole
phytoalexins. Isolation from plants is difficult and time
consuming and therefore the amounts required for screening
should be provided by synthesis. Approximately 30 indole
phytoalexins have been isolated to date4b and biological
screening has disclosed antifungal,6 canceroprotective7 and
antitumor8 activity of several compounds. The synthesis and
biological properties of cyclobrassinon (8), isolated in 1994
from kohlrabi,9 have not been described to date. Cyclo-
brassinin (4) was prepared by cyclization of brassinin (1)
with pyridinium tribromide10 or N-bromosuccinimide
(NBS).7b Sinalbin B (6) has been synthesized analogously,
by cyclization of 1-methoxybrassinin (2) with N-bromo-
succinimide.11 The subsequent oxidation of 6 with 3-chloro-
peroxybenzoic acid afforded sinalbin A (7).11 Treatment of
4-methoxybrassinin (3) with NBS, followed by dehydro-
bromination, employing 1,8-diazabicyclo[5.4.0]undec-7-
ene (DBU) afforded an unstable 4-methoxycyclobrassinin
(9), which after heating with 2,3-dichloro-5,6-
dicyanobenzoquinone (DDQ) in benzene gave a dehydro-
genated product, dehydro-4-methoxycyclobrassinin (10).12
1,3-Thiazino[6,5-b]indole skeleton was also prepared by
heating of 2-chloroindole-3-carboxaldehyde with thiourea
in ethanol.13 Derivatives of the types C and D have not been
described to date.
a close analog of natural product 8. Starting aldehydes
13a,16a 13e16c and 13f,16d afforded by oxidation with
KMnO4 the corresponding acids 14a,16a 14e16c and 14f in
good yields (Scheme 2). Acid chlorides 15a, 15e and 15f
were prepared by heating of acids with PCl3 in dry benzene.
It was important to keep the bath temperature between 85–
908C since at a lower temperature the reaction did not
proceed whereas over 908C decomposition had occurred.
Treatment of the crude products of highly instable acid
chlorides with KSCN in acetone afforded surprisingly stable
isothiocyanates 16a, 16e and 16f, showing in their IR
spectra a characteristic absorption bands of NvCvS group
at 1960 cm21. The isothiocyanates could be isolated by
flash chromatography as crystalline compounds, however
with significant loss, caused by decomposition on silica gel.
Therefore in the next reaction, crude isothiocyanates were
used. Reaction of 16a, 16e and 16f with methanol, ethanol,
2-propanol and piperidine afforded the key intermediates,
thiocarbamoyl compounds 17a–17f accompanied with side
products, carboxylic acid esters 18a–18c, 18e and 18f and
amide 18d (Scheme 2). The formation of 18a–18f can be
explained via the extrusion of the thiocyanate anion under
the reaction conditions, facilitated by the electron-donating
group, with the formation of a reactive ketene, which
immediately reacts with the nucleophilic reagent.
Analogous reactivity has been previously observed in the
reactions of indol-3-ylcarbonyl isothiocyanate with nucleo-
philic reagents.17 The formation of 18a–18f by direct
alcoholysis or aminolysis of 16a, 16e and 16f is less
probable, since the reaction of nucleophiles with 1-Boc-2-
chloroindole-3-ylcarbonylisothiocyanate (23) did not afford
analogous side products. Another possible way, alcoholysis
or aminolysis of 17a–17f can be excluded, since the
prolonged heating of 17a–17f in the presence of corre-
sponding alcohols or piperidine did not result in the
formation of corresponding esters or amide. Thiocarbamoyl
compounds 17a–17f were separated from the reaction
mixtures by simple crystallization from acetone/cyclo-
hexane.
2. Results and discussion
Within our continuing research in the field of the synthesis
of cruciferous phytoalexins and congeneric indoles,14 we
have recently focused our attention on investigation of the
first synthesis of the hitherto synthetically not available
phytoalexin cyclobrassinon (8) and its analogues as
interesting synthetic target.15 Our aim also included the
first examination of antitumor, antibacterial and antifungal
activity of a new group of thiazino[6,5-b]indol-4-one
derivatives. Retrosynthesis of cyclobrassinon (Scheme 1)
led us via thiocarbamate 11 to 2-chloroindole-3-carboxylic
acid (12), as a suitable starting compound.
After the preparation of key intermediates 17a–17f, the
possibility of their cyclization by intramolecular nucleo-
philic replacement of a chlorine atom in the 2-position of an
indole ring with sulfur, was studied. There are several
examples of 2 and 4-chloronicotionoyl thioureas with
activated carbon–chlorine bonds, where similar substitution
was achieved by heating with LiH in DMF.18 Many attempts
to cyclize thiocarbamoyl compounds 17a–17f under the
action of various bases (for example Et3N, NaH, MeONa)
and temperatures up to 1508C resulted in decomposition and
no cyclization took place. Another possibility for substi-
tution of the chlorine is the weakening of the carbon–
halogen bond by photochemical excitation. Despite several
examples of nucleophilic substitutions of chlorine with
The preparation of acid 12 was described by oxidation of
2-chloroindole-3-carboxaldehyde (21) with KMnO4 in
aqueous acetone in the presence of phosphate buffer16a or
NaClO2 in aqueous dioxane in the presence of 2-methyl-2-
butene.16b However, oxidation of 21 in our hands did not
afford acid 12 in a reasonable yield. It was decided to solve
the problem by introduction of protecting groups on the
indole nitrogen. For this purpose, electron-withdrawing acyl
groups are not suitable, since oxidation of the corresponding
aldehydes affords only low yields of the required acids.16a
Therefore benzyl and 4-methoxybenzyl groups were
selected and a methyl derivative was also investigated
with the aim to synthesize 9-methylcyclobrassinon (19a) as

P. Kutschy et al. / Tetrahedron 58 (2002) 9029–9039
9031
sulfur in the 2-position of the indole,13,19 the photochemical
approach has not been applied for this purpose.20 UV
spectra of compounds 17a–17f exhibit long wave absorp-
tion maxima at 308 nm, except for 17d (287 nm). Therefore,
their photolysis was realized by irradiation of a high-
pressure mercury discharge lamp through a Pyrex filter
(l.300 nm) under a nitrogen atmosphere. It was found that
the desired intramolecular photosubstitution of chlorine by
sulfur advantageously proceeds in methanolic solutions of
17a–17f (Scheme 2). Due to the decomposition of the final
products caused by liberated HCl, the addition of Et3N to
the reaction mixture was necessary. Since the thiazino-
indoles 19a–19f formed appeared to be photolabile, the
reaction time was optimised and the reaction was stopped,
before all the starting material was consumed. Analogues of
cyclobrassinon were isolated by column chromatography in
16–55% yield. The IR spectra of thiazinoindoles 19a–19f
lack the characteristic NH stretching frequency, present in
the starting thiocarbamoyl compounds in the region of
3400 cm21, while their characteristic CvO absorption
bands appeared at lower frequency by about 40 cm21 as a
result of conjugation with the endocyclic CvN bond. In
contrast to 13C NMR spectra of thiocarbamoyl compounds
17a–17f exhibiting signals of the thiocarbonyl group at
178–190 ppm, the photocyclization products 19a–19f
show the signals of CvN bond at 156–164 ppm. In the
mass spectra of thiazinoindoles 19a–19f their structure was
unambiguously confirmed by the presence of the expected
molecular ions and the base-peaks corresponding to retro
Diels–Alder splitting. In the next step we examined the
possibility of the removal of protecting Bn and 4-MeOC6-
H4CH2 groups from the indole nitrogen of thiazinoindoles
19e and 19f. The benzyl protecting group can be removed
from protected indoles by employing sodium in liquid
ammonia,21a AlCl3 in dry benzene,21b and by methyl lithium
or lithiumdiisopropylamide (LDA) in dry tetrahydro-
furan.21c The 4-methoxybenzyl protecting group can be
removed by DDQ in refluxing toluene and by trifluoroacetic
acid (TFA) in refluxing dichloromethane.22a Cerium–
ammonium nitrate (CAN) has been also applied for this
purpose.22b Except for the use of TFA, where dione 20
(Scheme 2) was formed by acidolysis of the 2-methoxy
group, the above methods produced intractable mixtures of
decomposition products.
Although a new photocyclization methodology for the
preparation of 1,3-thiazino[6,5-b]indol-4-one derivatives
substituted on the indole nitrogen with an electron-donating
substituent has been developed, it appeared not suitable for
the synthesis of cyclobrassinon (8). Since we did not
succeed with removal of the electron-donating protecting
groups, we turned our attention to the mere easily removed
electron-withdrawing groups. It is well known, that the tert-
butoxycarbonyl (Boc) group has a wide scope of application
for the protection of nitrogen23 and therefore we selected
1-Boc-2-chloroindole-3-carboxaldehyde (22) as a suitable
starting compound. As expected, its oxidation to 1-Boc-2-
chloroindole-3-carboxylic acid failed, as described for the
other 1-acyl-2-chloro-3-carboxaldehydes.16a The problem
was solved by application of radical bromination method-
ology (NBS, AIBN, tetrachloromethane, reflux),
previously used for the direct conversion of some aliphatic
and aromatic aldehydes to acid bromides.24 Whereas
unprotected aldehyde 21 completely decomposed under
these conditions, the radical bromination of 1-Boc-2-
chloroindole-3-carboxaldehyde (22) gave the corresponding
unstable acid bromide, which after treatment with KSCN
afforded stable 1-Boc-2-chloroindole-3-ylcarbonylisothio-
cyanate 23 (Scheme 3), showing in its IR spectrum the
characteristic absorption band of the NvCvS group at
Scheme 2. Reagents and conditions: (i) KMnO4, 14a,16a 14e,16c 14f: acetone/water, rt, 58%; (ii) PCl3, benzene, 85–908C, a: 35 min, e: 30 min, f: 1 h;
(iii) KSCN, acetone, rt, a: 20 min, 30%, e: 1 h, 40%, f: 1 h, 29%, the yields of isolated isothiocyanates are based on carboxylic acids 14; (iv) MeOH, 608C, 1 h,
26% (17a), 18% (18a); EtOH, 608C, 1 h, 23% (17b), 8% (18b); i-PrOH, 608C, 2 h, 21% (17c), 19% (18c); piperidine, acetone, rt, 20 min, 21% (17d), 20%
(18d); MeOH, 608C, 1 h, 42% (17e), 2% (18e); MeOH, 608C, 1 h, 40% (17f), 10% (18f), for 17 and 18 yields are based on carboxylic acids 14, using the crude
isothiocyanates 16; (v) hn, Pyrex filter, N2, Et3N, methanol, rt, a: 35 min, 33%, b: 75 min, 38%, c: 35 min, 55%, d: 6 h, 16%, e: 35 min, 47%, f: 35 min, 40%;
(vi) TFA, dichloromethane, reflux, 48 h, 29%.

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P. Kutschy et al. / Tetrahedron 58 (2002) 9029–9039
Scheme 3. Reagents and conditions: (i) Boc2O, DMAP, THF, 58C, 1 h, 68%; (ii) NBS, AIBN, tetrachloromethane, reflux, 10 min; (iii) KSCN, acetone, rt,
15 min, 41% (based on aldehyde 22); (iv) a: MeOH, acetone, rt, 2 h, b: EtOH, acetone, rt, 2.5 h, c: i-PrOH, acetone, reflux, 5 min, d: MeNH2, acetone, 08C,
10 min, 44%, e: 4-toluidine, acetone, 08C, 15 min f: piperidine, acetone, 08C, 5 min; (v) a: Et3N, acetone, rt, 1 h, 61%, b: Et3N, acetone, rt, 1.5 h, 64%, c: Et3N,
acetone, reflux, 1 h, 47%, d: Et3N, acetone, reflux, 3 h, 70%, e: Et3N, acetone, reflux, 75 min, 66%, f: Et3N, acetone, rt, 2 h, 67% (for 25a–25c, 25e and 25f
yields are based on isothiocyanate 23); (vi) a: 165–1708C, 40 min, 70%, b: 165–1708C, 30 min, 80%, c: 160–1658C, 30 min, 66%, d: 180–1858C, 20 min,
72%, e: 155–1608C, 25 min, 81%, f: 155–1608C, 25 min, 77%.
1967 cm21. Nucleophilic addition of methanol to isothio-
cyanate 23 afforded the corresponding monothiocarbamate
24a. Cyclization of 24a to 9-Boc-cyclobrassinon (25a)
proceeded smoothly in 61% yield (from isothiocyanate 23)
by nucleophilic substitution of the chlorine in the presence
of triethylamine, significantly facilitated by the activating
effect of the Boc group. It was found that monothiocarba-
mate 24a cannot be isolated in a pure state, since after its
formation and during isolation it undergoes spontaneous
partial cyclization to thiazinoindole 25a. This behaviour
can be explained by an unusually high reactivity of the
chlorine atom in the 2-position activated by the electron-
withdrawing effect of the carbonyl groups in the 1 and
3-positions. The best yield of 25a as a sole reaction product
was obtained when 24a was generated in situ and
cyclization completed by addition of triethylamine. The
last step in the synthesis of cyclobrassinon (8) was the
removal of the protecting group from 25a. The Boc group
can be removed thermally,25a,b under acidic25c or basic25d
conditions. We found that deprotection of 25a to 8 can be
effectively achieved in 70% yield by heating without
solvent to 165–1708C, whereas treatment with acidic or
basic reagents led to decomposition. The spectral data of
synthetic cyclobrassinon are in agreement with the data of
the natural product.9
The in vitro antimicrobial activity of the synthesized
compounds was tested against representative strains of
bacteria, yeasts and molds. The activity of most compounds
was very low or absent against bacteria (MIC$100
mg mL21) and fungi (MIC$200 mg mL21). Compouds
16a, 22 and 23, exhibited a good antifungal activity against
Cryptococcus neoformans (MIC¼75, 25 and 10 mg mL21
,
respectively). Compounds 17a and 24d exhibited activity
against Staphylococcus aureus (MIC¼50 mg mL21).
As a primary in vitro screening for growth inhibition and
cytotoxicity, selected compounds were submitted to NCI
(National Cancer Institute, Bethesda, Md) and evaluated for
their cytotoxic potency on three human cell lines, such as
NCI-H460 lung cancer, MCF7 breast cancer and SF-268
glioma. A compound is considered active when it reduces
the growth of any of the cell lines to 32% or less and it is
passed on for evaluation in the full panel of sixty cell lines.
Compounds 22, 25e, 25f, 26e and 26f were active in
this test. The panel of 60 human tumor cell lines is
organised into subpanels representing leukemia, melanoma
and cancers of lung, colon, kidney, ovary, breast, prostate
and central nervous system. The test compounds
were dissolved in DMSO and evaluated using five
concentrations at ten-fold dilutions, the highest being
1024 M and the others 1025–1028 M L21. They did not
show a level of activity sufficient to enter the subsequent in
vivo step.
After the successful synthesis of cyclobrassinon (8) we
studied the scope and limitations of our elaborated synthetic
sequence. Thus, isothiocyanate 23 was reacted with ethanol,
2-propanol, methylamine, piperidine and 4-toluidine
(Scheme 3). Thiocarbamates 24b and 24c and thiourea
derivatives 24d–24f cyclized smoothly to protected thia-
zinoindoles in 47–70% yield (based on isothiocyanate 23).
Except for 24d, thiocarbamoyl compounds 24b, 24c, 24e
and 24f appeared to be equally reactive as 24a and during
attempted isolation partially cyclized to the corresponding
thiazinoindoles. Analogously to 25a, the deprotection of
thiazinoindoles 25b–25f was advantageously performed
thermally in 70–80% yield. The structure of thiazino[6,5-
b]indol-4-one derivatives 25a–25f and 26b–26f was
confirmed by spectral data, fully corresponding with the
data for compounds 19a–19f.
3. Conclusion
The first total synthesis of indole phytoalexin cyclo-
brassinon (six steps, overall yield 12%) and some of its
analogues was achieved starting from 1-substituted
2-chloroindole-3-carboxaldehydes. The photochemical or
Et3N-mediated nucleophilic cyclization of indolyl thio-
carbamates and thioureas was employed as a key step.
Specific antifungal activity of compounds 16a, 22 and 23
against C. neoformans is very promising: in fact life-
threatening infections caused by this encapsulated fungal
pathogen have been increasing steadly over the past 10

P. Kutschy et al. / Tetrahedron 58 (2002) 9029–9039
9033
years because of the onset of AIDS and the expanded use of
immunosuppressive drugs. Compounds 17a and 24d
exhibited activity against S. aureus. It is noteworthy to
consider that this Gram-positive bacterium is one of the
major pathogens that causes hospital infections. It is
characterized by an extreme flexibility in adapting to
antibiotic pressure, acquiring resistance to practically all
antibiotics so far introduced in clinical practice. Therefore
there is large interest in every new molecule that can be used
in intensive antibacterial therapy. The results from the
antitumor tests will be taken into account for the design of
new potential antitumor agents.
Harom), 7.11 (2H, d, J¼8.6 Hz, Harom), 7.17–7.41 (2H, m,
Harom), 7.62 (1H, d, J¼7.0 Hz, Harom), 8.05 (1H, d,
J¼7.0 Hz, Harom).
4.1.2. 1-Substitued 2-chloroindole-3-ylcarbonylchlorides
15a, 15e and 15f. To a stirred suspension of carboxylic acid
(14a, 1.048 g, 5 mmol; 14e, 1.143 g, 4 mmol; 14f, 1.263 g,
4 mmol) in dry benzene (14a, 45 mL; 14e and 14f, 36 mL)
was added PCl3 (14a, 0.587 g, 0.44 mL, 5 mmol; 14e and
14f, 0.549 g, 0.35 mL, 4 mmol) and the reaction mixture
was stirred at 85–908C (bath) until all carboxylic acid has
dissolved (14a, 15 min; 14e, 10 min; 14f, 40 min) and
heating was continued for 20 min. The reaction mixture was
then cooled to room temperature, decanted from the
phosphorous acid, deposited on the flask walls, the flask
washed with dry benzene (10 mL) and the collected benzene
solution evaporated to dryness (bath temperature up to
408C). The chlorides 15a, 15e and 15f were obtained as
unstable orange oils, immediately used for the next
reaction.
4. Experimental
4.1. General
Melting points were determined on a Koffler hot-stage
apparatus and are uncorrected. Infrared spectra were
recorded on an IR-75 (Zeiss, Jena) spectrometer in the
1
region 400–4000 cm21. H and 13C NMR spectra were
4.1.3. 1-Substitued 2-chloroindole-3-ylcarbonyl isothio-
cyanates 16a, 16e and 16f. The freshly prepared acyl
chlorides 15a, 15e and 15f were dissolved in dry acetone
(16a, 20 mL; 16e and 16f, 10 mL) and added in one portion
to the solution of KSCN (16a, 0.49 g, 5 mmol; 16e and 16f,
0.389 g, 4 mmol) in dry acetone (30 mL). The reaction
mixture was stirred at room temperature for 20 min (16a) or
1 h (16e and 16f). After the evaporation of acetone the crude
isothiocyanates are pure enough for the next reaction,
however they can be isolated in a pure crystalline form as
follows: to the acetone solution of isothiocyanate, prepared
from 2.5 mmol of corresponding acid was added a small
amount of silica gel and after the evaporation of acetone the
preadsorbed isothiocyanate was chromatographed on 50 g
of SiO2, cyclohexane/acetone (3:1, 16a), dichloromethane/-
hexane (3:1, 16e and 16f). 1-Methyl-2-chloroindole-3-
ylcarbonyl isothiocyanate (16a). Yield 30%, colourless
needles, mp 135–1378C (acetone/cyclohexane); (Found: C,
52.5; H, 3.0; N, 11.0. C11H7ClN2OS requires C, 52.70; H,
2.81; N, 11.17%); nmax (CHCl3) 1670 (CvO), 1970 cm21
(NvCvS); dH (300 MHz, CDCl3) 3.80 (3H, s, CH3), 7.27–
7.52 (3H, m, Harom), 8.12–8.37 (1H, m, Harom). 1-Benzyl-2-
chloroindole-3-ylcarbonyl isothiocyanate (16e). Yield
40%, colourless needles, mp 106–1088C (dichloro-
methane/hexane); (Found: C, 62.7; H, 3.5; N, 8.3. C17H11-
ClN2OS requires C, 62.48; H, 3.39; N, 8.57%); nmax(CHCl3)
1667 (CvO), 1960 cm21 (NvCvS); dH (300 MHz,
CDCl3) 5.47 (2H, s, CH2), 6.95–7.08 (8H, m, Harom),
8.05–8.42 (1H, m, Harom). 1-(4-Methoxybenzyl)-2-chloro-
indole-3-ylcarbonyl isothiocyanate (16f). Yield 29%,
colourless needles, mp 91–928C (dichloromethane/hexane);
(Found: C, 60.4; H, 3.9; N, 7.8. C18H13ClN2O2S requires C,
60.59; H, 3.67; N, 7.85%); nmax (CHCl3) 1660 (CvO),
1960 cm21 (NvCvS); dH (300 MHz, CDCl3) 3.77 (3H, s,
CH3), 5.42 (2H, s, CH2), 6.84 (2H, d, J¼8.7 Hz, Harom), 7.08
(2H, d, J¼8.7 Hz, Harom), 7.30–7.35 (3H, m, Harom), 8.24–
8.27 (1H, m, Harom); dC (75 MHz, CDCl3) 42.7 (CH3), 50.8
(CH2), 101.4 (C), 106.0 (CH), 109.9 (CH), 116.8 (CH),
119.2 (CH), 119.9 (CH), 121.4 (C), 122.2 (C), 123.5 (CH),
130.1 (C), 130.9 (C), 141.2 (C), 150.0 (CvO), 155.0
(NvCvS); EIMS: m/z (%): 356 (Mþ, 5), 298 (6), 177 (6),
121 (100), 114 (7), 77 (7).
measured on Varian Gemini 2000 (300 MHz for 1H,
75 MHz for 13C). Chemical shifts (d) are reported in ppm
downfield from TMS, the coupling constants (J) are given in
Hz. The mass spectra were recorded on a SSQ-700
spectrometer (Finigan) at an ionization energy 70 eV. The
ultraviolet spectra were recorded on a Specord M42
spectrometer in methanol. Microanalyses were performed
with a Perkin–Elmer, Model 2400 analyser. The reaction
course was monitored by TLC on Silufol plates (Kavalierw,
Czech Republic). The preparative column chromatography
(flash chromatography) was performed over the Kieselgel
Merck Typ 9385, 230–400 mesh. The in vitro antibacterial
activity of synthesized compounds was investigated against
representative Gram positive and Gram negative bacteria,
such as Escherichia coli ATCC 11105, Proteus mirabilis
M81, Streptococcus faecalis SF 8043 and S. aureus 6538/P.
Minimal inhibitory concentrations (MIC mg mL21) were
determined by the 2-fold agar dilution method,26 culturing
all bacteria on Mueller-Hinton agar (Difco, Detroit, MI,
USA). The in vitro antimycotic activity was tested against
Candida albicans L16, Candida lypolitica L6, C. neo-
formans L35 and Aspergillus niger NRLL330 on Sabauroad
dextrose agar (Merck) by the 2-fold agar dilution method.
All experiments were performed in triplicate, using inocula
of 106 cfu (colony forming units)/mL of bacteria and yeasts
and 105 conidia mL21 of molds. Fungal and bacteria plates
were incubated at 308C for 48 h and 378C for 24 h,
respectively. All compounds were dissolved in DMSO:
the final concentration of DMSO in tested media never
exceeded 1%. This concentration produced no visible
inhibition of growth in the controls. The cytotoxic activity
was evaluated according to the NCI protocols.27
4.1.1. 1-(4-Methoxybenzyl)-2-chloroindole-3-carboxylic
acid (14f). The acid 14f was prepared following the
procedure
for
1-phenyl-2-chloroindole-3-carboxylic
acid.28 Yield 58%, white solid, mp 215–2198C (decomp.,
ethanol); (Found: C, 64.4; H, 4.7; N, 4.2. C17H14NO3Cl
requires C, 64.66; H, 4.47; N, 4.44%); nmax (CHCl3) 1664
(CvO), 2200–3300 cm21 (OH); dH (300 MHz, DMSO-d6)
3.69 (3H, s, CH3), 5.52 (2H, s, CH2), 6.87 (2H, d, J¼8.6 Hz,

9034
P. Kutschy et al. / Tetrahedron 58 (2002) 9029–9039
4.1.4. Reaction of isothiocyanates 16a, 16e and 16f with
methanol. To the crude isothiocyanate 16a, 16e and 16f,
freshly prepared from 5 mmol of corresponding carboxylic
acid was added dry methanol (50 mL) and the mixture was
stirred for 1 h at 608C (bath). After the evaporation of
methanol, the residue was dissolved in ethyl acetate
(100 mL) and washed with 4% solution of sodium
hydrogencarbonate (16a, 2£50 mL) or 5% solution of
pottasium carbonate (16e and 16f; 1£80 mL, 1£60 mL,
1£40 mL) and then with water (60 mL). The organic layer
was dried (Na2SO4), solvent evaporated and the residue
crystallized from acetone/cyclohexane (17a, rt, 30 min; 17e
and 17f, 38C, 12 h). The precipitated thiocarbamates
were filtered off, washed with cyclohexane and dried. The
mother liquor was evaporated and residue chromatographed
on 30 g SiO2, cyclohexane/acetone (3:1, 17a), 20 g SiO2,
cyclohexane/acetone (3:1, 17e) or 100 g SiO2,
cyclohexane/acetone (5:1, 17f), affording a small amount
of thiocarbamates 17a, 17e and 17f (4–10%) and methyl
carboxylates 18a, 18e and 18f. O-Methyl N-(1-methyl-2-
chloroindole-3-ylcarbonyl)thiocarbamate (17a). Yield
26%, colourless needles, mp 127–1298C (acetone/cyclo-
hexane); (Found: C, 51.1; H, 4.2; N, 9.8. C12H11ClN2O2S
requires C, 50.98; H, 3.92; N, 9.91%); nmax (CHCl3) 1690
(CvO), 3413 cm21 (NH); dH (300 MHz, CDCl3) 3.82 (3H,
s, CH3), 4.24 (3H, s, CH3), 7.30–7.34 (3H, m, Harom), 8.25–
8.26 (1H, m, Harom), 9.60 (1H, s, NH); dC (75 MHz, CDCl3)
30.5 (CH3), 59.4 (CH3), 105.6 (C), 109.6 (CH), 121.4 (CH),
123.2 (CH), 124.0 (CH), 126.0 (C), 128.3 (C), 135.6 (C),
157.7 (CvO), 190.4 (CvS); UV, lmax (1max): 308 (1850);
267 (1980); 217 (3520). 1-Methyl-2-chloroindole-3-methyl-
carboxylate (18a). Yield 18%, colourless needles, mp 111–
1138C (methanol); (Found: C, 59.3; H, 4.6; N, 6.1.
C11H10ClNO2 requires C, 59.01; H, 4.51; N, 6.26%); nmax
(CHCl3) 1687 cm21 (CvO); dH (300 MHz, CDCl3) 3.78
(3H, s, CH3), 3.95 (3H, s, CH3), 7.12–7.43 (3H, m, Harom),
7.97–8.27 (1H, m, Harom). O-Methyl N-(1-benzyl-2-chloro-
indole-3-ylcarbonyl)thiocarbamate (17e). Yield 42%,
colourless needles, mp 127–1298C (acetone/cyclohexane);
(Found: C, 60.5; H, 4.0; N, 7.6. C18H15ClN2O2S requires C,
60.25; H, 4.21; N, 7.81%); nmax (CHCl3) 1687 (CvO),
3400 cm21 (NH); dH (300 MHz, CDCl3) 4.23 (3H, s, CH3),
5.45 (2H, s, CH2), 7.08–7.10 (2H, m, Harom), 7.28–7.33
(6H, m, Harom), 8.27–8.31 (1H, m, Harom), 9.61 (1H, s, NH);
dC (75 MHz, CDCl3) 47.6 (CH2), 59.4 (CH3), 106.2 (C),
110.3 (CH), 121.5 (CH), 121.7 (C), 123.4 (CH), 124.0 (C),
124.3 (CH), 126.2 (C), 126.5 (CH), 128.0 (C), 128.2 (CH),
129.1 (CH), 135.1 (CH), 135.5 (C), 157.8 (CvO), 190.3
(CvS); EIMS: m/z (%): 358 (Mþ, 5), 323 (22), 268 (80),
177 (11), 114 (12), 91 (100), 65 (12); UV, lmax (1max): 308
(2100); 268 (2400); 209 (5700). 1-Benzyl-2-chloroindole-3-
methylcarboxylate (18e). Yield 2%, white solid, mp 88–
908C (hexane); (Found: C, 68.4; H, 4.9; N, 4.6.
C17H14ClNO2 requires C, 68.12; H, 4.71; N, 4.67%); nmax
(CHCl3) 1687 cm21 (CvO); dH (300 MHz, CDCl3) 4.00
(3H, s, CH3), 5.48 (2H, s, CH2), 6.90–7.75 (8H, m, Harom),
8.02–8.35 (1H, m, Harom). O-Methyl N-[1-(4-methoxy-
benzyl)-2-chloroindole-3-ylcarbonyl]thiocarbamate (17f).
Yield 40%, colourless needles, mp 120–1228C (acetone/
cyclohexane); (Found: C, 58.8; H, 4.3; N, 7.6. C18H17ClN2-
O3S requires C, 58.68; H, 4.41; N, 7.20%); nmax (CHCl3)
1687 (CvO), 3400 cm21 (NH); dH (300 MHz, CDCl3) 3.76
(3H, s, CH3), 4.23 (3H, s, CH3), 5.38 (2H, s, CH2), 6.83 (2H,
d, J¼8.6 Hz, Harom), 7.05 (2H, d, J¼8.6 Hz, Harom), 7.26–
7.32 (3H, m, Harom), 8.26–8.29 (1H, m, Harom), 9.61 (1H, s,
NH); dC (75 MHz, CDCl3) 47.1 (CH2), 55.3 (CH3), 59.4
(CH3), 106.1 (C), 110.3 (CH), 114.5 (CH), 121.5 (CH),
123.3 (CH), 124.3 (CH), 126.2 (C), 127.1 (C), 128.0 (CH),
135.4 (CH), 157.8 (CvO), 159.5 (C), 190.3 (CvS); UV,
lmax (1max): 308 (1440); 270 (1570); 209 (5400). 1-(4-
Metoxybenzyl)-2-chloroindole-3-methylcarboxylate (18f).
Yield 10%, white solid, mp 88–918C (hexane); (Found:
C, 65.3; H, 5.0; N, 4.5. C18H16ClNO3 requires C, 65.56; H,
4.89; N, 4.25%); nmax (CHCl3) 1687 cm21 (CvO); dH
(300 MHz, CDCl3) 3.77 (3H, s, CH3), 3.98 (3H, s, CH3),
5.42 (2H, s, CH2), 6.80 (2H, d, J¼8.4 Hz, Harom), 7.08 (2H,
d, J¼8.4 Hz, Harom), 7.18–7.40 (3H, m, Harom), 8.02–8.35
(1H, m, Harom).
4.1.5. Reaction of isothiocyanate 16a with ethanol. To the
isothiocyanate 16a, freshly prepared from 7.5 mmol of
carboxylic acid 16a was added dry ethanol (70 mL) and the
mixture was stirred for 1 h at 608C (bath). After evaporation
of ethanol, the residue was dissolved in ethyl acetate
(130 mL) and washed with 4% solution of sodium
hydrogencarbonate (2£70 mL) and water (60 mL). The
organic layer was dried (Na2SO4), solvent evaporated and
the residue crystallized from acetone/cyclohexane to afford
17b. The mother liquor was evaporated and the residue
crystallized from methanol to afford methyl carboxylate
18b.
O-Ethyl
N-(1-methyl-2-chloroindole-3-ylcarbo-
nyl)thiocarbamate (17b). Yield 23%, colourless needles,
mp 131–1338C (acetone/cyclohexane); (Found: C, 52.8; H,
4.3; N, 9.6. C13H13ClN2O2S requires C, 52.61; H, 4.42; N,
9.44%); nmax (CHCl3) 1686 (CvO), 3410 cm21 (NH); dH
(300 MHz, CDCl3) 1.48 (3H, t, J¼7.0 Hz, CH3), 3.78 (3H,
s, CH3), 4.71 (2H, q, J¼7.0 Hz, CH2), 7.13–7.50 (3H, m,
Harom), 8.05–8.30 (1H, m, Harom), 9.60 (1H, s, NH); UV,
lmax (1max): 308 (933); 274 (1122); 215 (2754). 1-Methyl-2-
chloroindole-3-ethylcarboxylate (18b). Yield 8%, white
solid, mp 79–808C (methanol); (Found: C, 60.5; H, 5.2; N,
6.0. C12H12ClNO2 requires C, 60.64; H, 5.09; N, 5.89%);
nmax (CHCl3) 1683 cm21 (CvO); dH (300 MHz, CDCl3)
1.45 (3H, t, J¼7.0 Hz, CH3), 3.76 (3H, s, CH3), 4.43 (2H, q,
J¼7.0 Hz, CH2), 7.07–7.30 (3H, m, Harom), 7.95–8.27 (1H,
m, Harom).
4.1.6. Reaction of isothiocyanate 16a with 2-propanol. To
the isothiocyanate 16a, freshly prepared from 5 mmol of
carboxylic acid 16a was added dry 2-propanol (50 mL) and
the mixture was stirred for 2 h at 608C (bath). After the
evaporation of 2-propanol, the residue was dissolved in
ethyl acetate (100 mL) and washed with 4% solution of
sodium hydrogencarbonate (2£50 mL) and water (60 mL).
The organic layer was dried (Na2SO4), solvent evaporated
and the residue chromatographed on 70 g SiO2, cyclo-
hexane/acetone (3:1), affording thiocarbamate 17c and
2-propyl carboxylate 18c. O-2-Propyl N-(1-methyl-2-
chloroindole-3-ylcarbonyl)thiocarbamate (17c). Yield
21%, colourless needles, mp 112–1148C (acetone/cyclo-
hexane); (Found: C, 53.9; H, 4.7; N, 9.2. C14H15ClN2O2S
requires C, 54.10; H, 4.86; N, 9.01%); nmax (CHCl3) 1687
(CvO), 3417 cm21 (NH); dH (300 MHz, CDCl3) 1.46 (6H,
d, J¼6.0 Hz, 2£CH3), 3.80 (3H, s, CH3), 5.70 (1H, septet,
J¼6.0 Hz, CH), 7.16–7.50 (3H, m, Harom), 8.10–8.40 (1H,
m, Harom), 9.54 (1H, s, NH); UV, lmax (1max): 308 (2188);

P. Kutschy et al. / Tetrahedron 58 (2002) 9029–9039
9035
272 (2455); 213 (5248). 1-Methyl-2-chloroindole-3-(2-
propyl)carboxylate (18c). Yield 19%, white solid, mp 57–
598C (hexane); (Found: C, 62.2; H, 5.5; N, 5.7.
C13H14ClNO2 requires C, 62.03; H, 5.61; N, 5.56%); nmax
(CHCl3) 1683 cm21 (CvO); dH (300 MHz, CDCl3) 1.43
(6H, d, J¼6.0 Hz, 2£CH3), 3.77 (3H, s, CH3), 5.35 (1H,
septet, J¼6.0 Hz, CH), 7.15–7.45 (3H, m, Harom), 8.00–
8.35 (1H, m, Harom).
(Found: C, 58.4; H, 3.9; N, 11.5. C12H10N2O2S requires C,
58.52; H, 4.09; N, 11.37%); nmax (CHCl3) 1580 and
1650 cm21 (CvN–CvO); dH (300 MHz, CDCl3) 3.69
(3H, s, CH3), 4.17 (3H, s, CH3), 7.13–7.50 (3H, m, Harom),
8.25–8.55 (1H, m, Harom); dC (75 MHz, CDCl3) 30.7 (CH3),
57.7 (CH3), 102.7 (C), 108.6 (CH), 121.8 (CH), 122.5 (CH),
124.1 (CH), 125.1 (C), 138.3 (C), 139.5 (C), 164.5 and
166.4 (CvN–CvO); EIMS: m/z (%): 246 (Mþ, 61), 189
(100), 160 (15), 117 (25), 89 (10). 2-Ethoxy-9-methyl-1,3-
thiazino[6,5-b]indol-4-one (19b). Yield 38%, colourless
needles, mp 198–2018C (acetone/cyclohexane); (Found: C,
60.1; H, 4.5; N, 10.6. C13H12N2O2S requires C, 59.98; H,
4.65; N, 10.76%); nmax (CHCl3) 1570 and 1640 cm21
(CvN–CvO); dH (300 MHz, CDCl3) 1.45 (3H, t,
J¼7.0 Hz, CH3), 3.65 (3H, s, CH3), 4.68 (2H, q,
J¼7.0 Hz, CH2), 7.13–7.55 (3H, m, Harom), 8.25–8.63
(1H, m, Harom); EIMS: m/z (%): 260 (Mþ, 61), 189 (100),
160 (20), 117 (44), 89 (15). 2-(2-Propoxy)-9-methyl-1,3-
thiazino[6,5-b]indol-4-one (19c). Yield 55%, colourless
needles, mp 193–1968C (acetone/cyclohexane); (Found: C,
61.0; H, 5.3; N, 10.5. C14H14N2O2S requires C, 61.29; H,
5.14; N, 10.21%); nmax (CHCl3) 1570 and 1640 cm21
(CvN–CvO); dH (300 MHz, CDCl3) 1.42 (6H, d,
J¼6.1 Hz, 2£CH3), 3.69 (3H, s, CH3), 5.75 (1H, septet,
J¼6.1 Hz, CH), 7.16–7.58 (3H, m, Harom), 8.28–8.62 (1H,
m, Harom); EIMS: m/z (%): 274 (Mþ, 100), 233 (20), 189
(66), 160 (26), 128 (10), 117 (61), 89 (15), 43 (21). 2-(1-
Piperidyl)-9-methyl-1,3-thiazino[6,5-b]indol-4-one (19d).
Yield 16%, colourless needles, mp 227–2298C (acetone/
cyclohexane); (Found: C, 64.4; H, 5.4; N, 13.2.
C16H17N3OS requires C, 64.19; H, 5.72; N, 14.04%); nmax
(CHCl3) 1540 and 1607 cm21 (CvN–CvO); dH
(300 MHz, CDCl3) 1.60–1.85 (6H, m, 3£CH2), 3.65 (3H,
s, CH3), 3.72–3.90 (4H, m, 2£CH2), 7.13–7.48 (3H, m,
Harom), 8.30–8.63 (1H, m, Harom); dC (75 MHz, CDCl3)
24.5 (CH2), 25.7 (CH2), 30.3 (CH3), 47.9 (CH2), 101.6 (C),
108.3 (CH), 121.9 (CH), 122.0 (CH), 123.2 (CH), 125.3 (C),
137.8 (C), 138.0 (C), 156.3 and 167.7 (CvN–CvO);
EIMS: m/z (%): 299 (Mþ, 28), 189 (100), 160 (15), 117 (32),
89 (10). 2-Methoxy-9-benzyl-1,3-thiazino[6,5-b]indol-4-
one (19e). Yield 37%, colourless needles, mp 187–1898C
(dichloromethane/hexane); (Found: C, 67.3; H, 4.6; N, 8.9.
C18H14N2O2S requires C, 67.06; H, 4.38; N, 8.69%); nmax
(CHCl3) 1567 and 1640 cm21 (CvN–CvO); dH
(300 MHz, CDCl3) 4.17 3H, (s, CH3), 5.35 (2H, s, CH2),
7.12–7.14 (2H, m, Harom), 7.31–7.38 (6H, m, Harom), 8.49–
8.52 (1H, m, Harom); dC (75 MHz, CDCl3) 48.4 (CH2), 57.8
(CH3), 103.2 (C), 109.2 (CH), 122.1 (CH), 122.8 (CH),
124.4 (CH), 125.4 (C), 126.8 (CH), 128.6 (CH), 129.2 (CH),
134.4 (C), 138.2 (C), 139.2 (C), 164.8 and 166.6 (CvN–
CvO); EIMS: m/z (%): 322 (Mþ, 36), 265 (100), 236 (61),
204 (5), 91 (5). 2-Methoxy-9-(4-methoxybenzyl)-1,3-thia-
zino[6,5-b]indol-4-one (19f). Yield 48%, colourless
needles, mp 134–1378C (dichloromethane/hexane);
(Found: C, 64.5; H, 4.7; N, 7.7. C19H16N2O3S requires C,
64.76; H, 4.58; N, 7.96%); nmax (CHCl3) 1572 and
1642 cm21 (CvN–CvO); dH (300 MHz, CDCl3) 3.76
(3H, s, CH3), 4.15 (3H, s, CH3), 5.26 (2H, s, CH2), 6.83 (2H,
d, J¼9.0 Hz, Harom), 7.06 (2H, d, J¼9.0 Hz, Harom), 7.33–
7.37 (3H, m, Harom), 8.47–8.50 (1H, m, Harom); dC
(75 MHz, CDCl3) 48.2 (CH2), 55.5 (CH3), 57.9 (CH3),
103.6 (C), 109.4 (CH), 114.8 (C), 122.2 (CH), 122.9 (CH),
124.5 (C), 125.6 (CH), 126.4 (CH), 128.6 (C), 138.3 (C),
4.1.7. Reaction of isothiocyanate 16a with piperidine. To
an acetone solution of isothiocyanate 16a, filtered with
charcoal, freshly prepared from 5 mmol of carboxylic acid
16a, piperidine (0.639 g, 0.74 mL, 7.5 mmol) was added
with stirring. The reaction mixture was stirred for 20 min at
room temperature. Acetone was evaporated, the residue
dissolved in ethyl acetate (100 mL), washed with 4%
solution of sodium hydrogencarbonate (2£50 mL) and
water (60 mL). The water layer was extracted with
ethylacetate (30 mL), combined extracts dried (Na2SO4),
solvent evaporated and the residue crystallized from
acetone/cyclohexane (08C, 30 min) to afford thiourea 17d.
The mother liquor was evaporated and the residue
chromatographed on 80 g SiO2, benzene/acetone (7:1),
affording a small amount of thiourea 17d and the carboxylic
acid amide 18d. 1-[N-(1-Methyl-2-chloroindole-3-ylcar-
bonyl)thiocarbamoyl]piperidine (17d). Yield 21%, colour-
less needles, mp 147–1498C (acetone/cyclohexane);
(Found: C, 57.1; H, 5.3; N, 12.4. C16H18ClN3OS requires
C, 57.22; H, 5.40; N, 12.51%); nmax (CHCl3) 1670 (CvO),
3400 cm21 (NH); dH (300 MHz, CDCl3) 1.60–1.92 (6H, m,
3£CH2), 3.65–3.92 (2H, m, CH2), 3.81 (3H, s, CH3), 4.07–
4.32 (2H, m, CH2), 7.15–7.50 (3H, m, Harom), 8.05–8.28
(1H, m, Harom), 8.56 (1H, s, NH); dC (75 MHz, CDCl3) 24.0
(CH2), 25.7 (CH2), 30.4 (CH3), 49.9 (CH2), 105.6 (C), 109.7
(CH), 120.9 (CH), 122.9 (CH), 123.8 (CH), 125.7 (C), 129.0
(C), 135.6 (C), 159.0 (CvO), 178.4 (CvS); UV, lmax
(1max): 289 (2188); 241 (2089); 214 (3631). 1-Methyl-2-
chloroindole-3-ylcarbonylpiperidine (18d). Yield 20%,
white solid, mp 117–1198C (dichloromethane/hexane);
(Found: C, 65.3; H, 6.0; N, 10.4. C15H17ClN2O requires
C, 65.09; H, 6.19; N, 10.12%); nmax (CHCl3) 1609 cm21
(CvO); dH (300 MHz, CDCl3) 1.65 (6H, m, 3£CH2), 3.59
4H, (m, 2£CH2), 3.72 (3H, s, CH3), 7.00–7.40 (3H, m,
Harom), 7.43–7.75 (1H, m, Harom).
4.1.8. Photochemical cyclization of thiocarbamoyl com-
pounds 17a–17f. To a stirred solution of thiocarbamoyl
compounds 17a–17f (1 mmol) in methanol (200 mL) was
added triethylamine (0.202 g, 0.28 mL, 2 mmol) and the
mixture was irradiated (high pressure mercury lamp, Tesla
RVK-125) through a pyrex filter in a water-cooled
immersed apparature equipped with a quarz-jacketed
discharge lamp, cooled with water for 35 min (17a, 17c
and 17e), 75 min (17b), 1 h (17f) or 6 h29 (17d). Nitrogen
gas was bubbled through a solution 15 min before
irradiation and during the reaction. After evaporation of
methanol the residue was dissolved in chloroform (5 mL)
and chromatographed on 50 g SiO2 (19a–19c, 19e and 19f,
benzene/acetone (7:1); 19d, cyclohexane/acetone (2:1)).
The eluate was evaporated and thiazinoindoles 19a–19f
were crystallized from an appropriate solvent. 2-Methoxy-9-
methyl-1,3-thiazino[6,5-b]indol-4-one (19a). Yield 33%,
colourless needles, mp 202–2058C (acetone/cyclohexane);

9036
P. Kutschy et al. / Tetrahedron 58 (2002) 9029–9039
139.4 (C), 160.0 and 165.0 (CvN–CvO); EIMS: m/z (%):
352 (Mþ, 44), 295 (8), 266 (5), 236 (6), 148 (7), 121 (100),
91 (5), 77 (7).
7.39–7.42 (2H, m, Harom), 8.00–8.03 (1H, m, Harom), 8.22–
8.25 (1H, m, Harom); dC (75 MHz, CDCl3) 28.1 (CH3), 87.3
(C), 111.4 (C), 114.5 (CH), 121.1 (CH), 124.9 (CH), 125.6
(C), 125.9 (CH), 133.2 (C), 135.0 (C), 146.6 and 147.8
(CvO), 155.4 (NvCvS); EIMS: m/z (%): 336 (Mþ, 22),
278 (10), 236 (10), 178 (21), 114 (10), 57 (100), 41 (14).
4.1.9. Reaction of thiazinoindole 19f with trifluoroacetic
acid. To a solution of thiazinoindole 19f (0.135 g,
0.38 mmol) in dichloromethane (5 mL) was added trifluoro-
acetic acid (4.44 g, 3 mL, 38.3 mmol) and the reaction
mixture was refluxed for 48 h. The solvent was evaporated
and the residue chromatographed on 50 g SiO2, chloro-
form/acetonitrile (11:1). 9-(4-Methoxybenzyl)-1,3-thia-
zino[6,5-b]indol-2,4-dione (20). Yield 0.04 g (30%), white
solid, mp 228–2328C (methanol); (Found: C, 64.1; H, 4.3;
N, 9.5. C18H14N2O3S requires C, 63.89; H, 4.17; N, 8.29%);
nmax (CHCl3) 1670 (CvO), 3350 cm21 (NH); dH
(300 MHz, CDCl3) 3.78 (3H, s, CH3), 5.25 (2H, s, CH2),
6.86 (2H, d, J¼9.0 Hz, Harom), 7.12 (2H, d, J¼9.0 Hz,
Harom), 7.33–7.41 (3H, m, Harom), 8.27–8.30 (1H, m,
Harom), 8.38 (1H, s, NH); dC (75 MHz, CDCl3) 48.7 (CH2),
55.6 (CH3), 100.3 (C), 110.0 (CH), 114.9 (CH), 121.5 (CH),
123.6 (CH), 124.6 (CH), 125.7 (C), 126.2 (C), 128.7 (CH),
138.8 (C), 139.2 (C), 159.7 (C), 160.2 and 161.6 (CvO);
EIMS: m/z (%): 338 (Mþ, 18), 266 (5), 236 (5), 146 (5), 121
(100), 91 (5), 77 (7).
4.1.12. Conversion of isothiocayanate 23 to thiazino[6,5-
b]indol-4-one derivatives via thiocarbamates 24a–24c.
To a solution of isothiocyanate 23 (1 mmol) in dry acetone
(24a and 24b, 13 mL; 24c, 10 mL) was added methanol
(13 mL), ethanol (13 mL) or 2-propanol (10 mL) and
mixture was stirred at room temperature for 2 h (24a),
2.5 h (24b) or refluxed for 5 min (24c). Then triethylamine
(0.203 g, 0.28 mL, 2 mmol) was added and mixture was
stirred at room temperature for 1 h (25a), 1.5 h (25b) with
the formation of white precipitate or refluxed for 1 h (25c).
To a formed suspension of 25a, 25b was added water
(40 mL) and mixtures were set aside for 1 h at 38C. The
precipitated thiazinoindoles were filtered with suction, dried
and crystallized. In the case of 25c, the solvent was
evaporated and the residue chromatographed on 40 g
SiO2, benzene. 2-Methoxy-9-tert-butoxycarbonyl-1,3-thia-
zino[6,5-b]indol-4-one (25a). Yield 61%, colourless
needles, mp 212–2148C (dichloromethane/hexane);
(Found: C, 58.1; H, 4.9; N, 8.5. C16H16N2O4S requires C,
57.82; H, 4.85; N, 8.43%); nmax (CHCl3) 1580 and 1653
(CvN–CvO), 1727 cm21 (CvO); dH (300 MHz, CDCl3)
1.78 (9H, s, 3£CH3), 4.19 (3H, s, CH3), 7.41–7.44 (2H, m,
Harom), 8.07–8.10 (1H, m, Harom), 8.53–8.56 (1H, m,
Harom); dC (75 MHz, CDCl3) 28.2 (CH3), 57.5 (CH3), 87.9
(C), 108.5 (C), 114.6 (CH), 122.1 (CH), 124.8 (CH), 125.9
(CH), 126.9 (C), 136.0 (C), 139.0 (C), 149.8 (CvO), 167.4
and 169.7 (CvN–CvO); EIMS: m/z (%): 332 (Mþ, 22),
276 (81), 232 (10), 175 (41), 146 (18), 57 (100), 41 (22).
2-Ethoxy-9-tert-butoxycarbonyl-1,3-thiazino[6,5-b]indol-
4-one (25b). Yield 64%, colourless needles, mp 214–2168C
(dichloromethane/hexane); (Found: C, 59.1; H, 5.3; N, 8.4.
C17H18N2O4S requires C, 58.94; H, 5.24; N, 8.09%); nmax
(CHCl3) 1570 and 1647 (CvN–CvO), 1723 cm21
(CvO); dH (300 MHz, CDCl3) 1.46 (3H, t, J¼7.0 Hz,
CH3), 1.77 (9H, s, 3£CH3), 4.68 (2H, q, J¼7.0 Hz, CH2),
7.40–7.43 (2H, m, Harom), 8.07–8.10 (1H, m, Harom), 8.52–
8.55 (1H, m, Harom); dC (75 MHz, CDCl3) 14.2 (CH3), 28.2
(CH3), 67.0 (CH2), 87.8 (C), 108.4 (C), 114.6 (CH), 122.1
(CH), 124.8 (CH), 125.8 (CH), 126.9 (C), 136.0 (C), 139.1
(C), 149.8 (CvO), 167.6 and 169.1 (CvN–CvO); EIMS:
m/z (%): 346 (Mþ, 26), 290 (100), 246 (13), 172 (78), 141
(24), 51 (44). 2-(2-Propoxy)-9-tert-butoxycarbonyl-1,3-
thiazino[6,5-b]indol-4-one (25c). Yield 47%, colourless
needles, mp 206–2088C (dichloromethane/hexane);
(Found: C, 60.1; H, 5.7; N, 8.0. C18H20N2O4S requires C,
59.98; H, 5.59; N, 7.77%); nmax (CHCl3) 1567 and 1647
(CvN–CvO), 1727 cm21 (CvO); dH (300 MHz, CDCl3)
1.45 (6H, d, J¼6.1 Hz, 2£CH3), 1.78 (9H, s, 3£CH3), 5.75
(1H, septet, J¼6.1 Hz, CH), 7.41–7.44 (2H, m, Harom),
8.09–8.11 (1H, m, Harom), 8.53–8.56 (1H, m, Harom); dC
(75 MHz, CDCl3) 21.7 (CH3), 28.2 (CH3), 75.1 (CH), 87.7
(C), 108.3 (C), 114.6 (CH), 122.1 (CH), 124.8 (CH), 125.7
(CH), 127.0 (C), 136.0 (C), 139.3 (C), 149.7 (CvO), 167.7
and 168.7 (CvN–CvO); EIMS: m/z (%): 360 (Mþ, 9), 304
(27), 262 (7), 260 (5), 218 (9), 175 (69), 146 (29), 120 (13),
102 (6), 57 (100), 41 (62).
4.1.10. 1-(tert-Butoxycarbonyl)-2-chloroindole-3-car-
boxaldehyde (22). To a solution of aldehyde 21 (4.32 g,
24 mmol) in dry tetrahydrofuran (100 mL), was added in
one portion a solution of di-tert-butyl dicarbonate (6.288 g,
28.8 mmol) in dry tetrahydrofuran (20 mL) and the mixture
was cooled to 58C. Then DMAP (0.058 g, 0.5 mmol) was
added and the mixture was stirred for 1 h at 58C. The solvent
was evaporated, the residue dissolved in ethyl acetate
(150 mL) and consecutivelly washed with a 70 mL portions
of 5% solution of potassium hydroxide, 1 M solution of HCl
and brine. The organic layer was dried (Na2SO4) and solvent
evaporated. Yield 4.38 g (65%), white solid, mp 88–908C
(ethanol/water); (Found: C, 60.3; H, 5.0; N, 4.8.
C14H14ClNO3 requires C, 60.11; H, 5.04; N, 5.01%); nmax
(CHCl3) 1673 and 1760 cm21 (CvO); dH (300 MHz,
CDCl3) 1.75 (9H, s, 3£CH3), 7.37–7.41 (2H, m, Harom),
8.05–8.08 (1H, m, Harom), 8.29–8.32 (1H, m, Harom), 10.29
(1H, s, CHO); dC (75 MHz, CDCl3) 28.1 (CH3), 86.9 (C),
114.9 (CH), 117.5 (C), 121.1 (CH), 124.6 (C), 125.0 (CH),
125.9 (CH), 135.3 (C), 140.0 (C), 148.0 and 185.8 (CvO);
EIMS: m/z (%): 279 (Mþ, 22), 206 (11), 181 (14), 179 (45),
150 (28), 114 (12), 57 (100), 41 (28).
4.1.11. 1-(tert-Butoxycarbonyl)-2-chloroindole-3-ylcar-
bonyl isothiocyanate (23). To a solution of 22 (0.55 g,
2 mmol) in dry tetrachloromethane (3 mL) was added a
catalytic amount of AIBN and NBS (0.464 g, 2.6 mmol).
The mixture was stirred at reflux for 10 min, cooled to 38C,
separated precipitate filtered off and the filtrate treated with
a solution of KSCN (0.196 g, 2 mmol) in dry acetone
(10 mL). After stirring for 15 min at room temperature and
filtration with charcoal, the filtrate was evaporated and the
residue flash chromatographed on 10 g SiO2, benzene, to
afford 23. Yield 0.28 g (41%), colourless needles mp 131–
1328C (dichloromethane/hexane); (Found: C, 53.6; H, 4.0;
N, 8.5. C15H13ClN2O3S requires C, 53.49; H, 3.89; N,
8.32%); nmax (CHCl3) 1673 and 1755 (CvO), 1967 cm21
(NvCvS); dH (300 MHz, CDCl3) 1.74 (9H, s, 3£CH3),

P. Kutschy et al. / Tetrahedron 58 (2002) 9029–9039
9037
4.1.13. N0-Methyl N-(1-tert-butoxycarbonyl-2-chloro-
indole-3-ylcarbonyl)thiourea (24d). To a stirred solution
of isothiocyanate 23 (0.36 g, 1.07 mmol) in acetone
(25 mL) cooled to 08C was added 30% water solution of
methylamine (0.144 g, 0.16 mL, 2.67 mmol), the mixture
was stirred for 10 min at 08C, then poured into 150 mL of
water and set aside for 24 h at 38C. The separated precipitate
was filtered with suction, dried and crystallized. Yield
0.172 g (44%), white solid, mp 250–2528C (acetone/water);
(Found: C, 52.6; H, 5.0; N, 11.5. C16H18ClN3O3S requires
C, 52.24; H, 4.93; N, 11.42%); nmax (CHCl3) 1658 and 1753
(CvO), 3260 and 3403 cm21 (NH); dH (300 MHz, CDCl3)
1.73 (9H, s, 3£CH3), 3.28 (3H, d, J¼4.8 Hz, CH3), 7.29–
7.43 (2H, m, Harom), 8.04–8.13 (2H, m, Harom), 9.53 (1H, s,
NH), 10.68 (1H, s, NH); dC (75 MHz, CDCl3) 28.1 (CH3),
32.3 (CH3), 87.1 (C), 111.4 (C), 114.8 (CH), 120.8 (CH),
124.7 (CH), 125.7 (C), 126.0 (CH), 126.3 (C), 135.0 (C),
147.9 and 162.3 (CvO), 180.8 (CvS); EIMS: m/z (%): 367
(Mþ, 1), 332 (6), 276 (18), 232 (5), 177 (5), 114 (6), 77 (6),
57 (99), 41 (100).
Harom), 7.38–7.40 (2H, m, Harom), 8.04–8.07 (1H, m,
Harom), 8.34–8.37 (1H, m, Harom); dC (75 MHz, CDCl3)
21.1 (CH3), 28.0 (CH3), 88.1 (C), 107.1 (C), 114.8 (CH),
121.4 (CH), 122.6 (CH), 125.1 (CH), 125.6 (CH), 126.5 (C),
130.6 (CH), 135.8 (C), 136.1 (C), 137.4 (C), 139.7 (C),
149.3 (CvO), 153.1 and 160.6 (CvN–CvO); EIMS:
m/z (%): 407 (Mþ, 4), 351 (5), 175 (6), 57 (10), 56 (38),
41 (100). 2-(1-Piperidyl)-9-tert-butoxycarbonyl-1,3-thia-
zino[6,5-b]indol-4-one (25f). Yield 68%, white solid, mp
310–3128C (dichloromethane/hexane); (Found: C, 62.6; H,
6.2; N, 11.0. C20H23N3O3S requires C, 62.32; H, 6.01; N,
10.90%); nmax (CHCl3) 1555 and 1627 (CvN–CvO),
1720 cm21 (CvO); dH (300 MHz, CDCl3) 1.35–1.86 (6H,
m, 3£CH2), 1.76 (9H, s, 3£CH3), 3.77–3.98 (4H, m,
2£CH2), 7.34–7.41 2H, (m, Harom), 8.0–8.03 (1H, m,
Harom), 8.55–8.58 (1H, m, Harom); dC (75 MHz, CDCl3)
24.5 (CH2), 25.8 (CH2), 28.2 (CH3), 47.8 (CH2), 87.1 (C),
107.4 (C), 114.5 (CH), 122.5 (CH), 124.6 (CH), 125.2 (CH),
127.4 (C), 135.5 (C), 137.3 (C), 150.3 (CvO), 160.6 and
168.0 (CvN–CvO); EIMS: m/z (%): 385 (Mþ, 4), 341
(17), 285 (100), 172 (47), 141 (14), 50 (41), 41 (46).
4.1.14. Synthesis of thiazino[6,5-b]indol-4-one deriva-
tives 25d–25f. To a solution of isothiocyanate 23 (1 mmol)
in acetone (25 mL) cooled to 08C was added 4-toluidine
(24e, 0.107 g, 1 mmol) or piperidine (24f, 0.086 g, 0.1 mL,
1 mmol) and the mixture was stirred at 08C for 15 min (24e)
or 5 min (24f). Then triethylamine (0.203 g, 0.28 mL,
2 mmol) was added and stirring was continued for 75 min
at reflux (24e) or for 2 h at room temperature (24f). In the
case of thiourea 24d, to a solution of 24d (1 mmol) in
acetone (25 mL) was added triethylamine (0.203 g,
0.28 mL, 2 mmol) and refluxed with stirring for 3 h with
the formation of white precipitate. Thiazinoindoles 25d–
25f were isolated as follows: To the mixture containing
precipitated 25d was added water and mixture was set aside
for 30 min at 38C. The precipitate was filtered with suction,
dried and crystallized. The solution containing thiazino-
indole 25e was concentrated to a half of its original volume,
water (20 mL) was added and mixture set aside for 1 h at
38C, formed precipitate filtered with suction, dried and
crystallized. The mixture containing thiazinoindole 25f was
evaporated and the residue chromatographed on 30 g SiO2,
benzene/acetone (7:1). 2-Methylamino-9-tert-butoxycar-
bonyl-1,3-thiazino[6,5-b]indol-4-one (25d). Yield 70%,
white solid, mp 290–2928C (methanol/water); (Found: C,
58.1; H, 5.3; N, 12.5. C16H17N3O3S requires C, 57.99; H,
5.17; N, 12.68%); nmax (KBr) 1607 and 1627 (CvN–
CvO), 1703 cm21 (CvO); dH (300 MHz, DMSO-d6) 1.79
(9H, s, 3£CH3), 3.03 (3H, s, CH3), 7.20–7.58 (2H, m,
Harom), 8.39–8.18 (1H, m, Harom), 8.18–8.50 (1H, m,
Harom), 8.92 (1H, s, NH); dC (75 MHz, DMSO-d6) 27.8
(CH3), 29.5 (CH3), 87.4 (C), 106.3 (C), 114.8 (CH), 121.0
(CH), 124.4 (CH), 125.1 (CH), 127.1 (C), 135.1 (C), 137.5
(C), 149.4 (CvO), 161.1 and 167.2 (CvN–CvO); EIMS:
m/z (%): 331 (Mþ, 15), 275 (49), 231 (41), 172 (100), 141
(25), 113 (17), 50 (90), 41 (91). 2-(4-Tolylamino)-9-tert-
butoxycarbonyl-1,3-thiazino[6,5-b]indol-4-one (25e). Yield
66%, white solid, mp 280–2828C (acetone/water); (Found:
C, 65.1; H, 5.3; N, 10.5. C22H21N3O3S requires C, 64.85; H,
5.19; N, 10.31%); nmax(CHCl3) 1560 and 1620 (CvN–
CvO), 1727 (CvO), 3433 cm21 (NH); dH (300 MHz,
CDCl3) 1.66 (9H, s, 3£CH3), 2.43 (3H, s, CH3), 6.61 (1H, s,
NH), 7.06 (2H, d, J¼8.1 Hz, Harom), 7.25 (2H, d, J¼8.1 Hz,
4.1.15. Removal of the protecting Boc group. The
protected thiazinoindoles 25a–25f (0.5 mmol) were heated
without solvent as follows: (25a, 1708C, 40 min); (25b,
1708C, 30 min); (25c, 1608C, 30 min); (25d, 1858C,
20 min); (25e, 1608C, 25 min); (25f, 1558C, 25 min) and
after cooling to room temperature crystallized from an
appropriate solvent, except for thiazinoindole 26c, which
was preadsorbed on silica gel and chromatographed on 15 g
SiO2, benzene/acetone (7:1). 2-Methoxy-1,3-thiazino[6,5-
b]indol-4-one; Cyclobrassinon (8). Yield 70%, white solid,
mp 221–2238C (methanol); (Found: C, 57.1; H, 3.6; N,
12.2. C11H8N2O2S requires C, 56.88; H, 3.47; N, 12.06%);
nmax (KBr) 1567 and 1627 cm21 (CvN–CvO); dH
(300 MHz, DMSO-d6) 4.18 (3H, s, OCH3), 7.37–7.42
(2H, m, H-6, H-7), 7.66 (1H, d, J¼7.5 Hz, H-5), 8.27 (1Hd,
J¼7.5 Hz, H-8), 12.69 (1H, s, NH); dC (75 MHz, DMSO-d6)
57.6 (CH3), 102.2 (C), 111.9 (CH), 120.4 (CH), 122.0 (CH),
124.0 (CH), 124.3 (C), 136.9 (C), 137.5 (C), 165.5 and
165.7 (CvN–CvO); EIMS: m/z (%): 232 (Mþ, 45), 175
(100), 146 (23), 120 (33). 2-Ethoxy-1,3-thiazino[6,5-
b]indol-4-one (26b). Yield 81%, white solid, mp 223–
2258C (ethanol/water); (Found: C, 58.7; H, 4.3; N, 11.5.
C12H10N2O2S requires C, 58.52; H, 4.09; N, 11.37%); nmax
(KBr) 1567 and 1630 cm21 (CvN–CvO); dH (300 MHz,
DMSO-d6) 1.48 (3H, t, J¼7.0 Hz, CH3), 4.67 (2H, q,
J¼7.0 Hz, CH2), 7.37–7.43 (2H, m, Harom), 7.65 (1H, d,
J¼7.7 Hz, Harom), 8.27 (1H, d, J¼7.7 Hz, Harom), 12.68 (1H,
s, NH); dC (75 MHz, DMSO-d6) 14.1 (CH3), 67.0 (CH2),
102.2 (C), 111.9 (CH), 120.4 (CH), 122.0 (CH), 124.0 (CH),
124.3 (C), 136.8 (C), 137.6 (C), 164.8 and 165.8 (CvN–
CvO); EIMS: m/z (%): 246 (Mþ, 46), 172 (100), 141 (17),
113 (19). 2-(2-Propoxy)-1,3-thiazino[6,5-b]indol-4-one
(26c). Yield 66%, white solid, mp 208–2108C (acetone/
cyclohexane); (Found: C, 60.1; H, 4.7; N, 10.6.
C13H12N2O2S requires C, 59.98; H, 4.65; N, 10.76%);
n
max(KBr) 1554 and 1614 cm21 (CvN–CvO); dH
(300 MHz, DMSO-d6) 1.49 (6H, d, J¼6.2 Hz, 2£CH3),
5.62 (1H, septet, J¼6.2 Hz, CH), 7.36–7.42 (2H, m, Harom),
7.65 (1H, d, J¼7.5 Hz, Harom), 8.26 (1H, d, J¼7.5 Hz,
Harom), 12.68 (1H, s, NH); dC (75 MHz, DMSO-d6) 21.5
(CH3), 75.1 (CH), 102.1 (C), 111.9 (CH), 120.4 (CH), 122.0

9038
P. Kutschy et al. / Tetrahedron 58 (2002) 9029–9039
Action. Daniel, M., Purkayastha, R. P., Eds.; Marcel Dekker:
New York, 1995; pp 229–261. (b) Pedras, M. S. C.; Okanga,
F. I.; Zaharia, I. L.; Khan, A. Q. Phytochemistry 2000, 53,
161–176.
(CH), 124.0 (CH), 124.3 (C), 136.8 (C), 137.7 (C), 164.3
and 165.9 (CvN–CvO); EIMS: m/z (%): 260 (Mþ, 23),
175 (100), 146 (38), 120 (44), 103 (11), 43 (28).
2-Methylamino-1,3-thiazino[6,5-b]indol-4-one
(26d).
Yield 72%, white solid, mp 290–2928C (methanol);
(Found: C, 57.2; H, 4.0; N, 18.4. C11H9N3OS requires C,
57.13; H, 3.92; N, 18.17%); nmax (KBr) 1540 and
1607 cm21 (CvN–CvO); dH (300 MHz, DMSO-d6) 3.03
(3H, s, CH3), 7.26–7.35 (2H, m, Harom), 7.56 (1H, d,
J¼7.0 Hz, Harom), 8.23 (1H, d, J¼7.0 Hz, Harom), 8.74 (1H,
s, NH), 12.36 (1H, s, NH); dC (75 MHz, DMSO-d6) 29.6
(CH3), 101.3 (C), 111.4 (CH), 120.3 (CH), 121.4 (CH),
123.0 (CH), 124.8 (C), 136.1 (C), 136.4 (C), 157.7 and
167.0 (CvN–CvO); EIMS: m/z (%): 231 (Mþ, 58), 172
(100), 141 (22), 113 (27). 2-(4-Tolylamino)-1,3-thia-
zino[6,5-b]indol-4-one (26e). Yield 81%, white solid, mp
282–2848C (acetone/cyclohexane); (Found: C, 66.7; H, 4.4;
N, 13.5. C17H13N3OS requires C, 66.43; H, 4.26; N,
13.67%); nmax (KBr) 1547 and 1607 cm21 (CvN–
CvO); dH (300 MHz, DMSO-d6) 2.32 (3H, s, CH3), 7.22
(1H, s, NH), 7.21–7.28 (6H, m, Harom), 7. 47 (1H, d,
J¼8.3 Hz, Harom), 8.07 (1H, d, J¼8.3 Hz, Harom), 12.27 (1H,
s, NH); dC (75 MHz, DMSO-d6) 20.7 (CH3), 101.3 (C),
111.7 (CH), 120.0 (CH), 121.3 (CH), 121.9 (CH), 123.3
(CH), 124.9 (C), 130.0 (CH), 133.7 (C), 147.2 and 160.8
(CvN–CvO); EIMS: m/z (%): 307 (Mþ, 96), 176 (100),
146 (52), 120 (93), 103 (18), 91 (30), 77 (22). 2-(1-
Piperidyl)-1,3-thiazino[6,5-b]indol-4-one (26f). Yield 77%,
white solid, mp 320–3228C (methanol/water); (Found: C,
63.4; H, 5.4; N, 15.0. C15H15N3OS requires C, 63.13; H,
5.30; N, 14.73%); nmax (KBr) 1547 and 1600 cm21 (CvN–
CvO); dH (300 MHz, DMSO-d6) 1.68–1.77 (6H, m,
3£CH2), 3.79–3.91 (4H, m, 2£CH2), 7.26–7.27 (2H, m,
Harom), 7.58 (1H, d, J¼7.5 Hz, Harom), 8.24 (1H, d,
J¼7.5 Hz, Harom), 12.54 (1H, s, NH); dC (75 MHz,
DMSO-d6) 24.1 (CH2), 25.4 (CH2), 47.5 (CH2), 101.1 (C),
111.5 (CH), 120.5 (CH), 121.5 (CH), 123.3 (CH), 124.4 (C),
136.2 (C), 136.3 (C), 157.0 and 166.5 (CvN–CvO);
EIMS: m/z (%): 285 (Mþ, 56), 172 (100), 141 (22), 113 (27).
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Acknowledgements
Lett. 2001, 42, 9281–9283.
We thank the Grant Agency for Science, Slovak Republic
(Grant No. 1/9246/02) for financial support of this work. We
are grateful to the National Cancer Institute (Bethesda, MD)
for the antitumor tests.
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