Dependence of fluorescence properties of compounds from psoralen, angelicin, and carbazole series on the character of their terminal substituents

Article abstract of DOI:10.1023/B:RUJO.0000003169.96393.1d

Absorption, luminescence excitation and emission spectra of nine compounds from 4,8,4′-trimethylpsoralen, 4,4′-dimethylangelicin, and 1,2,3,4-tetrahydrocarbazole series with various terminal substituents were studied in water and 2-propanol. Proceeding from the data obtained in the present and also some previous studies we have formulated the main rules concerning a general mechanism of changes in the fluorescence characteristics of DNA-specific dyes depending on their chemical structure, substrate properties, and measurement medium.

Full text of DOI:10.1023/B:RUJO.0000003169.96393.1d

Russian Journal of Organic Chemistry, Vol. 39, No. 6, 2003, pp. 881 889. Translated from Zhurnal Organicheskoi Khimii, Vol. 39, No. 6, 2003,
pp. 930 938.
Original Russian Text Copyright
,
2003 by Sibirtsev, Tolmachev, Suslov, Garabadzhiu, Traven .
Dependence of Fluorescence Properties of Compounds
from Psoralen, Angelicin, and Carbazole Series
on the Character of Their Terminal Substituents
1
2
2
V. S. Sibirtsev , A. Yu. Tolmachev , V. V. Suslov ,
,
3
2
A. V. Garabadzhiu , and V. F. Traven
1
Mendeleev Central Research Institute of Metrology, St. Petersburg, 198005 Russia
2
Mendeleev Russian Chemical Technological University, Moscow, Russia
3
St. Petersburg State Technological Institute, St. Petersburg, Russia
Received June 27, 2002
Abstract Absorption, luminescence excitation and emission spectra of nine compounds from 4,8,4 -tri-
methylpsoralen, 4,4 -dimethylangelicin, and 1,2,3,4-tetrahydrocarbazole series with various terminal
substituents were studied in water and 2-propanol. Proceeding from the data obtained in the present and also
some previous studies we have formulated the main rules concerning a general mechanism of changes in the
fluorescence characteristics of DNA-specific dyes depending on their chemical structure, substrate
properties, and measurement medium.
Nowadays the application expands of artificially
produced relatively low-molecular compounds
capable of specific bind with a certain nucleotide
sequence in a genome. Such compounds may be used
as radioprotectors [1], antibacterial [2, 3], and anti-
tumor [4 6] pharmaceuticals. When alongside a good
affinity to a substrate and a high specificity the
DNA-binding compound possesses also convenient
measurable properties (for instance, fluorescence that
sharply changes at binding with a nucleotide) the
application range of the compound becomes still
wider. In particular, compounds with these properties
can be used for study of nucleic acids structure,
enzymatic kinetics [7 10]. Besides these compounds
can be applied to development of highly sensitive and
specific express-analyses for diagnostics of various
diseases and harmful influence based on pathology
analysis from the viewpoint of changes in content and
structure of nucleic acids during various physiological
states of a body [11, 12].
favorable. These molecules add to the polynucleotide
from outside, not affecting the substrate structure
(although therewith occurs some compression, and
also local changes in allosteric conformation of the
DNA double helix in the region of binding). Such
substances are mainly specific to the primary and
secondary organization orders of the nucleic acids
structure [9, 13 16].
At the same time the intercalator molecule must
contain one or several fragments of fused aromatic
systems with two or more (preferably three or four)
rings (preferably 5- or 6-membered), and at least one
heteroatom (preferably endocyclic). These compounds
build in between the complementary pairs of bases of
the polynucleotide double helix (effecting local un-
winding of the helix and its elongation), and they are
specific mainly to the secondary structure of nucleic
acids and to extent of their superspiralization [17 20].
Quite a number of DNA-fluorophores is now
known and widely applied [7, 9, 16 18]. However
the target of developing theoretical foundation for
building up new compounds with higher sensitivity
and specificity with respect to substrate is still urgent.
To this end it is necessary in its turn to have an exact
understanding of connection between the chemical
structure of the nucleotide-specific dyes and their
spectral and complexing characteristics.
All multitude of DNA-binding compounds known
nowadays is divided by the type of interaction with
the substrate in two large classes: intercalators and
so-called externally binding compounds. Therewith
the requirements for the structure of the latter class
compound are the least stringent: in its molecule
should be present at least a single heteroatom, and
besides in case the compound should possess measur-
able spectral properties the presence in the molecule
of an extended system of conjugated bonds is
In the present study were chosen as objects 9 new-
ly synthesized derivatives of 4,8,4 -trimethyl-
1
070-4280/03/3906-0881$25.00 2003 MAIK Nauka/Interperiodica

882
SIBIRTSEV et al.
psoralen (I III), 4,4 -dimethyangelicin (IV VI), and
,2,3,4-tetrahydrocarbazole (VII IX) with various
terminal substituents in positions 5 (for compounds
We selected these compounds for both psoralens
and angelicins were recently frequently used in
photochemotherapy, in particular in the case of
psoriasis [21, 22], for blood sterilization [3], for site-
directed RNA modification [23] etc. This wide ap-
1
I VI) and
9
(for compounds VII IX) [see
Scheme (1)].
(
1)
C(CH )=NOH (I, IV), C(O)CH (II, V), CH(CH )OCH (III), CH(CH )OH (VI), C N (VII), C(O)OH (VIII),
3
3
3
3
3
C(O)NHNH (IX).
2
plication range originates from the behavior of these
compounds: The compounds of both mentioned series
first reversibly intercalete into the double helix of
the polynucleotide (therewith as a rule psoralens
predominantly are bound with the AT-rich regions,
and angelicins are nonspecific to the primary struc-
ture of nucleic acids), and then after sufficiently
long (up to 1 h) irradiation in the wavelength range
with covalent bonds. Therewith the angelicins are
less toxic than psoralens for the latter are more prone
to cross-linking two strands of DNA; this is evidently
due to shorter angular angelicin core compared to
linear psoralen one.
A number of carbazole derivatives also exhibit
significant antimicrobial [24] and antitumor [25, 26]
activity.
2
50 370 nm are linked to substrate irreversibly
The spectral characteristics of compounds I IX
were studied in two media: solutions in water and
2
-propanol. As was described earlier [27, 28] in
the latter solvent because of its low permittivity
fluorescence was observed from the planar core
proper of compound excluding the influence thereto
of electrondonor or electron-acceptor effect of the
terminal substituents.
As seen from Figs. 1 3 in the spectra of all
compounds studied in this work the wavelengths in
the absorption maxima ( ab) coincide with the wave-
lengths of the fluorescence excitation maxima ( ex).
An exclusion is the absorption peak with a maximum
at
210 nm that does not appear in the fluorescence
ab
excitation spectra and evidently corresponds to elec-
tron transitions in benzene rings in the structure of
the compounds under investigation [29]. The other
observed absorption maxima in the spectra of com-
pounds from furocoumarin series I VI are located at
2
55 and 330 nm regardless of the measuring
conditions; in the spectra of compounds of tetra-
hydrocarbazole series VII IX the maxima in
water medium are 240 and 290 nm, and in 2-propanol
ab
ab
Fig. 1. Adsorption (a, d), fluorescence excitation (b, e),
and emission spectra (c, f) of 4,8,4 -trimethylpsoralens
2
55 and 355 nm respectively.
(
(
I III) in aqueous buffer solution (a c) and in 2-propanol
d f).
Wavelengths at the maxima of the fluorescence
emission spectra ( em) for compounds of the furo-
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 39 No. 6 2003

DEPENDENCE OF FLUORESCENCE PROPERTIES OF COMPOUNDS FROM PSORALEN
coumarin series I VI were 455 nm in both
883
em
solvents, and for compounds VII IX from the tetra-
hydrocarbazole series they were as follows: in water
medium
The comparison of values of fluorescence quantum
efficiency for compounds I IX in 2-propanol ( A
380 nm and in 2-propanol
360 nm.
em
em
)
and in water ( W) shows that in all cases the former
value is somewhat larger than the latter (1.2
4
.2 times) (see table). Therewith the
values for
A
related compounds I III, IV VI, and VII IX were
more alike than W, and decreased in the order from
tetrahydrocarbazole to trimethylpsoralen to dimethyl-
angelicin series.
Fluorescence quantum efficiency ( ) of compounds
a
under investigation
Compd. no.
A/
W
W
A
I
II
0.096
0.113
0.193
0.069
0.092
0.131
0.334
0.383
0.413
0.293
0.296
0.301
0.282
0.285
0.289
0.484
0.490
0.496
3.05
2.62
1.56
4.20
3.11
2.20
1.45
1.28
1.20
III
IV
V
VI
VII
VIII
IX
Fig. 2. Absorption (a, d), fluorescence excitation (b, e),
and emission spectra (c, f) of 4,4 -dimethylangelicins
IV VI) in aqueous buffer solution (a c) and in
-propanol (d f).
(
2
a
With indices (W) and (A) are marked the parameters
measured for compounds I IX in the aqueous buffer solution
and 2-propanol, respectively.
As seen from the table in the series of compounds
I III and IV VI is also observed decrease in the ratio
A/ evidently originating from reduction of the
W
electron-acceptor effect with respect to the furo-
coumarin core in the series of groups HO N = >
O = > HO > H C O . Obviously the hydroxyimino
3
group is the first in this series due to both strengthen-
ing of the nitrogen electron-acceptor properties by the
hydroxy group and by the possibility for this hydroxy
group to form a hydrogen bond with the oxygen of
the furan ring as shown in Scheme (2).
(
2)
Fig. 3. Absorption (a, d), fluorescence excitation (b, e),
and emission spectra (c, f) of 1,2,3,4-tetrahydrocarbazoles
(VII IX) in aqueous buffer solution (a c) and in
Oxo group attracts electron density from the furo-
coumarin core to greater extent than the hydroxy
2-propanol
(d f).
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 39 No. 6 2003

884
SIBIRTSEV et al.
group because the former is bonded to the core by a
double bond, and methoxy group is a weaker electron-
acceptor than hydroxy group for its oxygen can
attract electron density not only from the furo-
coumarin core but also from the methyl group.
(1) Active fluorescence can be produced only from
the structures with developed electronic conjugation
system involving -electrons of multiple (double and
triple) bonds or unshared n-electrons of heteroatoms.
(2) This conjugation is fulfilled only when this
A similar decrease in the value of A/
was also
electron system possesses sufficiently rigid coplanar
structure (that is, all fragments involved thereto are
located in the same plane).
W
observed in the series of compounds VII IX. It was
caused evidently by reduced electron-acceptor
character toward the tetrahydrocarbazole core of
carboxy group compared to nitrile one and of
hydrazinocarbonyl group compared to carboxyl.
However the range of changes was considerably
smaller in compounds VII IX than in compounds
I VI apparently due to longer distance (through two
The latter condition is as a rule met by inter-
calators with a core that as was already mentioned
should by definition be built of several fused aromatic
rings. However for compounds externally binding to
DNA, e.g., netropsin, distamycin A [9], Hoechst-
3
3258, Hoechst-33258 [27] [see compounds X XIII
additional CH groups) from the modified terminal
2
in Scheme (3)] etc. the stabilization of the planar
molecular structure preventing reciprocal rotation of
separate building blocks of the molecule and there-
with increasing the overall quantum efficiency of
fluorescence can be achieved only in very viscous
media (e.g., in saccharose solution [30]), by insertion
of certain substituents (for instance, 3-dimethylamino-
propylcarbamoyl, as in compound XIV, Scheme 3
[27, 30]), or at specific interaction with a substrate.
substituents to the aromatic core.
Thus we found that for the intercalators were also
valid the statements which we had formulated before
27, 28, 30] by an example of the externally bound to
DNA compounds: In simulating the fluorescence
properties of dyes both specific and nonspecific to
nucleotides the factors listed below should be
considered.
[
(
3)
R = CH NHC(NH )=NH, n = 2 (X); R = H, n = 3 (XI); R = OH (XII); OCH CH (XIII);
2
2
2
3
C(O)NH[CH ] N(CH ) (XIV); R = H, R = NH , X = O (XV); R = R = NH , X = O (XVI); R = R = NH ,
2
3
3
2
2
2
2
1
X = S (XVII); R = COONa, R = NH , X = S (XVIII); R = COONa, R = N(CH ) , X = S (XIX); R =
2
3 2
3
2
3
1
2
3
1
2
1
CH , R = Im, R = H (XX); R = CH , R = H, R = Am (XXI); R = H, R = Im, R = H (XXII); R = H,
3
3
2
3
1
2
3
1
2
3
R = Am, R = H (XXIII); R = H, R = H, R = Im (XXIV); R = H, R = H, R = Am (XXV);
(
3) When the core of a molecule possesses a suf-
polar solvent can be significantly reduced by intro-
duction of substituents electron-acceptor with respect
to the core (or the quantum efficiency would be
increased if the introduced substituent would be an
electron-donor toward the core). On the contrary in
ficiently developed and rigid electronic conjugated
structure (as, for instance, in compounds I IX under
consideration in this study) the quantum efficiency of
fluorescence ( ) of this molecule in nonbound state in
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 39 No. 6 2003

DEPENDENCE OF FLUORESCENCE PROPERTIES OF COMPOUNDS FROM PSORALEN
885
the media of low permittivity (as, e.g., 2-propanol)
or at specific interaction with a substrate the above
mentioned effect of the substituents on the core of the
ligand is considerably reduced due to the changed
character of the microenvironment of the molecule,
and as a result the overall quantum efficiency of
fluorescence ( ) becomes close in value to of the
core of compound proper (i.e. without any substitu-
ents).
(4) However the pattern often is even more
complicated. That is, sufficiently often the com-
pounds in question can be present in solution in
several equilibrium forms (distinguished by extent of
ionization of the core and substituents, by the number
of intramolecular hydrogen bonds, etc.), and more-
over, the relative content of these forms can change
depending on pH and the solvent character.
For instance, taking phenylbenzazoles XV XIX
It is possible to build a series of substituents in the
order of decreasing electron-donor (and consequently
increasing electron-acceptor) activity with respect to
the core of the fluorophore (as we have for instance
done in [29, 30]) proceeding from certain simple
principles [28, 31] defined below:
The electrnegativity of heteroatoms with respect to
hydrocarbon moiety linked thereto decreases in the
series O> N> S> P;
(see Scheme 3) described in [33], it was found that
although nonionized NH group would have showed
2
electron-donor effect with respect to the aromatic
core, the fluorescence quantum efficiency in aqueous
buffer solution with no DNA was considerably
smaller for compound XVI ( W 0.12) than for com-
pound XV ( W 0.87) which was distinguished from
dye XVI only by the absence of a terminal amino
group at the benzoxazole fragment of its molecule. It
means that under the described conditions the amino
-
Electrons are capable to shift to the more electro-
negative atom to a greater extent than n-electrons, and
the latter shift better than -electrons;
group was present predominantly in the ionized
+
H N -form that behaved with respect to the bound
3
A heteroatom linked to an aromatic ring with an
ordinary bond shows with respect to the ring electron-
donor properties owing to its unshared electron pairs,
and the more the less is the electronegativity of the
heteroatom;
A heteroatom linked to a neutral nonaromatic
hydrocarbon moiety shows toward the latter electron-
acceptor properties, and the more the higher is the
proper electronegativity of the heteroatom;
The more a heteroatom withdraws the electron
density from one of groups bonded thereto, the less it
is prone to attract it from the other groups linked to it;
Positively charged substituents are electron-
acceptors with respect to a neutral aromatic core
bonded thereto, and negatively charged substituents
play part of electron-donors;
thereto benzoxazole core of compound XVI as elec-
tron-acceptor. Similarly nonionized carboxy group
should be an electron-acceptor toward the aromatic
fragment linked to it, and in ionized OOC-form the
character of the influence on the neutral core of the
compound becomes electron-donating. As a result at
replacement of amino group of compound XVII by a
carboxy group in compound XVIII the
value of
W
the latter considerably grew (from 0.07 in XVII to
0.55 in XVIII). Finally, for dyes XVIII and XIX
evidently the properties of the terminal dimethyl-
amino group in compound XIX were decisive: in
ionized state it was a weaker acceptor, and in non-
ionized form a stronger donor with respect to the
phenylbenzothiazole core than the corresponding
amino group in compound XVIII in the respective
therewith the greater delocalization of the charge, the les ss t ai st e ts h. eApparently this was the reason of higher
influence of this substituen, etc., and from the already
for dye XIX than for compound XVIII ( W 0.66 and
W
,
,
known Hammett s and Taft s constants [28, 32].
0.55 respectively).
It is logic to assume that in the same order as
decrease the electron-donor properties of substituents
would decrease the relative values of fluorescence
quantum efficiency of compounds having such
terminal substituents measured in water medium (at
high permittivity) in the absence of substrates capable
of specific interaction with these compounds, and also
at similar character of their core. On the contrary the
ratio of quantum efficiency of fluorescence measured
In the same way the quantum efficiencies of carb-
azoles XX XXV [34, 35] (see Scheme 3) in water
medium with no DNA may be compared using the
ratio 2/ = (I A )/(I A ) [27] [where I , I and A ,
1
2
1
1
2
1
2
1
A are intensities of fluorescence and optical density
2
of compounds (1) and (2) at wavelengths correspond-
ing to the maxima on their absorption and emission
spectra reduced to unit dye concentration]. After that
it is possible to establish that the derivatives with a
in alcohol and in water ( A
/
W) (or in water in the
methyl group in 9 position possess smaller
value
W
presence of DNA and in the absence of the poly-
nucleotide) would increase in the above indicated
order.
than analogous compounds with only a hydrogen in
the same position. Similarly the derivatives with
amidine (AM) groups in positions 2, 7 or 3, 6 have
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 39 No. 6 2003

8
86
SIBIRTSEV et al.
value than analogous compounds with
smaller
substrate solution was prepared by dissolving a dry
substance (Serva, Germany) in distilled water follow-
ed by treating with ultrasound (for homogenization
and reduction of light scattering) on UZDN-2 device
(Russia) for 15 s at 0.3 A current and with resonance
frequency 22 kHz. After the treatment the average
molecular weight of DNA was 3500 Da. The sodium
ethylenediaminetetraacetate and Tris (2-amino-2-
hydroxymethyl-1,3-propandiol) were also commercial
products from Serva, Germany. The other reagents
used were of chemically pure grade.
W
imidazoline (Im) substituents. This happens although
obviously in a neutral form of the compound the
methyl group is a stronger electron donor for the
carbazole core than single hydrogen, and Am groups
are stronger acceptors compared to Im groups. Thus
the observed pattern is apparently due to the presence
in the water medium at pH 7 (see the measuring
conditions in [34, 35]) alongside neutral forms of
compounds under study also positively-charged ones.
Since the basicity of nitrogen is known to decrease in
the series R N > R NH > RNH > NH [31] the
3
2
2
3
The absorption spectra of solutions under study
were recorded on spectrophotometer Beckman Model
relative equilibrium ratio of ionized and nonionized
forms should be larger for H C-N-derivatives than for
3
3
5 (Austria). The fluorescence was measured on a
analogous HN-derivatives, and for Im-derivatives
larger than for Am-derivatives. We believe that this
situation is reflected in the overall values (summing
spectrofluorimeter Hitachi model 850 (Japan). In
registering the spectra on fluorimeter the slots of
monochromators of excitation and emission were set
up all the forms) of
ation.
compounds under consider-
1
W
at 3 nm, rate of scanning 120 nm min , response
time 2 s, normal amplification of photomultiplier.
The excitation spectra were registered at wavelengths
corresponding to the maxima of the most longwave
among the registered excitation peaks of compound
under study. All the spectra were registered with
subtraction of background. The fluorescence spectra
were corrected using quantum counter basing on
alcoholic rhodamine B solution in keeping with the
instruction manual supplied with the spectrofluori-
meter. The spectral characteristics were measured
with the use of standard cells of square cross-section
The procedure of evaluating an overall electron-
donor or electron-acceptor effect of any substituent
on the core of fluorophore which is present in solu-
tion in N different equilibrium form we have treated
before [36]. In the most general case the combined
influence on the fluorescence of the compound in
question of all above mentioned factors should be
taken into account.
EXPERIMENTAL
1
cm thick.
Quantum efficiency of fluorescence ( ) of dyes
were determined by comparison with quinine sulfate
1
(
M solution in sulfuric acid taken as reference
0.55).
8-Acetyl-4,9-dimethyl-2H-furo[2,3-h]chromen-2-
2
00 MHz from solutions in deuterochloroform
one (I) and 2-acetyl-3,5,9-trimethyl-7H-furo[3,2-g]-
chromen-7-one (II). To a boiling solution of an ap-
propriate o-acetyl-7-hydroxycoumarin (2.3 mmol)
and potassium carbonate (1.58 g, 0.01 mmol) in
anhydrous acetonitrile (40 ml) was added dropwise at
vigorous stirring a solution of chloroacetone (0.24 g,
2.6 mmol) in anhydrous acetonitrile (15 ml). The
reaction mixture was stirred at reflux for 3 4 h. Then
the mixture was cooled to room temperature, aceto-
nitrile was evaporated in a vacuum. To the residue
10% water solution of hydrochloric acid was added
to neutralize potassium carbonate till pH 2 3. The
precipitate was filtered off,washed with water, and
dried in air. Then it was recrystallized from 95%
ethanol.
(
internal reference TMS) or from solutions in
dimethyl sulfoxide. Mass spectra were measured on
SSQ-710 instrument (Finnigan-MAT) at ionizing
electrons energy 70 eV. The reaction progress was
monitored by TLC on Silufol UV-254 plates (eluent
chloroform). The chromatographic separation was
carried out on silica gel(eluent chloroform).
All photometric and fluorometric measurements
were carried out at concentration of compounds I IX
equal to IX) C = 4 10 M and at temperature 20
6
L
2
5 C in an aqueous buffer solution of the following
composition: 0.01 M NaCl, 0.01 M Na EDTA, and
2
0
.01 M Tris (pH 7.4), and also in 2-propanol. As a
substrate was used calf tymus DNA (58% AT-pairs;
average molecular weight per one nucleotide 326Da,
Compound I was prepared from 8-acetyl-7-
hydroxy-4-methylcoumarin. Yield 0.38 g (65%),
1
1
molar extinction
=
6600 M cm ). The
2
60
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 39 No. 6 2003

DEPENDENCE OF FLUORESCENCE PROPERTIES OF COMPOUNDS FROM PSORALEN
887
1
mp231 232 C (ethanol). HNMR spectrum (CDCl ),
of reaction the mixture was poured into ice water
3
,
ppm: 2.51 d (3H, C CH , J 1.14 Hz), 2.92 s (3H,
(150 ml). The separated precipitate was filtered off,
washed with water, and dried in air. Then it was
recrystallized from methanol. Yield of alcohol V was
4
3
9
3
C CH ), 2.63 s (3H, COCH ), 6.30 d (1H, C H,
3
3
6
J 1.14 Hz), 7.43 d (1H, C H, J 8.84 Hz), 7.67 d (1H,
5
1
C H, J 8.86 Hz). Mass spectrum, m/z (I , %): 256
0.39 g (78%), mp 160 161 C (methanol). H NMR
rel
+
(
100) [M] . Found, %: C 70.26; H 4.68. C H O .
Calculated, %: C 70.31; H 4.72; O 24.97.
spectrum (CDCl3), , ppm: 1.65 d (3H, CH CH,
1
5
12
4
3
4
J 6.64 Hz), 2.12 br.s (1H, OH), 2.46 (3H, C CH , J
3
9
1
.18 Hz), 2.55 s (3H, C CH ), 5.12 q (1H, CH CH,
J 6.64 Hz), 6.21 d (1H, C H, J 1.18 Hz), 7.32 d
(1H, C H, J 8.68 Hz), 7.43 d (1H, C H, J 8.68 Hz).
Mass spectrum, m/z (I , %): 258 (46) [M] , 243
3
3
Compound II was prepared from 6-acetyl-7-
3
hydroxy-4,8-dimethylcoumarin. Yield 0.35 g (56%),
mp239 240 C (ethanol). HNMR spectrum (CDCl ),
6
5
1
+
3
5
rel
,
ppm: 2.53 d (3H, C CH , J 1.16 Hz), 2.63 s (3H,
+
3
9
3
(100) [M CH ] . Found, %: C 69.72; H 5.44.
3
C CH ), 2.64 s (3H, COCH ), 2.65 s (3H, C CH ),
3
3
3
6
4
C H O . Calculated, %: C 69.76; H 5.46.
1
5
14
4
6
.30 d (1H, C H, J 1.16 Hz), 7.69 s (1H, C H). Mass
+
spectrum, m/z (I , %): 270 (100) [M] . Found, %:
2-(1-Methoxyethyl)-3,5,9-trimethyl-7H-furo-
rel
C 71.16; H 5.23. C H O . Calculated, %: C 71.10;
H 5.22; O 23.68.
[3,2-g]chromen-7-one (VI). To a solution of com-
pound II (0.54 g, 2 mmol) in a mixture of dichloro-
methane (70 ml) and methanol (10 ml) was added
sodium borohydride (0.1 g, 2.6 mmol). The reaction
mixture was vigorously stirred at room temperature
for 6 8 h (TLC monitoring). On completion of reduc-
tion to the reaction mixture was added concn. HCl
1
6
14
4
8
-Acetyl-4,9-dimethyl-2H-furo[2,3-h]chromen-2-
one ketoxime (III) and 2-acetyl-3,5,9-trimethyl-7H-
furo[3,2-g]chromen-7-one ketoxime (IV). To a solu-
tion of an appropriate acetylfurocoumarin (2.3 mmol)
in a mixture of ethanol (30 ml) and pyridine (10 ml)
was added hydroxylamine hydrochloride (0.24 g,
(3 ml), and the mixture was stirred for 3 min more.
Then to the reaction mixture 200 ml of water was
added, and it was transferred into a separatory funnel.
The organic layer was separated, and the water layer
was extracted with dichloromethane (2 5 ml). The
organic solutions were combined and dried with
anhydrous sodium sulfate. The solvent was evaporat-
ed in a vacuum, and the residue was subjected to
column chromatography on silica gel (eluent chloro-
form). Yield of ether VI was 0.41 g (71%), mp
3
3
.5 mmol), and the mixture obtained was boiled for
4 h. Then the hot reaction mixture was poured into
ice water (150 ml). The separated precipitate was
filtered off, washed first with 10% solution of hydro-
chloric acid and then with water, and dried in air.
For compound III yield was 93%, mp 234 235 C
1
(
methanol). H NMR spectrum (DMSO-d ), , ppm:
6
4
2
.26 s (3H, CH C=NOH), 2.54 (3H, C CH ,
3
3
9
3
J 1.10 Hz), 2.70 s (3H, C CH ), 6.33 d (1H, C H,
1
3
128 129 C (methanol). H NMR spectrum (CDCl3),
6
J 1.10 Hz), 7.52 d (1H, C H, J 8.80 Hz), 7.72 d
,
ppm: 1.62 d (3H, CH CH, J 6.62 Hz), 2.31 s (3H,
5
3
(
1H, C H, J 8.80 Hz), 11.65 s (1H, OH). Mass
3
5
C CH ), 2.52 d (3H, C CH , J 1.12 Hz), 2.60 s (3H,
+
3
3
spectrum, m/z (I , %): 271 (80) [M] , 254 (100)
9
rel
C CH ), 3.30 s (3H, OCH ), 4.58 m (1H, CH CH),
+
3
3
3
[
M OH] . Found, %: C 66.38; H 4.82; N 5.14.
6
4
6
.24 d (1H, C H, J 1.12 Hz), 7.50 s (1H, C H).
C H NO . Calculated, %: C 66.41; H 4.83;
+
1
5
13
4
Mass spectrum, m/z (I , %): 286 (20) [M] , 271
54) [M CH ] , 255 (100) [M OCH ] . Found, %:
rel
N 5.16; O 23.59.
+
+
(
3 3
For compound IV yield was 92%, mp 234 235 C
C 71.27; H 6.32. C H O . Calculated, %: C 71.31;
17 18 4
1
(
2
(
methanol). H NMR spectrum (DMSO-d ), , ppm:
H 6.34.
-(1,2,3,4-Tetrahydro-9H-carbazol-9-yl)propio-
6
3
.21 s (3H, CH C=NOH), 2.45 s (3H, C CH ), 2.50
3
3
3
5
9
3H, C CH , J 1.16 Hz), 2.55 s (3H, C CH ), 6.23
3
3
nitrile (VIII). To acrylonitrile (28.12 g, 0.53 mol)
at 40 45 C was added dropwise phenylhydrazine
6
4
d (1H, C H, J 1.16 Hz), 7.69 s (1H, C H), 11.42 s
(
[
1H, OH). Mass spectrum, m/z (I , %): 285 (33)
rel
(
54.07 g, 0.5 mol) and Triton B (5 drops). Then the
+
+
M] , 269 (100) [M H CH ] . Found, %: C 67.32;
3
mixture was heated at 100 C for 1.5 h. On cooling
cyclohexanone (53.97 g, 0.55 mol) an ethanol
H5.32; N 4.90. C H NO . Calculated, %: C 67.36;
1
6
15
4
H 5.30; N 4.91; O 22.43.
(100 ml) was added. The mixture was heated again,
8
-(1-Hydroxyethyl)-4,9-dimethyl-2H-furo[2,3-h]-
and into the boiling reaction mixture was added
dropwise concn. HCl (50 ml). Then the mixture was
boiled for 30 min, cooled, the separated precipitate
was filtered off, washed with water, dried, and
recrystallized from alcohol. Yield of compound VIII
chromen-2-one (V). A suspension of 8-acetyl-4,9-
dimethylangelicin (0.51 g, 2.0 mmol) and sodium
borohydride (0.1 g, 2.6 mmol) in ethanol (40 ml) was
stirred at room temperature for 6 8 h. On completion
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 39 No. 6 2003

888
SIBIRTSEV et al.
was 95.33 g (85%), fine colorless crystals, mp117
Dabrowska, M., and Rozanski, A., Acta Biochim.
Polonica, 2000, vol. 47, p. 23.
7. Morgan, A.R., Lee, J.S., Pulleyblank, D.E., Mur-
ray, N.L., and Evans, D.H., Nucl. Acids Res., 1979,
vol. 7, p. 547.
1
1
2
7
19 C. H NMR spectrum (CDCl ), , ppm: 1.8
.0 m (4H), 2.6 2.7 m (4H), 2.7 t (2H), 4.3 t (2H),
.0 7.5 m (4H).
3
3
-(1,2,3,4-Tetrahydro-9H-carbazol-9-yl)pro-
pionic acid (VII). Compound VIII (67.23 g,
.3 mol) was dissolved in boiling concn. HCl, and
8
. Darzynkiewicz, Z., Traganos, F., Kapuscinski, J.,
Staiano-Coico, L., and Melamed, M.R., Cytometry,
0
1
984, vol. 5, p. 355.
the solution was boiled for 2 h more. On cooling to
room temperature it was diluted with water (300 ml),
the separated precipitate was filtered off, washed with
water, and dried. Yield of compound VII was 58.39
g (80%), fine colorless crystals, mp 119 120 C.
9
. Zimmer, C. and Wahnert, U., Prog. Biophys. Molec.
Biol., 1986, vol. 47, p. 31.
0. Campejohn, R.S., Macarthey, J.C., and Mor-
ris, R.W., Cytometry, 1989, vol. 10, p. 410.
11. Ivanov, S.D., Doctoral Sci. (Biol.) Dissertation,
1
1
H NMR spectrum (CDCl3), , ppm: 1.8 2.0 m
Leningrad: TsNIRRI, 1992, p. 306.
(
7
4H), 2.6 2.7 m (4H), 2.7 t (2H), 4.3 t (2H), 7.0
.5 m (4H), 9.1 © (1H).
1
2. Ivanov, S.D., Korytova, L.I., Yamshanov, V.A.,
Ilyn, N.V., and Sibirtsev, V.S., J. Exp. Clin. Cancer
Res., 1997, vol. 16, p. 183.
3
-(1, 2, 3, 4-Tetrahydro-9H-carbazol-9-yl)-
propanoylhydrazide (IX). A mixture of compound
VII (48.66 g, 0.2 mol), ethanol (100ml), toluene
13. Pjura, P.E., Grzeskowiuk, K., and Dickerson, R.E.,
J. Mol., Biol., 1987, vol. 197, p. 257.
(
150 ml), and toluenesulfonic acid (0.2 g) was boiled
14. Pelton, J.C. and Wemmer, D.E., J. Am. Chem. Soc.,
1990, vol. 112, p. 1393.
15. Eriksson, S., Kim, S.K., Kubista, M., and Nor-
den, B., Biochemistry, 1993, vol. 32, p. 2987.
16. Sibirtsev, V.S., Garabadzhiu, A.V., and Ivanov, S.D.,
Bio. Org. Khim., 2001, vol. 27, p. 64.
17. Muller, W. and Grothers, D.M., Eur. J. Biochem.,
1975, vol. 54, p. 267.
18. Muller, W., Bunemann, H., and Dattagupta, N.,
Eur. J. Bio. Chem., 1975, vol. 54, p. 279.
19. Miller, K.J., Newlin, D.D., Biopolymers., 1982,
vol. 21, p. 633.
for 15 h distilling off the water toluene azeotrope.
Then the reaction mixture was washed with water,
1
water. Then mixture was evaporated at reduced pres-
sure, and the residue was distilled in a vacuum. Thus
was obtained ethyl 3-(1,2,3,4-tetrahydro-9H-carbazol-
9
1
0% solution of sodium carbonate, and again with
-yl)propionate (XXVI) in 54.27 g (85%) yield, nD20
.568, bp 228 230 C (15 mm Hg). Then a mixture of
compound XXVI (13.56 g, 0.05 mol), hydrazine
hydrate (40 ml), and ethanol (150 ml) was heated to
1
00 C for 3 h. Then the mixture was evaporated at
reduced pressure and left standing for 10 h for
crystallization. The separated precipitate was filtered
off, washed with water, with alcohol, and dried.
20. Tomosaka, H., Omata, S., Hasegawa, E., and
Anzai, K., Biosci., Biotech. Biochem., 1997, vol. 61,
p. 1121.
Yield of compound IX was 11.83 g (92%), mp
21. Guiotto, A., Rodighiero, P., Manzini, G., Pasto-
1
1
1
42 143 C. H NMR spectrum (CDCl ), , ppm:
.8 2.0 m (4H), 2.4 t (2H), 2.6 2.7 m (4H), 3.7 s
rini, F., Bordin, F., Baccichetti, F., Carlassare, D.,
3
,
Vedaldi, D., Dall Acqua, F., Tamaro, M., Rec-
(
2H), 4.3 t (2H), 6.7 s (1H), 7.0 7.4 m (4H).
chia, G., and Cristofolini, M., J. Med. Chem., 1984,
vol. 27, p. 959.
REFERENCES
22. Gia, O., Anselmo, A., Consoni, M.T., Antonello, C.,
Uriarte, E., and Caffieri, S., J. Med. Chem., 1996,
vol. 39, p. 4489.
,
1
2
3
. Denison, L., Haigh, A., D Cunha, G., and Mar-
tin, R.F., Int. J., Radiat. Biol., 1992, vol. 61, p. 69.
. Gazzard, B.G., J. Antimicrob. Chemother., 1989,
vol. 23, p. 67.
23. Leonov, A.A., Sergiev, P.V., Dontsova, O.A., and
Bogdanov, A.A., Molek. Biol., 1999, vol. 33, p. 1063.
24. Patrick, D.A., Boykin, D.W., Wilson, W.D.,
Tanious, F.A., Spychala, J., Bender, B.C., Hall, J.E.,
Dykstra, C.C., Ohemeng, K.A., and Tidwell, R.R.,
Eur. J. Med. Chem., 1997, vol. 32, p. 781.
25. Kanzava, F., Nishio, K., Kubota, N., Cancer Res.,
1995, vol. 55, p. 2806.
. Van Iperen, H.P. and van Henegouwen, G.M.J.B.,
J. Photochem. Photobiol., B: Biology, 1997, vol. 39,
p. 99.
. Baguley, B.C., Molec. Cell. Biochem., 1982, vol. 43,
p. 167.
4
5
6
. Osashi, M. and Oki, T., Expert. Opin. Therp.
Patents, 1996, vol. 6, p. 1285.
. Bartulewicz, D., Markowska, A., Wolczynski, S.,
26. Norbert, H., Jbara, R., Khamadi, F., Heterocycles,
1998, vol. 48, p. 1609.
27. Sibirtsev, V.S., Garabadzhiu, A.V., and Ivanov, S.D.,
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 39 No. 6 2003

DEPENDENCE OF FLUORESCENCE PROPERTIES OF COMPOUNDS FROM PSORALEN
889
Bioorg. Khim., 1997, vol. 23, p. 969.
Cambridge Univ. Press, 1973.
2
2
8. Sibirtsev, V.S., Garabadzhiu, A.V., and Ivanov, S.D.,
Bioorg. Khim., 1994, vol. 20, p. 650.
9. Krasovitskii, B.M. and Bolotin, B.M., Organicheskie
lyuminofory (Organic Luminophores), Moscow:
Khimiya, 1984, p. 343.
0. Sibirtsev, V.S., Garabadzhiu, A.V., and Ivanov, S.D.,
Bioorg. Khim., 1995, vol. 21, p. 731.
1. Gaupman, Z., Grefe, Yu., and Remane, Kh., Orga-
nicheskaya khimiya (Organic Chemistry), Moscow:
Khimiya, 1979, 832 p.
33. Ivanov, S.D., Kvitko, I.Ya., Rtishchev, N.I.,
Fomina, E.I., and Nagorskaya, L.P., Bioorg. Khim.,
1989, vol. 15, p. 648.
34. Tanious, F.A., Ding, D., Patrick, D.A., Tidwell, R.R.,
and Wilson, W.D., Biochemistry, 1997, vol. 36,
p. 15315.
35. Tanious, F.A., Ding, D., Patrick, D.A., Bailly, Ch.,
Tidwell, R.R., and Wilson, W.D., Biochemistry,
2000, vol. 39, p. 12091.
36. Sibirtsev, V.S., Glibin, E.N., adn Ivanov, S.D.,
Zh. Org. Khim., 2000, vol. 36, p. 1865.
3
3
3
2. Johnson, C.D., The Hammett Equation, Cambridge:
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 39 No. 6 2003