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УДК 615.012.01. Received
RADICAL BINDING ACTIVITY PRODUCT OF THE INTERACTION OF THE METHYL
ESTER OF 3,4-DIHYDROXY-6-OXO-2,4-HEXADIENE ACID WITH 1,2DIAMINOBENZENE
Karmanova Olga Gennadevna1*, Zykov Svetlana Sergeevna2+,
Lyuboseev Vladislav 2, Peter Mukovoz3, Vladislav Kozminyh4
1
Department of General Chemistry. VPO "Moscow Institute of Physics and Technology (State
University)." st. Kerch, 1-A, building 1. Moscow, 117303, Russia
2
Department of animal science. FKOU VPO Perm Institute FSIN Russia. St. Karpinski, 125.
Perm, 614012. Russia. Tel .: (342) 228-60-77 (253). E-mail: [email protected]
3
Institute of Cellular and Intracellular Symbiosis, RAS. St. Pioneer, 11.
Orenburg, 460000, Russia.
4
Department of Chemistry. VPO «Perm State University of Humanities and Education." St.
Siberian,24 Perm, 614000, Russia.
* Leading direction; + Corresponding author
Tags: 1,3,4,6-tetracarbonyl compounds, methyl 3,4-dihydroxy-6-oxo-2,4-hexadiene acid, methyl
(2Z) - [3- (2-oxopentyl) quinoxalin-2- (1H) -ylidene] etanoat radical binding activity.
Annotation
Reaction of methyl 3,4-dihydroxy-6-oxo-2,4-hexadiene and 1,2-acid diaminobenzene obtained
methyl (2Z) - [3- (2-oxopentyl) quinoxalin-2 (1H) -ylidene] etanoat. On the basis of IR and 1H
NMR spectroscopy and mass spectroscopy the structure of the synthesized compounds was
found out. Its radical binding activity in the reaction with DPPH (DPPH) was studied, which was
at the level of the standard anti-radical activity - water-soluble form of vitamin E (Trolox).
Introduction
It is known that 1,3,4,6-tetracarbonyl compound (TCS) react with 1,2-diaminobenzene to
form bis-2,3-dioxo-1,3,4,6 oksoilidenproizvodnyh [1-6]. The aim of this work is to study the
interaction of 1,2-diaminobenzene with methyl 3,4-dihydroxy-6-oxo-2,4-hexadiene acid [7],
which is an intermediate class of tetracarbonyl compounds directly from 1,3,4,6 tetraoksogeksanov with traditional end-keto esters of hexadiene acid [7-9].
Experimental part
Method of synthesis of methyl (2Z) - [3- (2-oxopentyl) quinoxalin-2 (1H) -ylidene]
etanoate (2). Yield 3.44 g (48%), m.p. 94-95 ° C.
To 6.01 g (25 mmol) of the compound (1) in 50 ml of ethanol was added 2.7 g (25 mmol) of 1,2diaminobenzene after brief refluxing. The solvent was evaporated, the residue triturated with
diethyl ether and dried, then the yielding compound was found (2). Recrystallization from ethyl
acetate.
Found,%: C 67.12%; H 6.34%; N 9.67%; S16N18N2O3. Calculated,%: C 67.33%; H
6.53%; N 9.78%. M 286.33.
IR spectrum, ν, cm-1 (vases. Oil): 1732 (O = C (CH2) 2CH3), 1604 (O = COCH3), 1569 (C = C,
chelate, C6H4), 1462 δas (CH3), 1435 (N = C), 1374 δs (CH3), 1179 (C-OCH3), 1154, 1114,
1067, 998 νskeletnye (CC), the pendulum 854 δ (CH3).
Mass spectrum (m / z, Iotn.%): 287 M + 1˥ + (3,08), 286 M˥ + (15.60), 255 M + SN3O˥
O
Pr
H
N
N
H
(8.94), 243 M + S3N7˥ (6.46) 216
O
Pr
H
N
216
˥+ (100), 215 М-С3Н7СО˥+ (3.27),
N
H
˥ (100) 199
+
N
ÑÍ
N
CH COCH
2
2
3
˥+ (3.47), 184 М-С3Н7СО-СН3О˥+
N
N
(70.13), 158
˥+ (22.16), 103 С6Н5СN˥+ (8.50), 77 С6Н5˥+ (10.46), 71
С3Н7СО˥+ (42.46), 43 С3Н7˥+ (85.79).
1H NMR (DMSO-d6, δC, ppm): 0.97 t (3H, CH2CH2CH3, J 7.0 Hz, 2B), 1.63 m (2H,
CH2CH2CH3, 2B), 2.38 m (2H, CH2CH2CH3, J 7.0 Hz , 2B), 3.67 c (3H, OCH3, 2B), 4.00 a
(2H, CH2COOCH3, 2B), 5.67 c (1H, CH, 2B, 94%), m 7.41-7.75 (4H, S6H4, 2B), 14.70 with
(1H, NH, 2B).
1H NMR (CDCl3, δC, ppm): 0.96 t (3H, CH2CH2CH3, J 7.0 Hz, 2B, 2C), 0.97 t (3H,
CH2CH2CH3, J 7.0 Hz, 2D), 1.71 m (2H, CH2CH2CH3 , 2B, 2C), 1.73 m (2H, CH2CH2CH3,
2D), 2.44 m (2H, CH2CH2CH3, J 7.0 Hz, 2B, 2C), 2.61 m (2H, CH2CH2CH3, J 7.0 Hz, 2D),
3.75 c (3H , OCH3, 2B), 3.73 c (3H, OCH3, 2C), to 3.90 (3H, OCH3, 2D), 4.09 a (2H,
CH2COOCH3, 2B), 4.24 a (2H, CH2COOCH3, 2D), 5.27 a (1H , CH, 2D, 28%), from 5.49 (1H,
CH, 2B, 42%), from 5.65 (1H, CH, 2C, 30%), m 6.93-8.05 (4H, S6H4, 2B, 2C, 2D), 11.69 c
(1H, NH, 2D), 14.12 c (1H, NH, 2C), with 15.03 (1H, NH, 2B).
Results and discussion
The interaction of the compound (1) with 1,2-diaminobenzene was first studied, whereby
methyl(2Z)-[(3Z)-3-(2-oxopentiliden)-3,4-dihydrohinoxalin-2(1H)-ilidene]etanoat
(2)
[7]
(Scheme 1) were found out. Previously synthesized compound was not known.
Compound (2) is a yellow crystalline substance soluble in common organic solvents and
insoluble in water. . In contrast to 2,3-bis (aroylmethylene) quinoxalines, existing in the form of
bis-chelated NH system [3-6], the compound (2) are in the form of endo-2,3-C = N- isomer 2A,
as indicates the absence of the corresponding absorption bands in the region of amino 3100-3500
cm-1 in the IR spectra. Presence of RF signal of absorption nonconjugated carbonyl group of the
acyl chain at 1732 cm-1 and the absorption signal of the ester carbonyl group at 1604 cm-1 also
confirms the structure 2A.
Scheme 1. Synthesis of methyl (2Z) - [(3Z) -3- (2-oksopentiliden) -3,4dihydrohinoksalin-2 (1H) -ylidene] etanoat (2)
1
O
MeO
Me
O
+
MeO
2
OMe
+
Me
H
O
O
+
H
2:1:2
Alk
O
O
MeONa
Pr
- CH 3OH
Pr
H2N
+
H2N
- 2 H2O
N
H
2
O
OMe
O
OMe
O
N
H
H
O
1
In polar solvents (DMSO) in the compound (2) predominant is the form of monohelat 2B
(94%), a stable chelate ring NH, indicating that considerable stability of this solution of DMSO
is in the form (Scheme 2)
Scheme 2. The tautomeric form
dihydrohinoxalin-2(1H)-ilidene]etanoat (2)
O
Pr
H
O
Pr
of
methyl(2Z)-[(3Z)-3-(2-oxopentidilen)-3,4-
H
O
Pr
N
N
N
N
N
N
H
H
OMe
2Â O
OMe
2C O
Pr
O
OMe
N
N
2À O
O
OMe
2D
Apparently, DMSO molecules polarize the carbonyl group of the acyl-level, creating
conditions for the delocalization of double bonds in the chelate stabilized by intramolecular
hydrogen bonds in the steady-chelating heterocyclic NH.
At the same time, the ester moiety of the molecule does not form a chelate-NH
depolarizing influence due to the unshared electron pairs of oxygen alkoxy, weakening the
charge on the carbonyl carbon of the ester group unit and polarizing molecules DMSO strength
giving the result of insufficiency
This confirms 1H NMR spectrum of the compound (2) recorded in DMSO-d6, wherein
signals other than n-propyl level such as, methoxy, ester moiety, and groups of heterocyclic
proton signals mono methine proton at δ 5,67 ppm, mono amino proton NH-chelate at δ 14,70
ppm mono and two methylene protons of the methyl acetate moiety at δ 3,67 ppm integral with
comparable intensities are present.
Furthermore predominant form of monohelatnoy 2B is present in the spectrum signals of
the two minor forms 2C and 2D each containing with not more than 3%. Slight their content
probably is connected with the lower stability of the structure having the chelating moiety with
the ester units. This also may be caused by specific solvation of polar molecules DMSO oxygen
of the carbonyl group of the ester moiety, reducing it effective negative charge and preventing
the formation of NH ... O = C-chelate IMHB stabilizing NH-chelating heterocyclic.
Change the non-polar solvent to chloroform leads to significant changes in the ratios of
the tautomeric forms. Thus, the content of forms 2C and 2D increases to 30% and 28%,
respectively. The reason for this apparently is the absence of the polarizing effect of the solvent
molecules, which do not create conditions for delocalization of double bonds in the chelate
anymore. 2B form stability decreases, resulting in the formation of alternate tautomers 2C and
2D.
Thus, 2C correspond form two magnetically equivalent mono methine protons at δ 5,65
ppm mono and two magnetically equivalent protons of two amino groups NH-chelates at δ 14.12
ppm
Thus, form 2C correspond two magnetically equivalent mono methine protons at δ 5,65
ppm mono and two magnetically equivalent protons of two amino groups NH-chelates at δ 14.12
ppm. 2B form of content in deuterochloroform is 42%. This form correspond to mono methine
proton with the value of the chemical shift δ 5.49 ppm mono amino proton NH-chelate with the
value of the chemical shift δ 15.03 ppm and mono two methylene protons of the methyl acetate
with a fragment value of the chemical shift δ 4.09 ppm integral with comparable intensities.
A shift signal of the methine proton bis-chelated form of into the weak field 2C (at δ 0.16
ppm) as compared with the shape monohelatnoy 2B explains coupling and chain extension,
respectively, decrease in the intensity of the external magnetic field in the area of the two
anisotropic kvazigeterotsiklam conjugate in which are located methine protons. The presence of
such a pairing is evidenced by the fact that the two methine proton and two protons of two amino
groups of two different chelates are magnetic equivalent and give appropriate, unsplit mono
signals in the 1H NMR spectrum.
At the same time, the shift of the signal of two magnetically equivalent protons of two
amino bis-chelated form of 2C in strong fields (at δ 0.91 ppm) compared with the signal
monohelat form of 2B also well explained by chain extension interface, in which the electron
density on the atoms nitrogen increases, resulting in the screening of the NH protons.
The signal value of the two methylene protons monohelatnoy form of 2B shifted to a
strong field compared to the same signal form of 2D (at δ 0,15 ppm), which is probably due to
the shielding effect of lone pairs of oxygen metoksizvena these protons. In the form of 2D
methoxy link adjacent to the methylene group of the acetyl moiety is absent, so the
corresponding signal of the group shifted to weak fields.
Signals corresponding to the endocyclic form 2A 1H NMR spectra of the compound (2),
were not detected.
The nature of the mass fragmentation quinoxaline (2) is consistent with the previously
established general laws of the collapse of the well-known quinoxaline derivatives [7, 10-12].
The mass spectrum of quinoxaline (2) there is an intense molecular ion signal 287 M˥ +.
Further fragmentation of molecules by electron impact in the gas phase takes place in four main
areas (Figure 3).
The first direction is the fragmentation of the molecule quinoxaline (2) is associated with
the separation of the methyl acetate fragments M - SN2SOOSN3˥ + and the formation of
O
Pr
H
N
N
H
The second and third directions
correspond to the separation of the acyl ions S3N7SO˥ + ions and SN3O˥ + to
fragment ions with maximum intensity
H
N
CH2
N
form an intense signal ketene ion
The fourth line corresponds to the rule of
maximum emission radical and characterized by the formation of very intense ion
signals S3N7˥ +. Subsequent fragmentation associated decay process azine ring
O
N
and further destruction to the ions
N
and PhCN˥+.
Scheme 3. Mass fragmentation quinoxaline (2)
IV
II
O
Pr
H
N
III
N
H
2
I
O
OMe
To determine the antiradical activity reaction was used with a stable free radical DPPH
(DPPH) (production Sigma-Aldrich) [13]. The concentration of DPPH in 95% ethanol was
6.5h10-5M. As a reference standard of antiradical activity of Trolox was used (production
Sigma-Aldrich) (Table 1).
Table 1
Antiradical activity of methyl (2Z) - [(3Z) -3- (2- oksopentiliden) -3,4-digidrohinoksalin2 (1H) - ylidene] etanoat (2)
Compound
2
Trolox
Decrease of radicals Q,%
86.51 ± 6.42*
87.40 ± 5.18*
* p <0,05 compared to control
The reaction mixture, which consists of 3 ml of 0.3 mM DPPH solution, 1 ml Tris-HCl,
pH = 7.4 and 1x10-5M Trolox or substances dissolved in DMSO, incubated under standard
conditions of temperature (T = 293K) for 30 minutes. Then, at a wavelength of 517 nm in it
cuvettes with a 0.5 cm thick layer of the optical density of the reaction mixture at
photoelectrocolorimeter CPK-3-01. Control measurements were made of the synthesized
compounds without DPPH at 517 nm.
The inhibitory effect is a decrease of DPPH radicals (Q), which is calculated by the
formula:
Q= 100(D0 – Dx)/D0
where D0 - optical density of a control solution of DPPH, Dx - DPPH absorbance of the
solution in the presence of the test substance or reference standard solution. The resulting
measurements were treated by standard methods using Student's t test (confidence level was
0.95). The result represents the mean ± standard error of the mean.
Methyl (2Z) - [(3Z) -3- (2-oksopentiliden) -3,4-dihydro quinoxalin-2 (1H) -ylidene] etanoat (2) showed binding activity expressed radical standard level - soluble form of vitamin E trolox.
Findings
1.
studied, as a result of reaction of methyl 3,4-dihydroxy-6-oxo-2,4-hexadiene acid
with 1,2-diaminobenzene methyl (2Z) - [(3Z) -3- (2- oksopentiliden)
digidrohinoksalin -3,4-2 (1H) -ylidene] etanoat (2) was gained.
2.
Found that the compound (2) in solid state exists in the form 2A, solution of polar
solvents (DMSO) at a mono-NH-chelated 2B, and non-polar solvents
(chloroform) in three forms chelate 2B, 2C, 2D with advantage first shape.
3.
Revealed that the synthesized compound (2) exhibits pronounced radical binding
activity at the level of the standard - a water-soluble form of vitamin E
(Trolox).
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