Synthesis and antitubercular activity of first glucuronosyl phosphates and amidophosphates containing polymethylene chains

Article abstract of DOI:10.1134/S1070428017010092

Novel phosphorylated glycolipids based on glucuronic acid have been synthesized and shown to inhibit M. Tuberculosis H37Rv in vitro at a minimum inhibitory concentration of 12.5 μg/mL.

Full text of DOI:10.1134/S1070428017010092

ISSN 1070-4280, Russian Journal of Organic Chemistry, 2017, Vol. 53, No. 1, pp. 51–56. © Pleiades Publishing, Ltd., 2017.
Original Russian Text © E.S. Izmest’ev, O.V. Andreeva, R.R. Sharipova, M.A. Kravchenko, B.F. Garifullin, I.Yu. Strobykina, V.E. Kataev, V.F. Mironov,
2
017, published in Zhurnal Organicheskoi Khimii, 2017, Vol. 53, No. 1, pp. 56–61.
Synthesis and Antitubercular Activity
of First Glucuronosyl Phosphates and Amidophosphates
Containing Polymethylene Chains
a
a
a
b
a
E. S. Izmest’ev , O. V. Andreeva , R. R. Sharipova , M. A. Kravchenko , B. F. Garifullin ,
a
a
a
I. Yu. Strobykina , V. E. Kataev ,* and V. F. Mironov
a
Arbuzov Institute of Organic and Physical Chemistry, Kazan Scientific Center, Russian Academy of Sciences,
ul. Arbuzova 8, Kazan, 420088 Tatarstan, Russia
*
e-mail: kataev@iopc.ru
b
Ural Research Institute of Phthisiopulmonology, Ministry of Health Protection of the Russian Federation,
ul. 22 Parts’’ezda 50, Yekaterinburg, 620039 Russia
Received July 21, 2016
Abstract—Novel phosphorylated glycolipids based on glucuronic acid have been synthesized and shown to
inhibit M. Tuberculosis H37Rv in vitro at a minimum inhibitory concentration of 12.5 μg/mL.
DOI: 10.1134/S1070428017010092
Increased interest of researchers in glucuronic acid
Herein we report the synthesis of first glucuronosyl
phosphates and amidophosphates containing polymeth-
ylene chains (phosphorylated glycolipids) (Scheme 1).
As starting compound we used D-(+)-glucurono-3,6-
lactone (1) which was converted to glucuronic acid
methyl ester [22] by the action of sodium methoxide,
and the subsequent acylation afforded methyl
1,2,3,4-tetra-O-acetyl-D-glucopyranuronate (2). In the
conjugates, primarily in glucuronides, is determined by
the fact that they are the most important products of
second phase metabolism of xenobiotics, drugs, and
hormones in humans [1, 2]. Glucuronides are just those
compounds which are involved in transport from one
part of organism to another and excretion therefrom.
Glucuronidation not only ensures transformation of
xenobiotics into readily soluble forms that are bio-
logically friendly and conducive to excretion but also
gives rise to new biologically active compounds [2–7].
Furthermore, in recent time glucuronides have been
used as prodrugs for targeted delivery of, e.g., anti-
biotics or anticancer agents to damaged tissues where
they are hydrolyzed by the action of β-D-glucuronidase
to release the corresponding therapeutic agent [8–11].
Although glucuronidation of biomolecules in living
matter is effected by uridine diphosphate glucuronic
acid (UDP-glucuronosyltransferase) [2], phosphates
derived from glucuronic acid itself have received little
attention. On the other hand, numerous publications
deal with phosphorylated glycolipids based on glucose
1
H NMR spectrum of 2, the anomeric proton resonated
at δ 5.77 ppm as a single doublet with a vicinal cou-
3
pling constant J of 7.7 Hz, indicating stereospecific
opening of lactone 1 with formation of the β-anomer
of 2. The latter was converted to methyl 2,3,4-tri-
O-acetyl-1-bromo-1-deoxy-β-D-glucopyranuronate (3)
according to the procedure described in [23], and the
hydrolysis of 3 in aqueous acetone in the presence of
silver carbonate according to [24] gave methyl
2,3,4-tri-O-acetyl-D-glucopyranuronate (4) which was
isolated in quantitative yield as a mixture of α- and
1
β-anomers (the H NMR spectrum of 4 contained two
sets of signals). The anomeric proton of the α-anomer
of 4 resonated as a doublet at δ 5.48 ppm with a vicinal
coupling constant of 2.9 Hz, whereas the correspond-
ing signal of the β-anomer appeared as a doublet at
[
12, 13], glucosamine [13–15], galactose [13, 16, 17],
mannose [18, 19], and arabinofuranose [20, 21], which
have been synthesized with the goal of revealing
efficient cell wall synthesis inhibitors active against
pathogenic bacteria and mycobacteria, primarily
against Mycobacterium tuberculosis.
3
δ 4.79 ppm ( J 7.7 Hz). These findings were consistent
with published data [23]. In the next stage, methyl
ester 4 was subjected to phosphorylation at the ano-
meric hydroxy group according to a modified proce-
5
1

5
2
IZMEST’EV et al.
Scheme 1.
O
O
HO
H
H
(
(
1) MeONa, MeOH
33% HBr–AcOH
O
OAc
O
Br
O
2) Ac O, HClO4
MeO
AcO
CH Cl2
MeO
AcO
2
2
HO
OH
O
OAc
OAc
OH
OAc
OAc
1
2
3
O
O
OH
Ag CO , H O, Me CO
MeO
2
3
2
2
AcO
OAc
OAc
4
O
O
H
O
O
(
1) Et N, THF
O–
3
MeO
P
O
P
(2) H O
2
4
+
O
OAc
AcO
HNEt3
O
Cl
OAc
5
(
(
(
(
1) Me CC(O)Cl
3
2) ROH, pyridine
3) I2
O
O
OR
O–
OR
O–
O
O
H N
O
O
3
4) Na S O
MeO
P
N H ·H O, MeOH
N
H
P
2
2
3
2
4
2
O
O
AcO
O
OAc
HO
OH
HNEt3
OAc
OH
6
, 7
9, 10
O
(
(
(
1) Me SiCl, CH Cl –Et N
3 2 2 3
2) I2
NHR
O–
NHR
O–
O
O
H N
3
O
O
3) RNH , pyridine
MeO
P
N H ·H O, MeOH
N
H
P
2
2
4
2
O
O
AcO
OAc
HO
OH
HNEt3
OAc
OH
8
11
2
'
Me 11'
7'
6
, 9, R = C
7
H15; 7, 10, R =
23
; 8, 11, R = C11H .
1
'
Me
Me
Me
dure [25] with 2-chloro-2H,4H-1,3,2-benzodioxaphos-
phinin-4-one which was prepared in turn as described
in [26]. Phosphonate 5 was thus isolated in 64% yield.
phosphate 8. By analogy with the procedure described
in [27], phosphonate 5 activated by pivaloyl chloride
was brought into reactions with heptan-1-ol and
3,7,11-trimethyldodeca-2,6,10-trien-1-ol (farnesol).
The resulting alkyl phosphonates were oxidized in situ
with iodine, and subsequent treatment of the reaction
mixtures with sodium thiosulfate afforded phosphates
6 and 7, which were isolated by column chromatog-
raphy in 47 and 83% yield, respectively. In the
1
Its H NMR spectrum contained only one set of sig-
nals, and the anomeric proton resonated as a doublet of
3
3
doublets at δ 5.75 ppm ( J
= 9.1, J
= 3.2 Hz).
1
-H,2-H
1-H,P
The PH proton signal was located at δ 6.84 ppm (d,
1
31
JPH = 641.2 Hz). In the P NMR spectrum of 5
we observed only one singlet at δ 0.9 ppm. These
P
3
1
findings indicated formation of 5 as pure β-anomer.
P NMR spectra of 6 and 7, the phosphorus nucleus
In the final stage, compound 5 was converted to
target glucuronosyl phosphates 6 and 7 and amido-
resonated as a singlet at δ –3.08 and –3.83 ppm,
respectively, whereas no PH signal was observed in
P
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 53 No. 1 2017

SYNTHESIS AND ANTITUBERCULAR ACTIVITY
53
1
their H NMR spectra. Heptyl phosphate 6 displayed
Scheme 1). The reaction of amidophosphate 8 with
hydrazine hydrate under the same conditions was
accompanied by hydrolysis, and we failed to isolate
hydrazide 11 in the pure state. Nevertheless, the
formation of 11 followed from the presence of the
1
in the H NMR spectrum the anomeric proton signal as
a single doublet of doublets at δ 5.75 ppm with vicinal
coupling constants J1-H,2-H = 7.8 and J1-H,P = 3.3 Hz.
The corresponding signal of 7 was a doublet of
doublets at δ 5.72 ppm with similar coupling constants
–
molecular ion peak with m/z 440.4 [M] (calculated for
–
(
J1-H,2-H = 7.8, J1-H,P = 3.3 Hz). The above data
C H N O P : 440.2) in the ESI mass spectrum of the
17
35
3
8
indicated β-orientation of the glycoside bond in both
reaction mixture.
glucuronosides.
According to published data, cell wall synthesis
inhibitors active against Micobacterium tuberculosis
were sought for among synthetic glycophospholipids
based on mannose [18, 19] and arabinofuranose
Following the procedure reported in [25], phos-
phonate 5 was converted to amidophosphate 8. In this
case, unlike the synthesis of phosphates 6 and 7, phos-
phonate 5 was activated by treatment with chloro-
[
20, 21]. Therefore, phosphates 6 and 7 and amido-
(
trimethyl)silane, and the phosphorus oxidation with
phosphate 8 were tested for antitubercular activity.
Glycophospholipids 6–8 inhibited the growth of
Micobacterium tuberculosis H37Rv in vitro for
iodine preceded the reaction with undecylamine as
nucleophile. Amidophosphate 8 was isolated in 37%
yield by chromatography. Its P NMR spectrum con-
tained only one singlet at δ 5.83 ppm, and only one
set of signals from the carbohydrate moiety was
observed in the H NMR spectrum of 8. The anomeric
3
1
1
0 days at a minimum inhibitory concentration (MIC)
P
of 12.5 μg/mL. The observed activity can be regarded
as moderate since the antitubercular drug isoniazid is
active at 0.1 μg/mL under analogous conditions. How-
ever, the following is important. First, arabinofurano-
sides functionalized with a long-chain alkyl phosphate
group, including those containing a farnesyl group as
in phosphate 7, did not inhibit M. Tuberculosis at MIC
values of up to 100 μg/mL [21]. Second, the antituber-
cular activity of glucuronosides 6–8 was similar to the
activity of pyrazinamide (MIC 12.5 μg/mL) [28]. The
fact that phosphates 6 and 7 possessing no nitrogen
atoms showed antitubercular activity at a level com-
parable to the nitrogen-containing compound suggests
a different mechanism (target) of their action. Ob-
viously, this problem requires a separate study.
1
proton resonated at δ 5.72 ppm (J1
-H,2-H
= 7.9, J1-H,P
=
3
.2 Hz), indicating β-orientation of the glycoside bond.
1
The NH signal of 8 appeared in the H NMR spectrum
as a broadened singlet at δ 2.12 ppm.
The acetate protection was removed from the
carbohydrate moieties of 6–8 by treatment with hydra-
zine hydrate according to [6]. This procedure also
ensured simultaneous transformation of the ester group
into more reactive carbohydrazide [6]. The reactions
were carried out in methanol at room temperature for
2
4 h, the precipitate was filtered off, and excess
hydrazine hydrate and the solvent were removed under
reduced pressure. Hydrazides 9 and 10 were isolated in
7
5 and 67% yield, respectively. Their formation was
EXPERIMENTAL
confirmed by the IR spectra which contained absorp-
tion bands due to the amide group at 1667, 1632, 1625,
–
1
The IR spectra (400–4000 cm ) were recorded on
a Bruker Vector 22 spectrometer (Germany) with
Fourier transform from samples prepared as thin films.
–
1
1
536, 1515, 1404, 1380, and 1376 cm ; in addition,
1
their H NMR spectra lacked signals from acetate
protecting groups at δ 1.96–1.99 ppm and methoxy
protons at δ 3.64–3.66 ppm, whereas proton signals of
the carbohydrate ring shifted upfield. The anomeric
proton of 9 and 10 resonated at δ 5.49 and 5.51 ppm,
respectively, as a doublet of doublets with the vicinal
1
13
31
The H, C, and P NMR spectra were measured on
Bruker Avance-400 and Avance-600 spectrometers
(Germany). The mass spectra were obtained on
a Bruker Daltonik AmazonX mass spectrometer
(Bremen, Germany); negative electrospray ionization,
a.m.u. range 100–1500; capillary voltage 4500 V;
drying gas nitrogen, temperature 200°C, flow rate
8 L/min; samples were dissolved in chloroform or
3
3
coupling constants J
= 7.0, J
= 3.41 Hz (9)
1
-H,2-H
1-H,P
3
3
and J1
= 7.0, J
= 3.47 Hz (10) corresponding
-H,2-H
1-H,P
to β-orientation of the glycoside bonds in their mole-
1
–5
cules. The H NMR spectra of 9 and 10 lacked signals
methanol to a concentration of 10 M. The data were
assignable to triethylammonium counterion, which
were present in the spectra of initial phosphates 6
and 7. These data allowed us to presume that hydra-
zides 9 and 10 were isolated as inner salts (betaines;
processed using DataAnalysis 4.0 (Bruker Daltonik).
The optical rotations were measured on a Perkin Elmer
341 polarimeter (USA). The progress of reactions and
the purity of the isolated compounds were monitored
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 53 No. 1 2017

5
4
IZMEST’EV et al.
by TLC on Sorbfil PTSKh-AF-A plates (Krasnodar,
Russia); spots were detected by treatment with
1 mL of water was added, the mixture was allowed to
warm up to room temperature, and 254 mg (1.0 mmol)
of iodine was added. The mixture was stirred for 1 h,
5
% H SO or a solution of H PO ·12 MoO ·H O,
2 4 3 4 3 2
followed by heating. The products were isolated by
column chromatography on silica gel (0.06–0.20 mm;
Acros Organics).
and a 1 M solution of Na S O was added until the
2 2 3
mixture became colorless. The solvent was removed
under reduced pressure, and the product was isolated
by silica gel column chromatography (CH Cl –
2
2
Compound 2 was synthesized according to the
MeOH–Et N, 95:5:1).
3
procedure reported in [22]; mp 177°C (from MeOH),
2
0
[
α] = 8.1° (c = 1.1, CHCl ); published data:
Triethylammonium heptyl (2S,3R,4S,5S,6S)-
3,4,5-tris(acetyloxy)-6-(methoxycarbonyl)oxan-2-yl
phosphate (6). Yield 0.29 g (47%), yellow oily mate-
D
3
2
5
mp 176°C [22]; mp 177–178°C (from EtOH), [α]
=
D
7
.4 (c = 2.0, CHCl ) [29]. Compound 3 was prepared
3
2
0
1
according to [23]; mp 105–106°C (from EtOAc);
published data [29]: mp 106–107°C (from EtOH).
Compound 4 was synthesized as described in [24];
mp 83°C; published data [24]: mp 81°C.
rial, [α]
(400 MHz, CDCl
6.8 Hz), 1.18–1.28 m (8H, CH
NCH CH , J = 7.3 Hz), 1.49–1.61 m (2H, OCH
.96 s (3H, CH CO), 1.97 s (3H, CH CO), 1.99 s (3H,
D
= 50.0° (c = 1.0, CHCl
), δ, ppm: 0.82 t (3H, C H
), 1.31 t (9H,
CH ),
3
). H NMR spectrum
7
′
, J =
3
3
2
2
3
2
2
1
3
3
Triethylammonium (2S,3R,4S,5S,6S)-3,4,5-tris-
CH CO), 3.08 q (6H, NCH , J = 7.3 Hz), 3.66 s (3H,
3
2
(
acetyloxy)-6-(methoxycarbonyl)oxan-2-yl phospho-
CH O), 3.82 q (2H, OCH , J = 6.7 Hz), 4.57 d (1H,
3
2
nate (5). Compound 4, 1 g (3.0 mmol), was dissolved
in 8 mL of anhydrous THF, 3 mL of anhydrous tri-
ethylamine and 0.67 g (3.3 mmol) of 2-chloro-2H,4H-
5
4
-H, J = 10.2 Hz), 4.84–4.91 m (1H, 2-H), 5.11 t (1H,
-H, J = 9.8 Hz), 5.53 t (1H, 3-H, J = 9.8 Hz), 5.74 d.d
1
3
(
(
1H, 1-H, J = 7.8, 3.3 Hz). C NMR spectrum
100 MHz, CDCl ), δ , ppm: 8.56 s (3C, NCH CH ),
1
,3,2-benzodioxaphosphinin-4-one were added, the
3
C
2
3
mixture was stirred for 24 h at room temperature, 3 mL
of water was added, and the mixture was left to stand
for 1 h. The solvent was removed under reduced pres-
sure, and the residue was purified by column chroma-
tography on silica gel using methylene chloride–
methanol–triethylamine (50:1:0.5 to 50:2:0.5) as
eluent. Yield 0.95 g (64%), light yellow oily material,
7
′
1
2
3.92 s (C H ), 20.37 s (CH CO), 20.53 s (CH CO),
3 3 3
0.56 s (CH CO); 22.44 s, 25.58 s, 28.90 s, 31.67 s
3
(
(
2C, CH ); 30.58 d (OCH CH , J = 7.7 Hz), 45.91 s
2 2 2 CP
3C, NCH CH ), 52.54 s (CH O), 66.07 d (OCH ,
2
3
3
2
3
5
4
JCP = 6.2 Hz), 68.79 s (C ), 69.08 s (C ), 69.55 s (C ),
7
1
2
1
0.32 d (C , J = 7.3 Hz), 91.69 d (C , J = 5.1 Hz),
CP CP
2
0
1
68.07 s (COOCH ); 169.47 s, 169.71 s, 169.88 s
[
α] = 33.5° (c = 1.0, CHCl ). H NMR spectrum
3
D
3
3
1
(
CH CO). P NMR spectrum (400 MHz, CDCl ):
(
400 MHz, CDCl ), δ, ppm: 1.30 t (9H, CH CH , J =
3
3
3
2
3
δ –3.1 ppm, s. Mass spectrum, m/z (I , %): 511.3
7
1
7
1
9
1
.4 Hz), 1.93 s (3H, CH CO), 1.94 s (3H, CH CO),
P
rel
3
3
–
–
(
100) [M] . C H O P . Calculated: M 511.2.
.96 s (3H, CH CO), 3.02 q (6H, CH CH , J =
20 32 13
3
2
3
.5 Hz), 3.63 s (3H, CH O), 4.55 d (1H, 5-H, J =
Triethylammonium 3,7,11-trimethyldodeca-
2,6,10-trien-1-yl (2S,3R,4S,5S,6S)-3,4,5-tris(acetyl-
3
0.2 Hz), 4.89–4.94 m (1H, 2-H), 5.09 t (1H, 4-H, J =
.6 Hz), 5.52 t (1H, 3-H, J = 9.7 Hz), 5.77 d.d (1H,
-H, J = 9.1, 3.4 Hz), 6.87 d (1H, PH, J = 640.4 Hz).
C NMR spectrum (100.6 MHz, CDCl ), δ , ppm:
oxy)-6-(methoxycarbonyl)oxan-2-yl phosphate (7).
2
0
Yield 0.6 g (83%), yellow oily material, [α] = 19.2°
D
1
3
1
3
C
(c = 1.0, CHCl ). H NMR spectrum (400 MHz,
3
8
.41 s (3C, CH CH ), 20.27 s (CH CO), 20.43 s (2C,
CDCl ), δ, ppm: 1.37 t (9H, NCH CH , J = 7.4 Hz),
2
3
3
3
2
3
CH CO), 45.64 s (3C, CH CH ), 52.48 s (CH O),
3
2
3
3
1.52 s and 1.60 s (12H, 3′-CH , 7′-CH , 11′-CH ),
3 3 3
3
5
4
2
6
8.89 s (C ), 69.08 s (C ), 69.42 s (C ), 70.07 d (C ,
JCP = 5.5 Hz), 90.79 d (C , J = 4.8 Hz), 167.92 s
1
.85–2.04 m (17H, CH , CH CO), 3.14 q (6H,
2 3
1
CP
NCH CH , J = 7.4 Hz), 3.64 s (3H, CH O), 4.33 t (2H,
2 3 3
(
COOCH ), 169.37 s (CH CO), 169.66 s (2C,
3 3
OCH , J = 6.5 Hz), 4.50 d (1H, 5-H, J = 10.1 Hz),
2
3
1
CH CO). P NMR spectrum (400 MHz, CDCl ):
3
3
4.79–4.84 m (1H, 2-H), 4.97–5.05 m (2H, 6′-H,
10′-H), 5.08 t (1H, 4-H, J = 9.9 Hz), 5.25–5.29 m (1H,
δ 0.9 ppm, s. Mass spectrum: m/z 397.1 (I 100%)
P
rel
–
–
[
M] . C H O P . Calculated: M 397.1.
13 18 12
3-H), 5.49 t (2H, OCH , J = 9.8 Hz), 5.72 d.d (1H,
2
1
3
Phosphates 6 and 7 (general procedure). A solu-
tion of 0.5 g (1.0 mmol) of phosphonate 5 and
.0 mmol of heptan-1-ol or farnesol in 15 mL of anhy-
drous pyridine was cooled to –15°C, and a solution of
08 μL (3.3 mmol) of pivaloyl chloride in 3 mL of
pyridine was added. The mixture was stirred for 1.5 h,
1-H, J = 7.8, 3.2 Hz). C NMR spectrum (100 MHz,
CDCl ), δ , ppm: 8.66 s (3C, NCH CH ); 15.78 s,
16.18 s, 17.48 s (3′-CH , 7′-CH , 11′-CH ); 20.32 s,
20.47 s, 20.51 s (CH CO); 23.17 s, 25.48 s, 25.52 s,
25.97 s, 26.25 s, 26.36 s, 26.53 s, 27.01 s, 31.76 s,
39.37 s, 39.50 s, 39.64 s (CH ); 46.28 s (3C,
3
C
2
3
2
3
3
3
3
4
2
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 53 No. 1 2017

SYNTHESIS AND ANTITUBERCULAR ACTIVITY
55
NCH CH ), 52.62 s (CH O), 62.65 s (OCH ), 68.74 s
Hydrazides 9 and 10 (general procedure). Phos-
phate 6 or 7, 0.2 mmol, was dissolved in 7 mL of
methanol, 6.0 mmol of hydrazine hydrate was added,
and the mixture was stirred for 24 h at room tempera-
ture. The precipitate was filtered off, and the solvent
and excess hydrazine hydrate were removed under
reduced pressure.
2
3
3
2
3
5
4
2
(
(
C ), 68.83 s (C ), 69.34 s (C ), 70.27 s (C ), 91.66 d
1
C , J = 5.1 Hz); 123.50 s, 124.06 s, 124.26 s,
CP
1
1
1
31.12 s, 131.34 s, 135.21 s, 135.32 s, 140.00 s,
40.17 s (CH=C); 167.86 s (COOCH ); 169.33 s,
3
3
1
69.64 s, 169.78 s (CH CO). P NMR spectrum
3
(
(
400 MHz, CDCl ): δ –3.8 ppm, s. Mass spectrum
MALDI): m/z 616.70 (Irel 100%) [M] . C H O P .
28 42 13
3 P
–
–
(
2S,3R,4S,5S,6S)-6-(Diazan-2-ium-1-carbonyl)-
,4,5-trihydroxyoxan-2-yl heptyl phosphate (9).
Yield 0.13 g (75%), light yellow oily material. IR
Calculated: M 617.24.
3
Triethylammonium (2S,3R,4S,5S,6S)-3,4,5-tris-
acetyloxy)-6-(methoxycarbonyl)oxan-2-yl N-un-
–
1
(
spectrum, ν, cm : 3292 br (NH , OH), 1667, 1625,
1
2
1
decylphosphoramidate (8). A solution of 0.5 g
1.0 mmol) of phosphonate 5 in 10 mL of anhydrous
536, 1376 (HNC=O), 1219, 1073 (P=O). H NMR
(
spectrum (600 MHz, CD OD), δ, ppm: 0.89 t (3H,
3
methylene chloride was cooled to 0°C, and 1 mL of
anhydrous triethylamine and 0.38 mL (3.0 mmol) of
chloro(trimethyl)silane were added. The mixture was
stirred for 30 min, 0.25 g (1.0 mmol) of crystalline
iodine was added, the mixture was stirred for 20 min,
7′
C H , J = 6.8 Hz), 1.24–1.41 m (8H, CH ), 1.56–
3
2
1
.65 m (2H, OCH CH ), 3.45–3.50 m (1H, 2-H), 3.54 t
2 2
(
1H, 3-H, J = 9.8 Hz), 3.71 t (1H, 4-H, J = 9.3 Hz),
3
9
.89 q (2H, OCH , J = 6.5 Hz), 4.20 d (1H, 5-H, J =
2
.9 Hz), 5.49 d.d (1H, 1-H, J = 7.0, 3.41 Hz), 7.88 s
0
.24 mL (3.0 mmol) of anhydrous pyridine was added,
the mixture was stirred for 5 min, and 0.22 g
1.3 mmol) of undecylamine was added. The mixture
31
(
1H, NH). P NMR spectrum (400 MHz, CD OD):
3
δ –1.2 ppm, s. Mass spectrum: m/z 385.2 (I 100%)
P
rel
(
–
–
[
M] . C H N O P . Calculated: M 385.1.
13 26 2 9
was allowed to warm up to room temperature and
stirred for 24 h, the solvent was removed under
reduced pressure, the residue was dissolved in methy-
lene chloride (50 mL), and the solution was washed
(
2S,3R,4S,5S,6S)-6-(Diazan-2-ium-1-carbonyl)-
3
,4,5-trihydroxyoxan-2-yl 3,7,11-trimethyldodeca-
2,6,10-trien-1-yl phosphate (10). Yield 0.11 g (67%),
–
1
with water (50 mL) and dried over MgSO . The
light yellow oily material. IR spectrum, ν, cm :
3443 br (NH , OH), 1632, 1515, 1404, 1380
[C(O)NH], 1219, 1096 (P=O). H NMR spectrum
(400 MHz, CD OD), δ, ppm: 1.57–1.61 m (6H, CH ),
1.64–1.68 m (6H, CH ), 1.92–2.12 m (8H, CH ), 3.48–
4
solvent was removed, and the residue was purified by
silica gel column chromatography using methylene
chloride–methanol (10:1 to 5:1 with addition of
2
1
3
3
1
vol % of triethylamine) as eluent. Yield 0.25 g
3
2
2
0
(
37%), yellow oily material, [α] = 42.8 (c = 1.0,
3.53 m (1H, 2-H), 3.56 t (1H, 3-H, J = 9.5 Hz), 3.72 t
(1H, 4-H, J = 9.35 Hz), 4.21 d (1H, 5-H, J = 9.9 Hz),
D
1
CHCl ). H NMR spectrum (600 MHz, CDCl ), δ,
ppm: 0.83 t (3H, C H , J = 6.8 Hz), 1.16–1.24 m
3
3
11′
3
4.46 t (2H, OCH , J = 6.5 Hz), 5.05–5.14 m (2H, 6′-H,
2
(
16H, CH ), 1.29 t (9H, NCH CH , J = 7.4 Hz), 1.36–
2 2 3
10′-H), 5.36–5.41 m (1H, 2′-H), 5.51 d.d (1H, 1-H, J =
1
.44 m (2H, NHCH CH ), 1.96 s (3H, CH CO), 1.97 s
2 2 3
7.0, 3.47 Hz); 8.10–8.16 m, 8.58–8.63 m, 8.93–8.97 m
3
1
(
3H, CH CO), 1.99 s (3H, CH CO), 2.12 br.s (1H,
3
3
(3H, NH). P NMR spectrum (400 MHz, CD OD):
3
NH), 2.80 q (2H, NHCH , J = 7.6 Hz), 3.04 q (6H,
2
δ –1.0 ppm, s. Mass spectrum, m/z 491.3 (I 100%):
P
rel
–
–
NCH CH , J = 7.4 Hz), 3.66 s (3H, CH O), 4.61 d
2
3
3
[M] . C H N O P . Calculated: M 491.2.
21 36 2 9
(
(
5
1H, 5-H, J = 10.5 Hz), 4.86–4.91 m (1H, 2-H), 5.12 t
1H, 4-H, J = 9.8 Hz), 5.55 t (1H, 3-H, J = 9.7 Hz),
.72 d.d (1H, 1-H, J = 7.9, 3.2 Hz). C NMR spectrum
The antiturbercular activity of compounds 6–8 was
assayed by the vertical diffusion method on a Novaya
solid nutrition medium inoculated with Mycobacterium
tuberculosis H37Rv. Nutrition medium was placed into
test-tubes in 5-mL portions, inoculated with 0.1 mL of
a suspension of different mycobacteria strains diluted
to a turbidity value of 10 GKI units, and incubated for
1
3
(
150 MHz, CDCl ), δ , ppm: 8.42 s (3C, NCH CH ),
3 C 2 3
1
1′
1
2
2
7
5
3.96 s (C ); 20.39 s, 20.57 s, 20.60 s (CH CO);
3
2.54 s, 26.86 s, 29.21 s, 29.37 s, 29.51 s, 29.52 s,
9.57 s, 31.78 s (CH ); 32.09 d (NHCH CH , J =
2
2
2
CP
.2 Hz), 41.96 s (NHCH ), 45.47 s (3C, NCH CH ),
2
2
3
3
5
2
4 h at a constant temperature to grow Mycobacterium
2.49 s (CH O), 68.67 s (C ), 69.44 s (C ), 69.74 s
C ), 70.51 d (C , J = 7.2 Hz), 91.27 d (C , J
3
4
2
1
tuberculosis (MBT). After 24 h, the test tubes were
placed vertically, and 0.3 mL of a solution of 6–8 in
aqueous ethanol with a concentration of 12.5, 6.2, 3.1,
1.5, 0.7, 0.35, and 0.1 μg/mL was added to each test
tube. The test tubes were then placed in a thermostat
(
=
CP
CP
5
1
.3 Hz), 168.37 s (COOCH ); 169.52 s, 169.85 s,
3
3
1
69.90 s (CH CO). P NMR spectrum (400 MHz,
3
CDCl ): δ 5.8 ppm, s. Mass spectrum: m/z 566.4
3
P
–
–
(
I 100%) [M] . C H NO P . Calculated: M 566.2.
rel 24 41 12
RUSSIAN JOURNAL OF ORGANIC CHEMISTRY Vol. 53 No. 1 2017

5
6
IZMEST’EV et al.
and were incubated under sterile conditions at 37°C for
0 days. The growth of MBT was evaluated according
12. Greimel, P., Lapeyre, M., Nagatsuka, Y., Hiraba-
yashi, Y., and Ito, Y., Bioorg. Med. Chem., 2008, vol. 16,
p. 7210.
1
to the standard procedure. An MBT growth inhibition
zone of larger than 10 mm indicated tuberculostatic
activity. The size of the inhibition zone (mm) is pro-
portional to the tuberculostatic activity. An inhibition
zone of 100 mm and larger was considered as com-
plete inhibition of MBT growth. Isoniazid was used as
control drug.
1
3. Chen, C., Liu, B., Xu, Y., Utkina, N., Zhou, D., Dani-
lov, L., Torgov, V., Veselovsky, V., and Feng, L.,
Carbohydr. Res., 2016, vol. 430, p. 36.
1
1
4. Brockhausen, I., Larsson, E.A., and Hindsgaul, O.,
Bioorg. Med. Chem. Lett., 2008, vol. 18, p. 804.
5. Vinnikova, A.N., Torgov, V.I., Utkina, N.S., Veselov-
sky, V.V., Druzhinina, T.N., Wang, S., Brockhausen, I.,
and Danilov, L.L., Russ. J. Bioorg. Chem., 2015,
vol. 41, p. 105.
This study was performed under financial support
by the Russian Science Foundation (project no. 14-50-
0
0014).
16. Vinnikova, A.N., Demirova, K.A., Druzhinina, T.N., and
Veselovsky, V.V., Russ. Chem. Bull., Int. Ed., 2015,
vol. 64, p. 202.
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