82
BOLDYREV, MOLOTKOVSKY
and evaporated to dryness. The residue was extracted CH2), 1.58 (6 H, br. m, 3 × CH2), 2.28 (4 H, br. m, 2 ×
with ethyl acetate, and the extract was dried with CH2), 2.41 (6 H, s, 2 × CCH3), 2.45 (6 H, s, 2 × CCH3),
Na2SO4 and evaporated. The individual according to 2.95 (2 H, br. m, CCH2), 3.19 (9 H, s, 3 × NCH3), 3.64
the TLC amorphous Me4-BODIPY-8-alkanoic acids (2 H, m, CH2N), 4.03 (2 H, br. m, CH2OOC), 4.14 (1 H,
were obtained in yields of 85–95%. They were further br. m, CH2OP), 5.22 (1 H, br. m, CHOOC), and 6.06
used without additional purification.
(2 H, s, arom. H).
N-[7-(4,4-Difluoro-1,3,5,7-tetramethyl-4-bora-
3a,4a-diaza-s-indacene-8-yl)heptanoyl]sphingosine-
1-phosphocholine (XIX). BOP (5 mg) was added to a
solution of Me4-BODIPY-heptanoic acid (XII)
(3.5 mg, 9 µmol) and triethylamine (5 µl) in dry dichlo-
romethane (0.5 mg) and isopropanol (1 ml), and the
mixture was stirred for 1 h at 20°ë. Then, sphingosine-
1-phosphocholine (XVIII) (7 mg, 9 µmol), dichlo-
romethane (0.5 ml), and isopropanol (1 ml) were added
to the mixture. After stirring overnight in argon atmo-
sphere, the mixture was chromatographed on a silica gel
column eluted within chloroform–methanol (1 : 0
1 : 1) to yield sphingomyelin (XIX) (5 mg, 65%) as an
orange glassy solid, Rf 0.40 in 65 : 35 : 8 chloroform–
methanol–4 N NH4OH ammonia (detection with A, B,
and C); the chromatographic mobility is identical to
that of sphingomyelin from bovine brain; MALDI MS,
1-Acyl-2-[w-(4,4-difluoro-1,3,5,7-tetramethyl-4-
bora-3a,4a-diaza-s-indacene-8-yl)acyl]-sn-glycero-3-
phosphocholines (XV)–(XVII). Egg yolk lysophos-
phatidylcholine (10 mg, ~20 µmol), acid (XI)–(XIII)
(50 µmol), 4-aminopyridin (9 mg, 95 µmol), and 4-pyr-
rolidinopyridin (14 mg, 95 µmol) were dissolved in dry
chloroform (3 ml) under vigorous stirring in argon
atmosphere. DCC (20 mg, 95 µmol) was then added as
30% solution in CCl4, and the mixture was stirred for
~1 h and kept overnight at 20°ë. The mixture was chro-
matographed on a silica gel column eluted with CHCl3–
MeOH (from 5 : 1 to 1 : 1 ratio), and phosphatidylcho-
lines (XV)–(XVII) were isolated as glassy orange-red
solids in 20–25% yields; Rf 0.35–0.4 in 65 : 25 : 4
CHCl3–MeOH–water, detection with A, B, and C; the
chromatographic mobility is identical to that of egg
yolk phosphatidylcholine. The following compounds
were obtained:
1
m/z: 823.87 [M + H]+, 803.81 [M – F]+; H NMR
(CDCl3–CD3OD 2 : 1): 0.84 (3 H, t, CH2CH3), 1.22 (22
H, br. m, 11 × CH2), 1.37 (2 H, br. m, CH2), 1.51 (2 H,
br. m, CH2), 1.62 (4 H, br. m, 2 × CH2), 1.98 (2 H, br.
m, CH2), 2.17 (2 H, br. m, CH2), 2.41 (6 H, s, 2 ×
ëCH3), 2.45 (6 H, s, 2 × ëCH3), 2.96 (2 H, br. m,
ëCH2), 3.17 (9 H, s, 3 × NCH3), 3.59 (4 H, br. m, CH
and CH2CH2N), 4.27 (2 H, m, CHCH2OP), 5.41 (1 H,
br. m, CH=), 5.68 (1 H, br. m, CH=), and 6.06 (2 H, s,
arom. H).
N-[7-(4,4-Difluoro-1,3,5,7-tetramethyl-4-bora-
3a,4a-diaza-s-indacene-8-yl)heptanoyl]-1-O-b-D-gal-
actosyl sphingosine (XXI). BOP (8 mg, 18 µmol) was
added to a solution of Me4-BODIPY-heptanoic acid
(XII) (6.3 mg, 16,7 µmol) and 1-methylimidazole
(10 µl) in dry CH2Cl2 (2 ml), and the mixture was
stirred for 1.5 h. Then, 1-O-β-D-galactosyl sphingosine
(XX) (8 mg, 17.3 µmol) was added, and stirring was
continued overnight in argon atmosphere. Chromatog-
raphy of the mixture on a silica gel column eluted with
chloroform containing 5–20% of methanol yielded
7.6 mg (55%) of lipid (XXI) as an amorphous orange-
red solid, Rf 0.6 in 78 : 20 : 3 chlorophorm–MeOH–
AcOH; the chromatographic mobility was identical to
that of natural galactosyl ceramide.; UV (ethanol):
498 nm (ε 8.1 × 104 M–1 cm–1); MALDI MS, m/z: 820.4
[M + H]+, 800.42 [M – F]+; 1H NMR (CDCl3–CD3OD
4 : 1): 0.81 (3 H, t, CH2CH3), 1.18 (20 H, br. m, 5 ×
1-Acyl-2-[5-(4,4-difluoro-1,3,5,7-tetramethyl-4-
bora-3a,4a-diaza-s-indacene-8-yl)pentanoyl]-sn-gly-
cero-3-phosphocholine (XV): MALDI MS, m/z:
826.44 [M + H]+, 806.36 [M – F]+ (the major peaks are
1
presented, where acyl is palmitoyl; H NMR (CDCl3–
CD3OD 2 : 1): 0.84 (3 H, t, CH2CH3), 1.22 (24 H, br. m,
CH2), 1.52 (2 H, m, CH2), 1.64 (2 H, m, CH2), 1.79 (2
H, m, CH2), 2.22 (2 H, t, CH2), 2.31 (2 H, t, 12 × CH2),
2.41 (6 H, s, CCH3), 2.45 (6 H, s, CCH3), 2.97 (2 H, t,
CCH2), 3.16 (9 H, s, 3 × NCH3), 3.61 (2 H, m, CH2N),
4.04 (2 H, br. m, CH2OOC), 4.10 (1 H, m, CH2OP),
4.27 (2 H, br. m, CH2CH2N), 4.34 (1 H, m, CH2OP),
5.21 (1 H, m, CHOOC), and 6.05 (2 H, s, arom. H).
1-Acyl-2-[7-(4,4-difluoro-1,3,5,7-tetramethyl-4-
bora-3a,4a-diaza-s-indacene-8-yl)heptanoyl]-sn-gly-
cero-3-phosphocholine (XVI): MALDI MS, m/z:
854.44 [M + H]+, 834.4 [M – F]+ (the major peaks are
presented, where acyl is palmitoyl); 1H NMR (CDCl3–
CD3OD 2 : 1): 0.83 (3 H, t, CH2CH3), 1.21 (26 H, br. m,
13 × CH2), 1.37 (2 H, m, CH2), 1.49 (2 H, m, CH2), 1.54
(2 H, m, CH2), 1.61 (4 H, m, 2 × CH2), 2.26 (2 H, t,
CH2), 2.31 (2 H, t, CH2), 2.39 (6 H, s, 2 × CCH3), 2.45
(6 H, s, 2 × CCH3), 2.94 (2 H, t, CCH2), 3.17 (9 H, s, 3
× NCH3), 3.59 (2 H, m, CH2N), 4.01 (2 H, br. m,
CH2OOC), 4.11 (1 H, m, CH2OP), 4.26 (2 H, br. m,
CH2CH2N), 4.35 (1 H, m, CH2OP), 5.19 (1 H, m,
CHOOC), and 6.18 (2 H, s, arom. H).
1-Acyl-2-[9-(4,4-difluoro-1,3,5,7-tetramethyl-4- CH2), 1.26 (2 H, br. m, CH2), 1.32 (2 H, br. m, CH2),
bora-3a,4a-diaza-s-indacene-8-yl)nonanoyl]-sn-gly- 1.46 (2 H, br. n, CH2), 1.57 (4 H, br. m, 2 × CH2), 1.94
cero-3-phosphocholine (XVII): MALDI MS, m/z: CH2CH=), 2.13 (2 H, t, CH2), 2.36 (6 H, s, 2 × ëCH3),
882.48 [M + H]+, 862.43 [M – F]+ (the major peaks are 2.43 (6 H, s, 2 × ëCH3), 2.89 (2 H, t, ëCH2), 3.45 (1 H,
presented, where acyl is palmitoyl); 1H NMR (CDCl3– br. m, CH, Gal), 3.49 (1 H, br. m, CH, Gal), 3.51 (1 H,
CD3OD 2 : 1): 0.84 (3 H, t, CH2CH3), 1.23 (28 H, br. m, br. m, Gal H6), 3.53 (1 H, br. m, Gal H6), 3.72 and 3.75
14 × CH2), 1.32 (4 H, br. m, 2 × CH2), 1.48 (2 H, br. m, (2 H, two br. m, C–OCH2), 3.85 (1 H, s, CH, Gal), 3.93
RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY Vol. 32 No. 1 2006