K.J. Kayser-Bricker et al. / Tetrahedron 64 (2008) 7015–7020
7019
(Na2SO4), filtered, and concentrated in vacuo to afford a yellow oil.
This crude material was then purified by column chromatography
(0–45% ethyl acetate/hexanes) to afford 14 as a clear thick oil
(170 mg, 75%). 1H NMR (CDCl3, 500 MHz) d 7.28–7.12 (m, 30H), 5.04
(ABq, J¼12.1 Hz, 2H), 4.93–4.86 (m, 4H), 4.70 (ABq, J¼11.2 Hz, 2H),
4.64–4.56 (m, 5H), 4.23–4.20 (m, 2H), 3.83 (td, J¼9.5 and 2.3 Hz,
2H), 3.47 (td, J¼9.2 and 2.3 Hz, 1H), 2.37 (d, J¼2.4 Hz, 1H); 13C NMR
(CDCl3, 500 MHz) d 154.4, 138.3, 138.2, 138.1, 135.6, 135.6, 135.1,
128.6, 128.6, 128.5, 128.5, 128.5, 128.4, 128.3, 128.2, 127.9, 127.8,
127.8, 127.7, 127.7, 127.5, 79.6, 79.6, 78.9, 77.8, 77.8, 75.3, 75.2, 75.2,
74.4, 74.4, 69.8, 69.5, 69.5, 69.4, 69.3; 31P NMR (CDCl3, 202 Hz)
d ꢂ1.7; IR (film, cmꢂ1) 3387, 3089, 3064, 3032, 2946, 2880, 1744,
1495, 1450, 1262, 1013; TLC Rf 0.41 (40% ethyl acetate/hexanes);
exact mass calcd for [C49H49O11P]þ requires m/z 845.3085, found
845.3064 (ESIþ); [a]D þ26.4 (c 1.0, CHCl3).
1.23–1.15 (m, 16H), 0.82–0.77 (m, 6H); 13C NMR (CDCl3, 500 MHz)
d 173.0,172.7,172.6,154.2,154.2,138.0,138.0,137.9,137.8,135.9,135.9,
135.8,135.8,135.4,135.4,135.3,135.3,134.9,134.9,128.6,128.6,128.3,
128.3, 128.2, 128.1, 128.1, 127.9, 127.8, 127.8, 127.7, 127.7, 127.6, 127.6,
127.5, 127.5, 127.4, 127.3, 79.9, 79.8, 78.2, 78.2, 78.2, 78.1, 77.6, 77.5,
77.5, 77.5, 77.2, 76.6, 76.5, 76.3, 76.2, 75.4, 75.3, 74.7, 74.7, 69.9, 69.9,
69.8, 69.8, 69.6, 69.6, 69.3, 69.3, 69.2, 69.2, 69.2, 69.1, 65.8, 65.7, 65.6,
65.5, 61.4, 61.4, 34.0, 34.0, 33.9, 31.6, 29.0, 29.0, 28.9, 24.8, 24.7, 22.6,
14.0; 31P NMR (CDCl3, 202 Hz) d ꢂ1.6, ꢂ1.9, ꢂ2.0; IR (film, cmꢂ1); TLC
Rf 0.37 (30% ethyl acetate/hexanes); exact mass calcd for
[C75H91O18P2]þ requires m/z 1341.5675, found 1341.5661 (ESIþ); [a]D
þ8.8 (c 1.0, CHCl3).
4.2.4. PI5P–DiC8
Compound 16 (0.095 g, 0.071 mmol) was dissolved in t-BuOH/
H2O (5:1) (6 mL) and Chelex (Na form) resin was added till a thick
slurry. Palladium hydroxide on carbon (0.190 g) was added and the
chamber evacuated. The solution was placed under 1 atm of H2 and
stirred for 18 h. The reaction mixture was filtered over a Celite pad
and the Celite was washed with ethanol (20 mL), ethanol/water
(1:1) (15 mL), and H2O (15 mL). The filtrate was filtered through
a 0.22 mm filter and concentrated under reduced pressure (no
heating). The resulting solid was dissolved in a minimal amount of
water and run through a Sephadex G-10 size exclusion column
eluting with water. Fractions containing pure product were com-
bined and lyophilized. The solid was dissolved in a minimal amount
of waterand run through a Chelex (Na form) ion exchange column to
fully form the desired sodium salt. Fractions containing the desired
product were combined and lyophilized to yield the sodium salt of
PI5P–DiC8 as a white fluffy solid (0.032 g, 61% yield). 1H NMR (D2O,
500 MHz) d 5.32 (q, J¼7.7 Hz, 1H), 4.43 (dd, J¼12.2 and 1.5 Hz, 1H),
4.27 (dd, J¼12.1 and 8.2 Hz,1H), 4.22 (t, J¼2.6 Hz,1H), 4.08–4.03 (m,
3H), 3.93–3.85 (m, 2H), 3.80 (t, J¼9.3 Hz, 1H), 3.62 (dd, J¼10.0 and
2.4 Hz,1H), 2.48–2.30 (m, 4H),1.64–1.58 (m, 4H),1.37–1.25 (m,16H),
0.89–0.85 (m, 6H); 13C NMR (D2O, 500 MHz) d 175.8,175.7, 79.7, 79.7,
76.6, 76.5, 72.5, 71.7, 71.5, 71.5, 71.2, 63.9, 34.8, 34.8, 32.4, 29.7, 29.6,
29.6, 29.5, 25.5, 25.4, 23.2, 23.2, 14.5, 14.4; 31P NMR (D2O, 202 Hz)
d 4.3, ꢂ0.6; exact mass calcd for [C25H49O16P2]þ requires m/z
667.2496, found 667.2476 (ESIþ); [a]D þ2.75 (c 2.0, H2O, pH 9).
4.2.2. Compound 15
To a stirred solution of 14 (0.185 g, 0.219 mmol) in 14 mL of
acetone (reagent grade) was added LiBr (0.035 g, 0.408 mmol), and
the reaction mixture was refluxed for 10 h. It was then cooled to
room temperature and concentrated. The residue was purified by
silica gel chromatography eluting with 50% EtOAc/hexanes to 15%
CH3OH/CH2Cl2 to give the crude product as a lithium salt. The salt
was dissolved in a minimal amount of CH3OH and run through a Hþ
DOWEX 50ꢁ2-200 ion exchange column. Fractions containing the
desired product were combined and concentrated to give the crude
phosphoric product. The crude residue was then dissolved in THF
(1.1 mL), and diacylglycerol 8 (0.151 g, 0.437 mmol) and triphenyl-
phosphine (0.115 g, 0.437 mmol) were added and the reaction was
cooled to 0 ꢀC. DEAD (68 mL, 0.437 mmol) was then added and the
reaction mixture was stirred at 0 ꢀC under N2 for 48 h. The mixture
was then concentrated under reduced pressure and purified by
silica gel chromatography eluting with 0–40% EtOAc/hexanes to
yield 15 as a colorless thick oil (0.165 g, 74%, over two steps). 1H
NMR (CDCl3, 400 MHz) d 7.29–7.15 (m, 25H), 5.07–4.88 (m, 5H),
4.76–4.57 (m, 7H), 4.28–3.81 (m, 8H), 3.50 (m, 1H), 2.48 (s, 1H),
2.17–2.10 (m, 4H), 1.48–1.46 (m, 4H), 1.18–1.15 (m, 16H), 0.79–0.77
(m, 6H); 13C NMR (CDCl3, 400 MHz) d 173.0, 173.0, 172.6, 154.3,
154.3, 138.2, 138.2, 138.1, 138.0, 137.9, 135.4, 135.4, 135.4, 135.3,
135.0, 135.0, 128.7, 128.5, 128.3, 128.2, 128.2, 128.2, 128.0, 127.9,
127.8, 127.8, 127.7, 127.7, 127.6, 127.5, 127.5, 79.5, 79.4, 79.4, 78.9,
78.9, 77.9, 77.9, 77.8, 77.8, 77.8, 77.2, 76.8, 75.3, 75.2, 75.1, 74.4, 69.8,
69.7, 69.6, 69.5, 69.5, 69.2, 69.1, 65.7, 65.6, 65.4, 65.4, 61.5, 61.4, 34.0,
34.0, 33.9, 31.6, 29.0, 28.9, 28.8, 24.7, 24.7, 22.5, 14.0; 31P NMR
(CDCl3, 162 Hz) d ꢂ1.7, ꢂ1.8; IR (film, cmꢂ1) 3026, 2951, 2925, 2849,
1738, 1456, 1260, 1158, 1113, 1022; TLC Rf 0.21 (30% ethyl acetate/
hexanes); exact mass calcd for [C61H78O15P]þ requires m/z
1081.5073, found 1081.5035 (ESIþ); [a]D þ11.6 (c 1.0, CHCl3).
4.2.5. ent-PI5P–DiC8
Synthesis and spectral data were identical to PI5P–DiC8. [a]D
ꢂ8.75 (c 2.0, H2O, pH 8).24
Acknowledgements
We thank the National Institutes of General Medical Sciences of
the National Institutes of Health for support (GM-068649).
4.2.3. Protected PI5P–DiC8 (16)
To a stirred solution of 15 (0.100 mg, 0.092 mmol) in CH2Cl2
(18.5 mL) was added dibenzyl diisopropylphosphoramidite (311 mL,
0.92 mmol) followed by 4,5-dicyanoimidazole (0.131 g,1.109 mmol).
The reaction was stirred at room temperature under a nitrogen
atmosphere for 15 h. The reaction was then cooled to 0 ꢀC and 30%
H2O2 (8.6 mL) was added. The reaction was stirred at 0 ꢀC for 1 h at
which time the reaction was quenched with saturated Na2SO3
(w70 mL) until no peroxides were detected via starch paper. The
reaction was then extracted with DCM (3ꢁ50 mL) and the organic
layers were combined, dried (Na2SO4), filtered, and concentrated
under reduced pressure. The resulting residue was then purified by
silica gel column chromatography (0–38% ethyl acetate/hexanes)
and a second column (70% diethyl ether/hexanes) to afford pure 16 as
a thick oil (0.105 g, 85%). 1H NMR (CDCl3, 500 MHz) d 7.315–7.32 (m,
31H), 6.98–6.94 (m, 4H), 4.98 (s, 2H), 4.94–4.61 (m, 14H), 4.45–4.15
(m, 3H), 4.04–3.72 (m, 6H), 2.17–2.09 (m, 4H), 1.51–1.44 (m, 4H),
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