Organic & Biomolecular Chemistry
Paper
spirit–TmsCl (9 : 1) under nitrogen; a single signal at δP −19.0 HRMS (ESI+) m/z found [M
+
Na]+
=
1,122.7013,
confirmed complete exchange of the cyanoethyl phosphate C62H102NO13PNa requires 1,122.6987.
esters. The filtrate was evaporated to dryness and the 1-O-[(Cyanoethyloxy)(sn-1-O-stearoyl-2-O-arachidonoylglyceryl-
residue stirred in 1 M methanolic ammonia (5 mL) for 20 min. oxy)phosphoryl]-4-O-(dicyanoethyloxyphosphoryl)-2,3:5,6-O-
The solvents were again evaporated in vacuo, and the residue dicyclohexylidene-myo-inositol, 24d. 1-O-[(Cyanoethyloxy)(sn-
taken up in acetic acid–water (2 : 3, 3 mL). After stirring for 1-O-stearoyl-2-O-arachidonoylglycerol)phosphoryl]-2,3:5,6-O-
24 h, the solution was diluted with water and freeze-dried to dicyclohexylidene-myo-inositol (23d, 225 mg, 0.2 mmol) was
give a white solid (60 mg, 102% assuming mono-ammonium evaporated from pyridine (3 × 2 mL) then dissolved in
salt). δH (400 MHz, D2O) 4.13 (1H, t, J 2.7, Ins 2-CH), 3.82 (1H, CH2Cl2–pyridine (3 : 2, 2.5 mL) to which N-methyl imidazole
dt, J 2.7, 9.1, Ins 1-CH), 3.63 (1H, t, J 9.6, Ins 6-CH), 3.53 (1H, (81 µL, 1.00 mmol, 5.0 eq.) and 0.34 M dicyanoethylpho-
t, J 9.6, Ins 4-CH), 3.45 (1H, dd, J 2.8, 10.0, Ins 3-CH), 3.22 (1H, sphorochloridite in CH2Cl2 (15, 2.36 mL, 0.80 mmol, 4.0
t, J 9.3, Ins 5-H); δP (162 MHz, D2O) 0.43; δC (125 MHz, D2O) eq.) were added. After 16 h cyanoethanol (41 µL, 0.67 mmol,
75.2 (d, J 4.2), 74.0, 72.2, 71.7 (d, J 4.2), 71.3, 70.7; HRMS 3.1 eq.) was added and the mixture stirred for 30 min. The
(ESI−) m/z found [M − H]− = 259.0210, C6H12O9P requires solution was then cooled to 0 °C and 5 M tert-butyl hydro-
259.0219.
peroxide in decanes (200 µL, 1.00 mmol, 5.0 eq.) was added.
1-O-[(Cyanoethyloxy)(sn-1-O-stearoyl-2-O-arachidonoylglycer- After 12 h water (5 mL) was added and the solution was con-
yloxy)phosphoryl]-2,3:5,6-O-dicyclohexylidene-myo-inositol, centrated in vacuo. The resulting mixture was suspended in
23d. To
1-O-(dicyanoethyloxyphosphoryl)-2,3:5,6-O-dicyclo- MeCN–water (1 : 9, 100 mL) and fractionated through a
hexylidene-myo-inositol (11, 203 mg, 0.354 mmol) were added column of silanised silica, eluting with a gradient of MeCN–
CH2Cl2 (3 mL), MeCN (1 mL) and triethylamine (3 mL). After water (1 : 4 → 7 : 3, and flushed with ethyl acetate). The
stirring the solution for 36 h, the solvent was removed in vacuo appropriate fractions were combined, dried (MgSO4), and
to give the putative phosphodiester salt as a white solid adsorbed onto flash silica. This was poured onto a column
(169 mg). To this were added sn-1-O-stearoyl-2-O-arachidonoyl- of silica and fractionated, eluting with a gradient of first
glycerol (19d, 683 mg, 1.06 mmol) and 3-nitro triazole (22, diethyl ether–pet. spirit (0 : 1 → 1 : 0) then methanol–ethyl
323 mg, 2.83 mmol, 8.0 eq.), and the mixture was co-evapor- acetate (0 : 1 → 1 : 1), to afford the title compound as a white
ated from pyridine (3 × 2 mL). The residue was dissolved in greasy solid (219 mg, 85%). Rf (EtOAc) 0.26; δH (400 MHz,
CH2Cl2–MeCN–pyridine (2 : 2 : 1, 5 mL) and a solution of mesi- CDCl3) 5.39–5.28 (8H, m, 4 × HCvCH), 5.28–5.20 (1H, m, Glyc
tylene sulfonyl chloride (309 mg, 1.41 mmol, 4.0 eq.) in pyri- 2-CH), 4.76–4.69 (1H, m, Ins 1-CH), 4.64–4.56 (2H, m, Ins 4-CH
dine (1 mL) was added dropwise over 25 min. The reaction + Ins 2-CH), 4.38–4.00 [10H, m, (3 × OCH2CH2CN) + Glyc
mixture was stirred for a further 2 h after which water (2 mL) 3-CH2 + Ins 3-CH + Ins 6-CH], 3.46 (1H, t, J 10.4, Ins 5-CH),
was added. The mixture was diluted with ethyl acetate 2.75 [12H, m, (3 × CH2CN) + (3 × (CHvCH)2CH2)], 2.32 (2H, t,
(100 mL), washed with water (3 × 300 mL), dried (MgSO4) and J 7.5, CH2CO2), 2.27 (2H, t, J 7.5 Hz, CH2CO2), 2.09 (2H, q, J
flash silica was added before stripping off the solvent. The 7.0, CH2CH2CHvCH), 2.04 (2H, q, J 7.0, CH2CH2CHvCH),
silica was poured onto a flash column that was eluted with a 1.70–1.17 (68H, m, 34 × CH2), 0.83 (6H, 2 × t, J 7.0, 2 × CH3); δP
gradient of EtOAc–pet. spirit (0 : 1 → 1 : 0) to afford the title (202 MHz, CDCl3) −2.4 (1P, 4-P), −2.83 (0.5P, 1-P), −3.0 (0.5P,
compound as a white solid (351 mg, 90%). Rf (EtOAc) 0.90; δH 4-P); δC (125 MHz, CDCl3) 173.1, 172.4 (2C, CvO), 130.4, 128.9,
(400 MHz, CDCl3) 5.45–5.36 (8H, m, 4 × HCvCH), 5.34–5.26 128.58, 128.50, 128.2, 127.93, 127.69, 127.40 (4 × HCvCH),
(1H, m, Gly 2-CH), 4.80–4.72 (1H, m, Ins 1-CH), 4.61–4.58 (1H, 116.18, 116.08, 116.00 (3 × CuN), 114.3, 111.9 (2 × acetal C),
m, Ins 2-CH), 4.41–4.25 [6H, m, (2 × POCH2) + Gly 1-CH2], 80.8 (d, Ins 4-CH, JC–P 5.0), 79.2, 75.97, 75.63, 75.0, 74.5 (5 ×
4.23–4.15 (1H, m, Ins 3-CH), 4.08 (1H, t, J 6.0, Ins 3-CH), Ins-CH), 69.2 (Gly 2-CH), 66.1 (Gly 3-CH2), 62.5–62.0 (3 × P–O–
4.06–4.01 (2H, m, Gly 3-CH2), 3.91 (1H, dd, J 10.5, 6.5, Ins CH2), 61.5–61.3 (m, Gly 1-CH2), 37.4, 36.1 (2 × CH2CO2),
4-CH), 3.39 (1H, t, J 10.0, Ins 6-CH), 2.85 [8H, m, CH2CN + (3 × 35.0, 33.8, 33.4, 31.8, 31.4, 30.3, 29.58 (7C), 29.54 (3C),
(CHvCH)2CH2)], 2.38 (2H, t, J 7.5, CH2CO2), 2.33 (2H, t, J 7.5, 29.38, 29.24 (2C), 29.20, 29.18, 29.01, 27.1, 26.3, 25.5, 24.71,
CH2CO2), 2.14 (2H, q, J 7.0, CH2CH2CHvCH), 2.07 (2H, q, J 24.59, 23.88, 23.81, 23.74, 23.68, 23.63, 22.57, 22.45, 19.44,
7.0, CH2CH2CHvCH), 1.74–1.25 (64H, m, 32 × CH2), 0.90 (6H, 19.38, 19.32 [(24 × fatty acid CH2,) + (10 × cyclohexylidene
2 × t, J 7.0, 2 × CH3); δP (162 MHz, CDCl3) −2.81 (0.5P), −2.94 CH2,) + (3 × CH2CN)], 14.07, 14.01 (2 × CH3,); HRMS (ESI+) m/z
(0.5P); δC (125 MHz, CDCl3) 173.3, 172.6 (2 × CvO), 130.5, found [M
+ = 1,286.7357, C68H110N3O16P2 requires
H]+
129.1, 128.7, 128.6, 128.34, 128.07, 127.8, 127.5 (4 × HCvCH), 1,286.7361.
116.4 (0.5C), 116.1 (0.5C) (CuN), 113.8, 111.4 (2 × acetal C),
sn-1-Stearoyl-2-arachidonoyl phospatidylinositol 4-phos-
81.5, 77.5, 76.0, 75.4, 74.96, 74.83 (6 × Ins-CH), 69.3 (Gly phate, triethylammonium salt, 2d. 1-O-(sn-1-O-Stearoyl-2-ara-
2-CH2), 66.2 (d, JC–P 6.3, Gly 3-CH2), 62.1–61.7 (2C, Gly 1-CH2 + chidonoyl glycer-3-yloxy)(2-cyanoethyloxy)phosphoryl-4-O-di(2-
P–O–CH2), 37.8, 36.36 (2 × CH2COORs), 36.28, 35.12, 35.07, cyanoethyloxy)phosphoryl-2,3:5,6-O-dicyclohexylidene-myo-ino-
34.0, 33.6, 31.9, 31.5, 29.7 (6C), 29.5 (2C), 29.38, 29.32, 29.16, sitol (26d, 148 mg, 0.115 mmol) was evaporated from MeCN
27.2, 26.5, 25.6 (4C), 24.8 (3C), 24.79, 24.72, 23.99, 23.79, (3 × 2 mL), and dissolved in CH2Cl2–MeCN (1 : 1, 6 mL). To
23.72, 23.65, 22.71 (2C), 22.59, 19.4 [(24 × fatty acid CH2,) + (10 this was added trimethylsilyl chloride (1 mL) then N,N,N′,N′-
× cyclohexylidene CH2,) + CH2CN], 14.15, 14.10 (2 × CH3); tetramethyl-N″-tert-butyl-guanidine (90%, 774 µL, 5.92 mmol,
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