HETEROCYCLES

Short Paper | Special issue | Vol. 88, No. 1, 2014, pp. 689-704
Received, 3rd June, 2013, Accepted, 23rd July, 2013, Published online, 24th July, 2013.
DOI: 10.3987/COM-13-S(S)23

■ Synthetic Studies on Glycosphingolipids from Protostomia Phyla: Synthesis of Glycosphingolipid from the Marine Sponge Spheciospongia vesparia and Its Analogue

Noriyasu Hada, Akira Miyamura, Isao Ohtsuka, and Fumiyuki Kiuchi*

Faculty of Pharmacy, Keio University, 1-5-30 Shiba-Kouen, Minato-ku, Tokyo, Japan

Abstract

Stereocontrolled syntheses of a neutral glycosphingolipid found from a marine sponge Spheciospongia vesparia and its analogue have been accomplished. Disaccharide glycosphingolipids, β-D-Arap-(1→6)-β-D-Glcp-(1↔1)-Cer (1) and α-D-Arap-(1→6)-β-D-Glcp-(1↔1)-Cer (2), were synthesized from suitable monosaccharide donors and an acceptor by stepwise synthesis.

In our continuing systematic studies on the role and biological functions of glycosphingolipids, we have synthesized glycolipids found in various lower animal species.1 Glycolipids found from various marine sponges are especially interesting molecules from the viewpoint of drug discovery. For example, it is well established that sponges of the genus Agelas produce α-galactosyl ceramides (α-GalCer),2 which are potent ligands of the MHC class I-like CD1d protein and found to activate iNKT cells strongly.3 On the other hand, marine sponges are a rich source of novel glycosphingolipids which are often characterized by unprecedented structural features.4 Therefore, we are interested in the structure and function of glycosphingolipids found from various marine sponges. In our previous papers, we synthesized a glycosphingolipid, β-D-GalNAcp(1→4)[α-D-Fucp(1→3)]-β-D-GlcNAcp(1↔1)Cer (A), isolated from a marine sponge Aplysinella rhax and five analogues and examined their structure-activity relationships on LPS-induced NO production.1e,1g Recently, Mangoni et al. isolated and characterized a novel neutral glycosphingolipid, β-D-Arap-(1→6)-β-D-Glcp(1↔1)-Cer (1, Scheme 3) from a marine sponge Spheciospongia vesparia.4d The carbohydrate part of 1 consists of a D-arabinose attached to the reducing-end D-glucose through an β1→6 linkage and this was the first example of a natural diglycosylceramide whose carbohydrate chain contains a pentose unit. In this study, we report the synthesis of glycosphingolipid 1 and its structural analogue 2.

Phenyl 2,3,4-tri-O-benzyl-1-thio-α-D-arabinopyranoside (5) was selected as a glycosyl donor at first, and 2-(trimethylsilyl)ethyl 2,3,4-tri-O-benzoyl-β-D-glucopyranoside (8) as a glycosyl acceptor (Scheme 1). The glycosyl donor (5) was prepared from D-arabinose by a sequence of acetylation, thioglycosylation, deacetylation and subsequent benzylation using standard conditions. On the other hand, the acceptor 8 was prepared from 2-(trimethylsilyl)ethyl β-D-glucopyranoside (6)5 by the following three-step procedure. Regioselective tritylation of the starting material 6 with triphenylmethyl chloride, followed by benzoylation (7) and acid hydrolysis of the trityl group, gave compound 8. We first carried out glycosylation of 8 with 5 under argon atmosphere with several combinations of solvents and promoters. The results are summarized in Table 1.

When a combination of NIS and TfOH was used as a promoter6 in 1:1 ether-CH2Cl2, the ratio of 9α : 9β was 1.2 : 1.0 (entry 1), and the ratio of 9β was higher with MeOTf,7 although the yield was not so good (entry 3). When a combination of NIS/AgOTf in cyclopentyl methyl ether (CPME) was used,8 the ratio of α : β was 1.0 : 1.1 (entry 2), but the yield was excellent (100%). The ratio of the β-arabinoside was improved with dimethyl(methylthio)sulfonium trifluoromethanesulfonate (DMTST, entry 4) as a promoter and the yield was also excellent.9 However, sufficient stereoselectivity of desired β-arabinoside was not obtained (α : β = 1.2 : 1.0〜1.0 : 2.5). Thus, the activation of thioglycoside 5 under these conditions led to non stereoselective glycosylation.

Li et al. reported a rational design of a highly α-selective galactopyranosyl donor based on the influence of remote protecting groups.10 They demonstrated that a galactosyl donor with acetyl groups at 3- and 4-positions and a benzyl group at 2-position showed excellent stereoselectivity, probably though blockage of β-side attack to the cationic intermediate by intramolecular acetal formation with the acetyl groups, to give only α-product. Therefore, we applied this method to the β-selective D-arabinosylation which corresponds to α-selective D-galactosylation. Compound 12 was chosen as a new glycosyl donor and was prepared from phenyl thio-α-D-arabinopyranoside 10 by a four-step procedure. Protection of hydroxyl groups at 3- and 4-positions of 10, which was obtained from deacetylation of 4, with an isopropylidene group, followed by benzylation gave compound 11. Removal of the isopropylidene group of 11 under acidic condition followed by acetylation afforded desired glycosyl donor 12. Glycosylation of the acceptor 8 with 12 in the presence of DMTST and MS 3Å in CH2Cl2-Et2O afforded desired disaccharide 13 in 95% yield (Scheme 2). The 1H NMR spectra of 13 showed a doublet at δ 5.04 for anomeric proton with J1,2 = 3.0 Hz, which confirmed the β-stereochemistry.

Removal of the benzyl protecting group of 13 by catalytic hydrogenation over 10% Pd-C in THF-MeOH and acetylation gave protected disaccharide 14. Selective removal of the 2-(trimethylsilyl)ethyl (TMS-ethyl) group in 14 with TFA, followed by exposure of resulted hemiacetal to CCl3CN and 1,8-diazabicyclo[5,4,0]-7-undecene (DBU)11 afforded corresponding α-trichloroacetimidate 15. Glycosylation of (2S,3S,4R)-2-azido-3-O-benzoyl-4-octadecane-1,3,4-triol (16)12 with the glycosyl donor 15 was carried out in the presence of trimethylsilyl trifluoromethanesulfonate (TMSOTf)13 and MS 4Å for 8 h at 0 °C, to afford desired β-glycoside 17 in 93% yield. Selective reduction14 of the azido group in 17 with triphenylphosphine in THF-water gave corresponding amine, which on condensation with stearic acid using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) in CH2Cl2, gave fully protected derivative 18 (54%, 2-steps). Finally, standard deacetylation of 18 and purification by column chromatography on Sephadex LH-20 furnished glycolipid 1 (Scheme 3). The structure and purity of 1 were demonstrated by its 1H NMR and HR-FABMS data.

As we had a good amount of 9α as a glycosyl donor in hand, the synthesis of α-D-Arap-(1→6)-β-D- Glcp(1↔1)-Cer (2) was also carried out in order to compare biological activities of natural and non natural products. Deprotection of benzyl groups of 9α by catalytic hydrogenation over 10% Pd-C in THF-MeOH and acetylation provided 19. Compound 19 was exposed to TFA and resulted hemiacetal was converted to glycosyl imidate 20 using standard conditions. Glycosylation of the azidosphingosin 16 with 20 produced glycosyl azidosphingosin 21 which was further converted into glycosyl ceramide 2 by a series of reactions as described for 1 (Scheme 4). The structure and purity of 2 were demonstrated by its 1H NMR and HR-FABMS data.

In conclusion, a stereoselective synthesis of the glycosphingolipid 1 found from Spheciospongia vesparia together with a synthesis of its analogue 2 have been accomplished. We have succeeded in the first total synthesis of D-arabinose-containing glycosphingolipid found from invertebrate species in good yield.

EXPERIMENTAL
General
Optical rotations were measured with a Jasco P-1020 digital polarimeter. 1H and 13C NMR spectra were recorded with a Varian 400 or 500 FT NMR spectrometer. Me4Si was used as an internal standard. MALDI-TOFMS was recorded on an AB SCIEX Voyager RP mass spectrometer. High-resolution mass spectra were recorded on a JEOL JMS-700 under FAB conditions. TLC was performed on Silica Gel 60 F254 (E. Merck) with detection by quenching of UV fluorescence and by charring with 10% H2SO4. Column chromatography was carried out on Silica Gel 60 (E. Merck). 2-(Trimethylsilyl)ethyl β-D-glucopyranoside (7)5 was prepared as reported. (2S,3S,4R)-2-Azido-3-O-benzoyl-4-octadecane-1,3,4-triol (16) was prepared from phytosphingosine, which was purchased from Degussa (The Netherlands), according to the synthesis of (2S,3R,4E)-2-azido-3-O-benzoyl-4-octadecene-1,3-diol.12
Phenyl 2,3,4-tri-O-acetyl-1-thio-α-D-arabinopyranoside (4)
A solution of D-arabinopyranose 3 (5.0 g, 33.3 mmol) and acetic anhydride (12 mL) in pyridine (18 mL) was stirred for 20 h at room temperature. After the reaction was quenched with MeOH, toluene was added and co-evaporated several times. To a solution of the residue in CH2Cl2 (30 mL) cooled at 0 °C were added PhSH (5.13 mL, 50.0 mmol) and BF3・OEt2 (6.27 mL, 50.0 mmol). The reaction mixture was stirred for 2 h at room temperature. The mixture was poured into ice-water and extracted with CHCl3. The extract was successively washed with aq NaHCO3 and brine, dried (MgSO4), and concentrated. The residue was purified by silica gel column chromatography (10:1 hexane-EtOAc) to give 4 (11.1 g, 2 steps 91%). [α]D25 +18.1 (c 2.9, CHCl3). HR-FABMS: calcd for C17H21O7S: m/z 369.1008, found: m/z 369.0990 [M+H]+. 1H NMR (500 MHz, CDCl3): δ 7.50—7.26 (m, 5H, Ph), 5.26 (dd, 1H, J3, 4 = 3.0 Hz, H-4), 5.24 (t, 1H, J1, 2 = J2, 3 = 8.3 Hz, H-2), 5.10 (dd, 1H, H-3), 4.81 (d, 1H, H-1), 4.15 (dd, 1H, J4, 5a = 4.2 Hz, J5a, 5b = 12.7 Hz, H-5a), 3.67 (dd, 1H, J4, 5b = 2.0 Hz, H-5b). 13C NMR (125 MHz, CDCl3): δ 170.1, 169.8, 169.3, 132.2, 132.0, 129.0, 128.9, 127.9, 86.7, (C-1), 70.4 (C-3), 68.4 (C-2), 67.4 (C-4), 75.2 (C-5), 20.79, 20.76, 20.6.

Phenyl 2,3,4-tri-O-benzyl-1-thio-α-D-arabinopyranoside (5)
To a solution of 4 (11.1 g, 30.0 mmol) in MeOH (40 mL) was added NaOMe (300 mg) and the mixture was stirred for 1.5 h at room temperature. After completion of the reaction, the reaction mixture was neutralized with Amberlite IR 120 [H+]. After filtration, the solution was concentrated, and the residue was dissolved in dry N,N-dimethylformamide (DMF, 30 mL). To the stirred solution was added sodium hydride (NaH, 7.2 g, 180 mmol: 60% oil dispersion), and the mixture was stirred for 30 min at 0 °C and then benzyl bromide (16.5 mL, 122.0 mmol) was added. The stirring was continued for 4 h at 0 °C and then MeOH was added to destroy excess NaH. The mixture was poured into iced-water and extracted with EtOAc. The extract was successively washed with aq NaHCO3 and brine, dried (MgSO4), and concentrated. The product was purified by silica gel column chromatography (10:1 hexane-EtOAc) to give 5 (8.29 g, 2 steps 54%). [α]D25 +24.3 (c 1.6, CHCl3). HR-FABMS: calcd for C32H32O4S: m/z 512.2021, found: m/z 512.2059 [M]+. MALDI-TOFMS: m/z 535.9 [M+Na]+ (C32H32O4SNa). 1H NMR (500 MHz, CDCl3): δ 7.53—7.19 (m, 20H, 4×Ph), 4.91 (d, 1H, J1, 2 = 6.0 Hz, H-1), 4.70—4.58 (m, 6H, 3×CH2), 4.26 (dd, 1H, J4, 5a = 5.8 Hz, J5a, 5b = 11.9 Hz, H-5a), 3.93 (t, 1H, J2, 3 = 6.5 Hz, H-2), 3.82 (dd, 1H, J3, 4 = 2.5 Hz, H-4), 3.68 (dd, 1H, H-3), 3.44 (br.d, 1H, H-5b). 13C NMR (125 MHz, CDCl3): δ 138.14, 138.09, 137.97, 135.5, 131.1, 128.8, 128.4, 128.31, 128.30, 128.0, 127.8, 127.73, 127.71, 127.6, 126.9, 87.1 (C-1), 78.4 (C-3), 77.2 (C-2), 74.1, 72.3 (C-4), 72.2, 71.0, 63.1 (C-5).

2-(Trimethylsilyl)ethyl 2,3,4-tri-O-benzoyl-6-O-trityl-β-D-glucopyranoside (7)
A solution of 6 (1.0 g, 3.57 mmol) and NaOMe (200 mg) in MeOH (50 mL) was stirred for 30 min, and then neutralized with Amberlite IR 120 [H+]. The mixture was filtered and concentrated. A mixture of the resulted deacetyl compound (2.89 g) and triphenylmethyl chloride (TrCl, 1.39 g, 4.99 mmol) in pyridine (10 mL) was stirred for 4 h at 80 °C. After completion of the reaction, the reaction mixture was added benzoyl chloride (1.49 mL, 12.8 mmol), and the mixture was stirred for 20 h at 80 °C. Toluene was added and the mixture was concentrated to dryness, then the residue was dissolved in CHCl3, washed successively with 5% HCl, aq NaHCO3 and water, dried (MgSO4), and concentrated. The product was purified by silica gel column chromatography (8:1 hexane-EtOAc) to give 7 (2.26 g, 2 steps 76%).
[α]D25 +0.4 (c 1.9, CHCl3). HR-FABMS: calcd for C51H50O9SiNa: m/z 857.3122, found: m/z 857.3104 [M+Na]+. 1H NMR (500 MHz, CDCl3): δ 7.97—7.09 (m, 30H, 6×Ph), 5.78 (t, 1H, J2, 3 = J3, 4 = 9.7 Hz, H-3), 5.60 (t, 1H, J4, 5 = 9.7 Hz, H-4), 5.48 (dd, 1H, J1, 2 = 8.1 Hz, H-2), 4.83 (d, 1H, H-1), 4.15—4.10 (m, 1H, OCH2CH2), 3.86 (dt, 1H, J5, 6a = 2.6 Hz, J5, 6b = 5.1 Hz, H-5), 3.75—3.70 (m, 1H, OCH2CH2), 3.33 (dd, 2H, J6a, 6b = 10.6 Hz, H-6a, 6b), 1.04—0.96 (m, 2H, OCH2CH2), –0.02 (s, 9H, Si(CH3)3). 13C NMR (125 MHz, CDCl3): δ 165.9, 165.1, 164.8, 143.6, 133.03, 132.99, 129.77, 129.75, 129.63, 129.57, 129.2, 129.0, 128.6, 128.23, 128.19, 128.1, 127.9, 127.7, 126.9, 100.5 (C-1), 86.7, 73.9 (C-5), 73.4 (C-3), 72.2 (C-2), 69.6 (C-4), 67.3 (OCH2CH2), 62.5 (C-6), 18.0 (OCH2CH2), −1.4 (Si(CH3)3).

2-(Trimethylsilyl)ethyl 2,3,4-tri-O-benzoyl-β-D-glucopyranoside (8)
A solution of 7 (2.26 g, 2.71 mmol) in 80% AcOH (9 mL) was stirred at 90 °C for 1 h, then diluted with toluene and concentrated. The product was purified by silica gel column chromatography (4:1 hexane-EtOAc) to give 8 (1.29 g, 80%). [α]D25 −4.8 (c 0.8, CHCl3). HR-FABMS: calcd for C32H36O9SiNa: m/z 615.2026, found: m/z 615.2023 [M+Na]+. 1H NMR (500 MHz, CDCl3): δ 7.96—7.26 (m, 15H, 3×Ph), 5.93 (t, 1H, J2, 3 = J3, 4 = 9.8 Hz, H-3), 5.52—5.48 (m, 2H, H-2, 4), 4.85 (d, 1H, J1, 2 = 8.1 Hz, H-1), 4.08—4.03 (m, 1H, OCH2CH2), 3.89—3.74 (m, 3H, H-5, 6a, 6b), 3.66—3.60 (m, 1H, OCH2CH2), 0.96—0.87 (m, 2H, OCH2CH2), –0.06 (s, 9H, Si(CH3)3). 13C NMR (125 MHz, CDCl3): δ 166.0, 165.8, 165.0, 133.6, 133.2, 133.1, 129.9, 129.73, 129.69, 129.4, 128.8, 128.6, 128.5, 128.3, 100.6 (C-1), 74.5 (C-5), 72.9 (C-3), 71.9 (C-2), 69.6 (C-4), 67.7 (OCH2CH2), 61.4 (C-6), 18.0 (OCH2CH2), −1.5 (Si(CH3)3).

2-(Trimethylsilyl)ethyl 2,3,4-tri-O-benzyl-D-arabinopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-β-D-glucopyranoside (9)
entry 1
A mixture of 8 (200 mg, 0.34 mmol), 5 (223 mg, 0.44 mmol) and powdered MS AW300 (300 mg) in dry CH2Cl2-Et2O (1:1, 3 mL) was stirred under Ar atmosphere for 2 h at room temperature, then cooled to −40 °C. NIS (195 mg, 0.87 mmol) and TfOH (10.9 μL, 0.09 mmol) were added to the mixture, which was stirred for 3 h at −40 °C, then neutralized with Et3N. The precipitates were filtered off and washed with CHCl3. The combined filtrate and washings were successively washed with saturated aqueous Na2S2O3 and water, dried (MgSO4), and concentrated. The residue was separated by silica gel column chromatography (20:1 toluene-EtOAc) to give 9α (110 mg, 33%) and 9β (90 mg, 27%).
9α [α]D25 −17.6 (c 0.65, CHCl3). HR-FABMS: calcd for C58H62O13SiNa: m/z 1017.3857, found: m/z 1017.3890 [M+Na]+. 1H NMR (500 MHz, CDCl3): δ 7.96—7.14 (m, 30H, 6×Ph), 5.85 (t, 1H, J2,3 =J3,4 = 9.6 Hz, H-3), 5.51 (t, 1H, J4,5 = 9.6 Hz, H-4), 5.47 (t, 1H, J1,2 = 8.0 Hz, H-2), 4.76 (d, 1H, H-1), 4.71—4.55 (m, 6H, 3×benzyl methylene), 4.39 (d, 1H, J1',2' = 6.2 Hz, H-1'), 4.07—3.94 (m, 3H, H-6a, 5’a, CH2CH2Si(CH3)3), 3.83 (br. d, 1H, H-6b), 3.72 (dd, 1H, H-2'), 3.66 (br.s, 1H, H-4’), 3.66-3.53 (m, 1H, CH2CH2Si(CH3)3), 3.49 (dd, 1H, H-3'), 3.20(d, 1H, H-5'a), 0.93—0.80 (m, 2H, CH2CH2Si(CH3)3), –0.10 (s, 9H, Si(CH3)3). 13C NMR (125 MHz, CDCl3): δ 165.8, 165.1, 165.0, 138.7, 138.4, 133.2, 133.03, 132.97, 129.8, 129.72, 129.71, 129.5, 129.1, 129.0, 128.9, 128.31, 128.27, 128.22, 128.20, 128.16, 127.84, 127.82, 127.60, 127.55, 127.50, 127.47, 103.3 (C-1'), 100.5 (C-1), 78.8 (C-3’), 78.4 (C-2’), 74.6, 73.8, 73.3 (C-3), 72.2, 72.0 (C-2), 71.1 (C-5), 70.2 (C-4), 68.3(C-6), 67.3, 62.1 (C-5’), 17.8, –1.5 (Si(CH3)3).
9β [α]D25 −40.7 (c 2.3, CHCl3). HR-FABMS: calcd for C58H62O13SiNa: m/z 1017.3857, found: m/z 1017.3859 [M+Na]+. 1H NMR (500 MHz, CDCl3): δ 7.95—7.22 (m, 30H, 6×Ph), 5.84 (t, 1H, J2,3 = J3,4 = 9.7 Hz, H-3), 5.50—5.46 (m, 2H, H-2, 4), 4.89 (d, 1H, J1',2' = 3.0 Hz, H-1'), 4.79 (d, 1H, J1,2 = 7.6 Hz, H-1), 4.78—4.49 (m, 6H, 3×benzyl methylene), 4.07 (br.t, 1H, H-5), 4.02—3.98 (m, 1H, CH2CH2Si(CH3)3), 3.96 (dd, 1H, J2',3'= 9.0 Hz, H-2'), 3.86—3.76 (m, 3H, H-6a, 6b, 3’), 3.70 (br.s, 1H, H-4’), 3.67-3.53 (m, 3H, H-5’a, 5'b, CH2CH2Si(CH3)3), 0.89—0.81 (m, 2H, CH2CH2Si(CH3)3), –0.11 (s, 9H, Si(CH3)3). 13C NMR (125 MHz, CDCl3): δ 165.8, 165.2, 165.0, 138.9, 138.7, 138.3, 133.1, 133.0, 129.8, 129.72, 129.68, 129.5, 129.0, 128.9, 128.4, 128.3, 128.22, 128.18, 127.8, 127.6, 127.54, 127.52, 127.43, 127.39, 100.4 (C-1), 99.1 (C-1'), 76.8 (C-3’), 76.3 (C-2’), 73.9 (C-4’), 73.7 (C-5), 73.22 (C-3), 73.18, 72.6, 71.9, 71.7 (C-2), 70.2 (C-4), 67.7 (C-6), 67.3, 60.6 (C-5’), 17.8, –1.5 (Si(CH3)3).
MALDI-TOFMS: m/z 1018.4 [M+Na]+ (C58H62O13SiNa).

entry 2 (Table 1)
A mixture of 8 (150 mg, 0.25 mmol), 5 (155 mg, 0.30 mmol) and powdered MS AW300 (300 mg) in dry cyclopentyl methyl ether (CPME, 3 mL) was stirred under Ar atmosphere for 2 h at room temperature, then cooled to 0 °C. NIS (102 mg, 0.45 mmol) and AgOTf (93 mg, 0.36 mmol) were added to the mixture, which was stirred for 1 h at 0 °C, then neutralized with Et3N. The precipitates were filtered off and washed with CHCl3. The combined filtrate and washings were successively washed with saturated aqueous Na2S2O3 and water, dried (MgSO4), and concentrated. The residue was separated by silica gel column chromatography (20:1 toluene-EtOAc) to give 9α (120 mg, 48%) and 9β (130 mg, 52%).

entry 3 (Table 1)
A mixture of 8 (50 mg, 0.08 mmol), 5 (52 mg, 0.09 mmol) and MS 3Å (150 mg) in dry CH2Cl2-Et2O (1:1, 1.0 mL) was stirred for 1.5 h at room temperature. MeOTf (50 μL, 0.38 mmol) was added, and the mixture was stirred for 2 h at room temperature, then neutralized with Et3N. The precipitates were filtrated off and washed with CHCl3. The combined filtrate and washings were washed with water, dried (MgSO4), and concentrated. The residue was separated by silica gel column chromatography (20:1 toluene-EtOAc) to give 9α (10 mg, 13%) and 9β (22 mg, 27%).

entry 4 (Table 1)
A mixture of 8 (135 mg, 0.23 mmol), 5 (140 mg, 0.28 mmol) and MS 3Å (300 mg) in dry Et2O-CPME (1:1, 2.0 mL) was stirred for 2 h at room temperature, then cooled to 0 °C. Dimethyl(methylthio)sulfonium triflate (DMTST, 570 mg, 1.1 mmol) was added, and the mixture was stirred for 2 h at room temperature, then neutralized with Et3N. The precipitates were filtrated off and washed with CHCl3. The combined filtrate and washings were washed with water, dried (MgSO4), and concentrated. The residue was separated by silica gel column chromatography (20:1 toluene-EtOAc) to give 9α (62 mg, 27%) and 9β (155 mg, 69%).

Phenyl 3,4-O-isopropylidene-2-O-benzyl-1-thio-α-D-arabinopyranoside (11)
A mixture of 10 (4.46 g, 18.4 mmol) in acetone (30 mL), camphorsulfonic acid (CSA, 2.14 g, 9.20 mmol) and 2,2-dimethoxypropane (DMP, 4.5 mL, 36.8 mmol) was stirred for 1.5 h at 40 °C, then neutralized with Et3N. The precipitates were filtrated off and the filtrated was concentrated. The product was purified by silica gel column chromatography (1:1 hexane-EtOAc) to give an isopropylidene derivative. To a solution of this compound (3.97 g, 14.1 mmol) in DMF (60 mL) was added NaH (1.12 g, 28.1 mmol: 60% of oil dispersion), and the mixture was stirred for 30 min at 0 °C and then BnBr (6.9 mL, 28.1 mmol) was added. The stirring was continued for 3 h at 0 °C and then MeOH was added to destroy excess NaH. The mixture was poured into iced-water and extracted with EtOAc. The extract was successively washed with aq NaHCO3 and brine, dried (MgSO4), and concentrated. The product was purified by silica gel column chromatography (10:1 hexane-EtOAc) to give 11 (4.41 g, 2 steps 64%). [α]D25 +18.9 (c 2.3, CHCl3). HR-FABMS: calcd for C21H24O4S: m/z 372.1395, found: m/z 372.1420 [M]+. 1H NMR (500 MHz, CDCl3): δ 7.52—7.24 (m, 10H, 2×Ph), 4.82(d, 1H, J1,2 = 7.7 Hz, H-1), 4.79 and 4.68 (each d, Jgem = 11.5 Hz, benzyl methylene), 4.28 (dt, 1H, J3,4 = 7.7 Hz, J4, 5a = J4, 5b =3.9, H-4), 4.24 (t, 1H, J2,3 = 5.5 Hz, H-3), 4.18 (dd, 1H, Hz, J5a, 5b = 12.9 Hz, H-5a), 3.75 (dd, 1H, H-5b), 3.62 (dd, 1H, H-2), 1.47, 1.37 (each s, 6H, 2×CH3). 13C NMR (125 MHz, CDCl3): δ 137.7, 134.1, 131.8, 128.8, 128.3, 128.1, 127.7, 127.3, 109.9, 86.3 (C-1), 78.1 (C-2), 78.0 (C-3), 73.3, 72.4 (C-4), 64.5 (C-5), 27.7, 26.1.

Phenyl 3,4-di-O-acetyl-2-O-benzyl-1-thio-α-D-arabinopyranoside (12)
A solution of 11 (4.41 g, 11.8 mmol) in 80% AcOH (40 mL) was stirred at 50 °C for 18 h, then diluted with toluene and concentrated. The residue was acetylated with acetic anhydride (15 mL) in pyridine (20 mL). The reaction was quenched with MeOH, then toluene was added and co-evaporated several times. The product was purified by silica gel column chromatography (4:1 hexane-EtOAc) to give 12 (4.53 g, 2 steps 92%). [α]D25 −1.5 (c 1.2, CHCl3). HR-FABMS: calcd for C22H24O6S: m/z 416.1294, found: m/z 416.1328 [M]+. MALDI-TOFMS: m/z 440.1 [M+Na]+ (C22H24O6SNa). 1H NMR (500 MHz, CDCl3): δ 7.55—7.25 (m, 10H, 2×Ph), 5.29 (br.d, 1H, H-4), 5.11 (dd, 1H, J2,3 = 3.1 Hz, J3,4 = 8.1 Hz, H-3), 4.85 (d, 1H, J1,2 = 7.5 Hz, H-1), 4.81 and 4.61 (each d, Jgem = 11.1 Hz, benzyl methylene), 4.12 (dd, 1H, J4, 5a = 4.0 Hz, Hz, J5a, 5b = 12.6 Hz, H-5a), 3.81 (t, 1H, H-2), 3.65 (dd, 1H, J4, 5b = 1.9 Hz, H-5b), 2.11, 2.01 (each s, 6H, 2×COCH3). 13C NMR (125 MHz, CDCl3): δ 170.1, 169.9, 137.6, 134.0, 131.8, 129.0, 128.4, 127.84, 127.79, 127.6, 87.8 (C-1), 75.9 (C-2), 72.2 (C-3), 67.9 (C-4), 65.0 (C-5), 20.9, 20.8.

2-(Trimethylsilyl)ethyl 3,4-di-O-acetyl-2-O-benzyl-β-D-arabinopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-β-D-glucopyranoside (13)
A mixture of 8 (150 mg, 0.25 mmol), 12 (126 mg, 0.30 mmol) and MS 3Å (300 mg) in dry CH2Cl2-Et2O (1:1, 3.0 mL) was stirred for 1 h at room temperature, then cooled to 0 °C. DMTST (623 mg, 1.21 mmol) was added, and the mixture was stirred for 3 h at 0 °C, then neutralized with Et3N. The precipitates were filtrated off and washed with CHCl3. The combined filtrate and washings were washed with water, dried (MgSO4), and concentrated. The product was purified by silica gel column chromatography (2:1 hexane-EtOAc) to give 13 (214 mg, 95%). [α]D25 −58.3 (c 2.1, CHCl3). HR-FABMS: calcd for C48H54O15SiNa: m/z 921.3130, found: m/z 921.3171 [M+Na]+. 1H NMR (500 MHz, CDCl3): δ 8.03—7.32 (m, 20H, 4×Ph), 5.93 (t, 1H, J2, 3 = J3, 4 = 9.6 Hz, H-3), 5.56 (t, 1H, J1,2 = 8.1 Hz, H-2), 5.51 (t, 1H, J4, 5 = 9.9 Hz, H-4), 5.38 (dd, 1H, J2’, 3’ = 10.1 Hz, J3’, 4’ = 3.3 Hz, H-3’), 5.35 (br.s, 1H, H-4’), 5.04 (d, 1H, J1’,2’ = 3.0 Hz H-1’), 4.87 (d, 1H, H-1), 4.76 and 4.72 (each d, 2H, Jgem = 12.3 Hz, benzyl methylene), 4.14 (br.t, 1H, H-5), 4.11—4.07 (m, 1H, CH2CH2Si(CH3)3), 3.96—3.82 (m, 4H, H-6a, 6b, 2’, 5’a), 3.67-3.60 (m, 2H, H-5'b, CH2CH2Si(CH3)3), 2.17, 2.07 (each s, 6H, 2xCOCH3), 0.95—0.88 (m, 2H, CH2CH2Si(CH3)3), –0.03 (s, 9H, Si(CH3)3). 13C NMR (125 MHz, CDCl3): δ 170.2, 169.9, 165.7, 165.3, 165.0, 138.2, 133.4, 133.1, 133.0, 129.8, 129.74, 129.70, 129.5, 128.9, 128.8, 128.44, 128.35, 128.3, 128.21, 128.20, 127.70, 127.66, 127.4, 100.4 (C-1), 98.5 (C-1’), 74.0 (C-5), 73.9 (C-2’), 73.2 (C-3), 72.6 (benzyl methylene), 71.9 (C-2), 70.0 (C-4), 69.3 (C-4’), 69.0 (C-3’), 67.7 (C-6), 67.5 (OCH2CH2-), 60.5 (C-5’), 20.9, 20.8, 17.8, –1.5 (Si(CH3)3).

2-(Trimethylsilyl)ethyl 2,3,4-tri-O-acetyl-β-D-arabinopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-β-D-glucopyranoside (14)
Compound 13 (202 mg, 0.22 mmol) in MeOH–THF (1:1, 4.0 mL) was hydrogenolyzed (0.4 MPa) under hydrogen in the presence of 10% Pd/C (200 mg) for 3 h at room temperature, then the mixture was filtered and concentrated. The residue was acetylated with acetic anhydride (2.0 mL) in pyridine (3.0 mL). The reaction mixture was poured into ice-water and extracted with CHCl3. The extract was successively washed with 5% HCl, aq NaHCO3 and brine, dried (MgSO4), and concentrated. The residue was purified by silica gel column chromatography (10:1 toluene-EtOAc) to give 14 (165 mg, 2 steps 88%). [α]D25 −73.5 (c 2.0, CHCl3). HR-FABMS: calcd for C43H50O16SiNa: m/z 873.2766, found: m/z 873.2771 [M+Na]+. 1H NMR (500 MHz, CDCl3): δ 7.94—7.18 (m, 15H, 3×Ph), 5.85 (t, 1H, J2, 3 = J3, 4 = 9.7 Hz, H-3), 5.55 (t, 1H, J4,5 = 9.7 Hz, H-4), 5.49 (t, 1H, J1,2 = 8.0 Hz, H-2), 5.37 (dd, 1H, J2’, 3’ = 9.0 Hz, J3’, 4’ = 3.3 Hz, H-3’), 5.35 (br.s, 1H, H-4’), 5.19 (dd, 1H, J1’,2’ = 3.5 Hz, H-2’), 5.10 (d, 1H, H-1'), 4.81 (d, 1H, H-1), 4.07—4.02 (m, 1H, CH2CH2Si(CH3)3), 3.99—3.95 (m, 2H, H-6a, 5), 3.89 (d, 1H, H-5'), 3.68—3.60 (m, 2H, H-6b, CH2CH2Si(CH3)3), 3.67-3.60 (m, 2H, H-5'b, CH2CH2Si(CH3)3), 2.01, 1.99 (each s, 6H, 2xCOCH3), 0.94—0.88 (m, 2H, CH2CH2Si(CH3)3), –0.05 (s, 9H, Si(CH3)3). 13C NMR (125 MHz, CDCl3): δ 170.4, 170.3, 169.8, 165.8, 165.0, 164.9, 133.4, 133.09, 133.10, 129.7, 129.5, 129.0, 128.9, 128.4, 128.21, 128.19, 100.5 (C-1), 96.9 (C-1’), 73.6 (C-5), 73.3 (C-3), 71.8 (C-2), 69.3 (C-4), 69.4 (C-4), 69.0 (C-3'), 68.0 (C-2'), 67.4 (C-4’), 67.2 (OCH2CH2-), 66.5 (C-6), 60.3 (C-5’), 30.8, 20.9, 20.74, 20.66, 17.8, –1.5 (Si(CH3)3).

2,3,4-Tri-O-acetyl-β-D-arabinopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-α-D-glucopyranosyl trichloroacetimidate (15)
A solution of 14 (165 mg, 0.19 mmol) in CH2Cl2 (1.0 mL) cooled to 0 °C was added CF3CO2H (1.0 mL), and the mixture was stirred for 2 h at room temperature and then concentrated. EtOAc and toluene (1:2) were added and evaporated to give corresponding reducing sugar. To a solution of the residue in CH2Cl2 (1.0 mL) cooled at 0 °C were added DBU (15.0 mL, 0.10 mmol) and CCl3CN (0.30 mL, 3.00 mmol). The reaction mixture was stirred for 1 h at 0 °C. After completion of the reaction, the mixture was concentrated. The product was purified by silica gel column chromatography (3:1 hexane-EtOAc) as eluent to give 15 (139 mg, 2 steps 82%). 1H NMR (500 MHz, CDCl3): δ 8.65 (s, 1H, NH), 7.96—7.26 (m, 15H, 3×Ph), 6.82 (d, 1H, J1,2 = 3.5 Hz, H-1), 6.24 (t, 1H, J2, 3 = J3, 4 = 10.0 Hz, H-3), 5.77 (t, 1H, J4,5 = 10.0 Hz, H-4), 5.60 (dd, 1H, H-2), 5.40—5.36 (m, 2H, H-3', 4'), 5.21 (dd, 1H, J1’,2’ =3.5 Hz, J2’, 3’ = 10.5 Hz, H-2’), 5.04 (d, 1H, H-1'), 4.42 (dt, 1H, H-5), 3.95 (d, 1H, H-5’a), 3.94 (dd, 1H, J5,6a = 2.2 Hz, J5, 6b = 11.8 Hz, H-6a), 3.65 (dd, 1H, H-5’b), 3.59 (dd, 1H, J5,6a = 4.1 Hz, H-6a). 2.14, 2.13, 2.03 (each, s, 9H, 3×OAc).

2,3,4-Tri-O-acetyl-β-D-arabinopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-β-D-glucopyranosyl-(1→1)-(2S,3S,4R)-2-azido-3,4-di-O-benzoyl-octadecane-1,3,4-triol (17)
A mixture of 15 (139 mg, 0.16 mmol), 16 (167 mg, 0.30 mol) and MS 4Å (300 mg) in dry CH2Cl2 (1.3 mL) was stirred for 16 h at room temperature, then cooled to 0 °C. TMSOTf (25 mL, 0.14 mmol) was added, and the mixture was stirred for 1.5 h at 0 °C, then neutralized with Et3N. The precipitates were filtrated off and washed with CHCl3. The combined filtrate and washings were washed with water, dried (MgSO4), and concentrated. The product was purified by silica gel column chromatography (10:1 toluene-EtOAc) to give 17 (185 mg, 93%). [α]D25 −23.9 (c 0.54, CHCl3). HR-FABMS: calcd for C70H81O20N3Na: m/z 1306.5311, found: m/z 1306.5334 [M+Na]+. 1H NMR (500 MHz, CDCl3): δ 4.95 (d, 1H, J1’,2’ = 3.5 Hz, H-1'), 4.78 (d, 1H, J1, 2 = 7.8 Hz, H-1).

2,3,4-Tri-O-acetyl-β-D-arabinopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-β-D-glucopyranosyl-(1→1)-(2S,3S,4R)-3,4-di-O-benzoyl-2-hexadecanamido-octadecane-1, 3, 4-triol (18)
To an emulsion of 17 (185 mg, 0.14 mmol) in THF-H2O (5:1, 4.8 mL) was added triphenylphosphine (113 mg, 0.43 mmol), and the mixture was stirred for 20 h at 50 °C. The mixture was concentrated, and the residue was stirred with palmitic acid (108 mg, 0.42 mmol) and 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC: 80 mg, 0.42 mmol) in dry CH2Cl2 (2.0 mL) for 24 h at rt. The mixture was diluted with CHCl3, washed with water, dried (MgSO4), and concentrated. The product was purified by silica gel column chromatography (15:1 toluene-acetone) to give 18 (117 mg, 2 steps 54%). [α]D25 −31.4 (c 1.7, CHCl3). MALDI-TOFMS: m/z 1534.5 [M+K]+ (C86H113NO21K). 1H NMR (500 MHz, CDCl3): δ 8.13—7.16 (m, 25H, 5×Ph), 6.30 (d, 1H, J = 9.4 Hz, NH), 5.78 (t, 1H, J2, 3 = J3, 4 = 9.7 Hz, H-3), 5.64 (dd, 1H, H-3 of sphingosin), 5.43—5.35 (m, 2H, H-2, 4), 5.27—5.24 (m, 2H, H-3’, 4’), 5.09 (dd, 1H, J1’,2’ = 3.4 Hz, J2’, 3’ = 10.1 Hz, H-2’), 4.90 (d, 1H, H-1'), 4.71 (d, 1H, J1,2 = 7.8 Hz, H-1), 4.64 (br.t, 1H, CH(NHR)). 13C NMR (125 MHz, CDCl3): δ 206.9, 178.6, 173.1, 170.6, 170.2, 169.8, 166.1, 165.6, 165.1, 165.02, 164.96, 133.4, 133.3, 133.2, 133.0, 130.1, 129.84, 129.76, 129.71, 129.6, 128.99, 128.93, 128.8, 128.7, 128.5, 128.43, 128.37, 128.3, 128.21, 128.17, 125.3, 100.4 (C-1), 97.0 (C-1'), 73.8, 73.6, 72.8, 71.8, 69.2, 69.0, 68.1, 67.1, 66.9, 66.4, 60.4, 47.8, 36.3, 35.3, 33.9, 31.9, 29.7, 29.64, 29.61, 29.55, 29.5, 29.40, 29.37, 29.3, 29.21, 29.15, 29.0, 28.9, 28.8, 25.5, 24.7, 24.2, 22.6, 20.9, 20.73, 20.65, 14.1.

β-D-Arabinopyranosyl-(1→6)-β-D-glucopyranosyl-(1→1)-(2S,3S,4R)-2-hexadecanamido-octadecane-1,3,4-triol (1)
A solution of 18 (68 mg, 0.05 mol) and NaOMe (40 mg) in 1,4-dioxane-MeOH (1:1, 4 mL) was stirred at 50 °C for 48 h, and then neutralized with Amberlite IR 120 [H+]. The mixture was filtered and concentrated. The product was purified by Sephadex LH-20 column chromatography with 1:1 CHCl3-MeOH to give 1 (23 mg, 60%). [α]D25 −28.0 (c 0.46, CHCl3-MeOH). HR-FABMS: calcd for C45H87NONa: m/z 872.6075 found: m/z 872.6031 [M+Na]+. 1H NMR (500 MHz, CDCl3-CD3OD): δ 4.89 (br.d, 1H, H-1'), 4.31 (d, 1H, J1,2 = 7.8 Hz, H-1). 13C NMR (125 MHz, CDCl3-CD3OD): δ 175.2, 104.2 (C-1), 100.3 (C-1'), 76.8, 75.9, 74.8, 72.5, 70.7, 70.16, 70.10, 69.9, 69.8, 63.8, 63.5, 61.3, 50.8, 36.9, 32.6, 32.4, 30.3, 30.2, 30.14, 30.06, 30.0, 29.9, 29.8, 26.5, 26.4, 23.1, 14.3.

2-(Trimethylsilyl)ethyl 2,3,4-tri-O-acetyl-α-D-arabinopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-β-D-glucopyranoside (19)
Compound 19 was prepared from 9α (181 mg, 0.18 mmol) as described for preparation of 14. The product was purified by silica gel column chromatography (8:1 toluene-EtOAc) to give 19 (161 mg, 2 steps, quant.). [α]D 25 +9.2 (c 0.17, CHCl3). HR-FABMS: calcd for C43H50O16SiNa: m/z 873.2766, found: m/z 873.2784 [M+Na]+. 1H NMR (500 MHz, CDCl3): δ 7.94—7.27 (m, 15H, 3×Ph), 5.84 (t, 1H, J2, 3 = J3, 4 = 9.6 Hz, H-3), 5.54 (t, 1H, J4,5 = 9.6 Hz, H-4), 5.45 (dd, 1H, J1,2 = 7.9 Hz, H-2), 5.22 (br.s, 1H, H-4’), 5.17 (dd, 1H, J1’, 2’ = 6.2 Hz, J2’, 3’ = 8.6 Hz, H-2’), 5.06 (dd, 1H, J3’,4’ = 3.6 Hz, H-3’), 4.80 (d, 1H, H-1), 4.61 (d, 1H, H-1’), 4.08—4.03 (m, 1H, CH2CH2Si(CH3)3), 4.01—3.96 (m, 4H, H-5, 6a, 6b, 5’a), 3.86 (br.d, 1H, H-6b), 3.66—3.61 (m, 1H, CH2CH2Si(CH3)3), 3.55 (dd, 1H, H-5'), 2.12, 2.09, 2.06 (each s, 9H, 3×COCH3), 0.93—0.87 (m, 2H, CH2CH2Si(CH3)3), –0.05 (s, 9H, Si(CH3)3). 13C NMR (125 MHz, CDCl3): δ 170.2, 170.1, 169.3, 165.8, 165.00, 164.9, 133.3, 133.1, 133.0, 129.73, 129.71, 129.65, 129.4, 129.0, 128.9, 128.3, 128.22, 128.20, 100.6 (C-1), 100.4 (C-1’), 73.8 (C-5), 73.3 (C-3), 71.9 (C-2), 69.7 (C-3’), 69.4 (C-4), 69.0 (C-2’), 67.5, 67.2 (C-4), 67.0 (C-6), 62.1 (C-5’), 20.82, 20.76, 20.7, 17.9, –1.5 (Si(CH3)3).

2,3,4-Tri-O-acetyl-α-D-arabinopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-α-D-glucopyranosyl trichloroacetimidate (20)
Compound 20 was prepared from 19 (159 mg, 0.19 mmol) as described for preparation of 15. The product was purified by silica gel column chromatography (2:1 hexane-EtOAc) to give 20 (146 mg, 2 steps 87%). [α]D 25 +52.3 (c 0.52, CHCl3). HR-FABMS: calcd for C40H38O16NCl3Na: m/z 916.1154, found: m/z 916.1160 [M+Na]+.1H NMR (500 MHz, CDCl3): δ 8.62 (s, 1H, NH), 7.96—7.26 (m, 15H, 3×Ph), 6.82 (d, 1H, J1,2 = 2.1 Hz, H-1), 6.21 (t, 1H, J2, 3 = J3, 4 = 10.0 Hz, H-3), 5.76 (t, 1H, J4,5 = 10.0 Hz, H-4), 5.53 (dd, 1H, H-2), 5.21—5.19 (m, 2H, H-4', 2'), 5.05 (dd, 1H, J2’, 3' = 3.0 Hz, J3’, 4' = 7.5 Hz, H-3’), 4.61 (d, 1H, J1', 2' = 6.0 Hz, H-1'), 4.46 (dt, 1H, H-5), 4.00 (d, 1H, J5,6a = 4.5 Hz, H-6a), 3.92 (dd, 1H, J5, 6b = 4.0 Hz, H-6b), 3.86 (dd, 1H, H-5’a), 3.56 (dd, 1H, J4', 5' = 2.5 Hz, H-5'b). 2.14, 2.11, 2.07 (each, S, 9H, 3×OAc). 13C NMR (125 MHz, CDCl3): 170.3, 170.2, 169.3, 165.7, 165.4, 164.8, 160.5, 133.5, 133.4, 133.2, 129.9, 129.8, 129.7, 128.9, 128.5, 128.40, 128.37, 128.3, 100.2 (C-1'), 93.2 (C-1), 90.7, 72.1, 70.7, 70.4, 69.7, 68.9, 68.1, 67.3, 65.8, 62.2, 20.9, 20.8, 20.7.

2,3,4-Tri-O-acetyl-α-D-arabinopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-β-D-glucopyranosyl-(1→1)-(2S,3S,4R)-2-azido-3,4-di-O-benzoyl-octadecane-1,3,4-triol (21)
Compound 21 was prepared from 20 (233 mg, 0.26 mmol) and 16 (169 mg, 0.29 mmol) as described for preparation of 17. The product was purified by silica gel column chromatography (10:1 toluene-EtOAc) to give 21 (294 mg, 88%). [α]D 25 +3.8 (c 0.50, CHCl3). HR-FABMS: calcd for C70H81O20N3Na: m/z 1306.5311, found: m/z 1306.5277 [M+Na]+. 1H NMR (500 MHz, CDCl3): δ 7.99—7.26 (m, 25H, 5×Ph), 5.80 (t, 1H, J2,3 = J3,4 = 9.6 Hz, H-3), 5.56 (t, 1H, J4,5 = 9.5 Hz, H-4), 5.50—5.45 (m, 2H, H-2, CH(OBzR)), 5.21 (br.d, 1H, H-4’), 5.16 (dd, 1H, J1',2' = J2',3' = 6.5 Hz, H-2'), 5.04 (dd, 1H, J3',4' = 3.4 Hz, H-3'), 4.82 (d, 1H, J1,2 = 7.8 Hz, H-1), 4.61 (d, 1H, H-1'), 3.95-3.90 (m, 2H, H-5, 6a), 3.54 (d, 1H, J5,6 = 11.3 Hz, H-6b). 13C NMR (125 MHz, CDCl3): δ 170.3, 170.1, 169.5, 165.8, 165.7, 165.1, 164.9, 164.8, 133.5, 133.3, 133.22, 133.18, 133.11, 129.9, 129.78, 129.75, 129.7, 129.3, 129.2, 129.1, 128.8, 128.6, 128.5, 128.34, 128.26, 100.6 (C-1,C-1’), 74.1 (C-5), 73.1 (C-3), 72.9 (C-2), 72.6, 71.6, 70.0 (C-3'), 69.2 (C-2'), 68.9 (C-4), 67.5 (C-4'), 66.5, 62.6 (C-6), 61.0 (C-5’), 31.9, 29.8, 29.7, 29.62, 29.58, 29.5, 29.41, 29.36, 29.34, 29.26, 22.7, 20.9, 20.8, 20.7, 14.1.

2,3,4-Tri-O-acetyl-α-D-arabinopyranosyl-(1→6)-2,3,4-tri-O-benzoyl-β-D-glucopyranosyl-(1→1)-(2S,3S,4R)-3,4-di-O-benzoyl-2-hexadecanamido-octadecane-1,3,4 triol (22)
Compound 22 was prepared from 21 (168 mg, 0.13 mmol) as described for preparation of 18. The product was purified by silica gel column chromatography (9:1 toluene-EtOAc) to give 22 (71 mg, 2 steps 36%). [α]D 25 +16.1 (c 1.4, CHCl3). MALDI-TOFMS: m/z 1517.6 [M+Na]+ (C86H113NO21Na). 1H NMR (500 MHz, CDCl3): δ 8.05—7.18 (m, 25H, 5×Ph), 6.23 (d, 1H, J = 9.4 Hz, NH), 5.76 (t, 1H, J2,3 = J3,4 = 9.6 Hz, H-3), 5.53 (t, 1H, J4,5 = 9.7 Hz, H-4), 5.37—5.31 (m, 2H, H-2, CH(OBzR)), 4.63 (d, 1H, J1,2 = 7.9 Hz, H-1). 13C NMR (125 MHz, CDCl3): δ 173.0, 170.3, 170.0, 169.5, 166.1, 165.7, 165.1, 165.0, 164.7, 133.4, 133.3, 133.21, 133.17, 132.97, 133.04, 129.84, 129.80, 129.76, 129.74, 129.66, 129.1, 129.00, 128.96, 128.8, 128.6, 128.44, 128.35, 128.3, 128.24, 128.19, 101.1, 100.5, 74.0, 73.7, 72.8, 72.5, 72.1, 70.0, 69.5, 68.7, 68.1, 67.6, 66.6, 62.4, 47.9, 36.4, 31.9, 29.70, 29.67, 29.65, 29.62, 29.60, 29.56, 29.4, 29.34, 29.28, 28.4, 25.52, 25.50, 22.7, 20.9, 20.7, 14.1.

α-D-Arabinopyranosyl-(1→6)-β-D-glucopyranosyl-(1→1)-(2S,3S,4R)-2-hexadecanamido-octadecane-1,3,4-triol (2)
Compound 2 was prepared from 22 (133 mg, 0.09 mmol) as described for preparation of 1. The product was purified by Sephadex LH-20 column chromatography in 1:1 CHCl3-MeOH to give 2 (58 mg, 78%). [α]D 25 +4.4 (c 0.86, CHCl3-MeOH). HR-FABMS: calcd for C45H87NONa: m/z 872.6075 found: m/z 872.6118 [M+Na]+. 1H NMR (500 MHz, CDCl3-CD3OD): δ 4.32 (d, 1H, J1’,2’ = 8.0 Hz, H-1'), 4.27 (d, 1H, J1,2 = 7.8 Hz, H-1). 13C-NMR (500 MHz, CDCl3-CD3OD): δ 175.1, 104.2 (C-1), 103.6 (C-1'), 76.6, 75.4, 74.7, 74.2, 73.1, 72.3, 71.6, 70.1, 69.9, 68.3, 67.7, 66.0, 50.8, 36.9, 32.40, 32.25, 30.25, 30.16, 30.13, 30.0, 29.94, 29.83, 26.4, 23.1, 14.3.

ACKNOWLEDGEMENTS
This work was supported by a Grant-in-Aid for Scientific Research (No. 25460131) and by Platform for Drug Discovery, Informatics, and Structural Life Science from the Ministry of Education, Culture, Sports, Science and Technology of Japan (MEXT). The authors are grateful to Ms. J. Hada for measurements of HR-FABMS data.

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