Efficient Synthesis of the Lewis A Tandem Repeat

Abstract

The convergent synthesis of the Lewis A (Le^a) tandem repeat is described. The Le^a tandem repeat is a carbohydrate ligand for a mannose binding protein that shows potent inhibitory activity against carcinoma growth. The Le^a unit, {β-d-Gal-(1→3)-[α-l-Fuc-(1→4)]-β-d-GlcNAc}, was synthesized by stereoselective nitrile-assisted β-galactosylation with the phenyl 3-O-allyl-2,4,6-tri-O-benzyl-1-thio-β-galactoside, and ether-assisted α-fucosylation with fucosyl (N-phenyl)trifluoroacetimidate. This common Le^a unit was easily converted to an acceptor and donor in high yields, and the stereoselective assembly of the hexasaccharide and dodecasaccharide as the Le^a tandem repeat framework was achieved by 2-trichloroacetamido-assisted β-glycosylation and the (N-phenyl)trifluoroacetimidate method.

1. Introduction

The Lewis A (Le^a) trisaccharide, {β-d-Gal-(1→3)-[α-l-Fuc-(1→4)]-β-d-GlcNAc}, is a component of glycolipids that have been identified as antigens of the Lewis blood group. Recently, N-linked glycoproteins with Le^a tandem repeats (Figure 1) consisting of four or more repeated sequences were isolated from the SW1116 human colorectal carcinoma cell line. This carbohydrate ligand forms a mannose binding protein–carbohydrate complex, which shows potent inhibitory activity against growth of human colorectal carcinoma cells [1,2,3]. To elucidate the inhibitory mechanism, it is necessary to determine and synthesize the minimum structure of the carbohydrate ligand; however, synthesis of the Le^a tandem repeat has not been reported. We have recently developed and reported a new synthetic strategy for core 2 decasaccharide with four repeated type-II N-acetyllactsamines using a benzyl-protected N-trichloroacetyllactsaminyl imidate with high β-selectivity and high yield [4]. In this paper, we describe a synthetic method for the tetrameric Le^a tandem repeat motif and the hexasaccharide and dodecasaccharide, via the synthesis of type-I lactosamine by β-selective galactosylation [5] and convergent synthesis with N-trichloroacetyllactosaminyl imidate.

2. Results and Discussion

In the retrosynthetic analysis of the Le^a tetramer, we planned the convergent synthesis with a Le^a trisaccharide common intermediate, which would be constructed with β-d-Gal-(1→3)-β-d-GlcNTCA synthesized by using β-selective galactosylation and 2-p-methoxybenzyl fucosyl imidate. The trisaccharide was designed as the suitably protected form equipped with TBDPS group on the 1-position of the glucosamine and an allyl group on the 3-position of the galactose for divergent synthesis of the acceptor and the donor. In addition, the trichloroacetyl (TCA) group on the 2-position of the glucosamine and the benzyl groups were expected to ensure high stereoselectivity and high yield during the later glycosylation [4,6,7,8,9,10,11,12,13]. We envisioned that the tetramer could be obtained by glycosylation promoted by a catalytic Lewis acid with haxasaccharyl acceptor and donor, which could be prepared from the hexasaccharide Le^a dimer, and the hexasaccharide could be synthesized by coupling the acceptor and N-phenyl trichloroacetimidyl donor provided from the Le^a trisaccharide common intermediate (Scheme 1).

Le^a trisaccharide 11 was synthesized from galactosyl donor 2 [5], 2-azidoglucosyl acceptor 3 [14], and fucosyl donor 8 [15,16] (Scheme 2). First, disaccharide 4 was constructed from 2 and 3 by propionitrile-mediated β-selective galactosylation [5] under BSP-Tf₂O-TTBP [17] conditions in 89% yield. Then, 4 was transformed into disaccharide acceptor 7 by reducing the azide group to give 5, TCA group protection to give 6, and reductive ring opening of the benzylidene to give 7 in 79% yield (three steps). The glycosylation of 7 with 8 was conducted in cyclopentyl methyl ether-dichloromethane (1:1) [16,18] at −40 °C to obtain trisaccharide 9 in 95% yield. Le^a common intermediate 11 was provided by deprotection of a 4-methoxybenzyl group and subsequent acetylation.

Le^a trisaccharide 11 was readily converted to donor 13 and acceptor 14 (Scheme 3). After desilylation of 11, the resulting hemiacetal 12 was treated with (N-phenyl)trifluoroacetimidoyl chloride [19] and K₂CO₃ to obtain (N-phenyl)trifluoroacetimidate 13. Acceptor 14 was prepared by selective deallylation with iridium-catalyzed olefin migration, followed by treatment with HgCl₂ and HgO in aqueous acetone solution [20]. The glycosylation of 13 and 14 promoted by catalytic TMSOTf proceeded at −78 °C to give hexasaccharide 15 in 93% yield [4,21,22]. By using the same procedure, hexasaccharide donor 17 and acceptor 18 were prepared in high yields, respectively. In the next coupling, the oligosaccharides showed lower reactivity, and the reaction occurred at 0 °C to afford dodecasaccharide 19 (88%). Next, the linker for sugar probes was introduced (Scheme 4). Dodecasaccharide 19 was converted to (N-phenyl)trifluoroacetimidate 21 (85% over two steps) in an analogous manner, and coupled with N-Boc aminopropanol to give 22 in 79% yield. After deallylation, microwave-assisted reductive dehalogenation of the TCA group was attempted with excess Zn and AcOH in several solvents (ethyl acetate, 1,4-dioxane, AcOH, and THF) [10]. However, Boc group was cleaved, giving the mixture of aminopropyl and acetamidopropyl derivatives as the main products. It was found that THF was most suitable to obtain aminopropyl derivative 24 almost predominantly. Then, the terminal amino group of 24 was re-protected with Boc group to afford fine 25 in 76% over two steps.

3. Experimental Section

3.1. General Methods

¹H- and ¹³C-NMR spectra were recorded with a spectrometer (Avance III 500, Bruker, Billerica, MA, USA). Chemical shifts are expressed in ppm (δ) relative to the Me₄Si signal as an internal standard. Electrospray ionization time-of-flight high-resolution mass spectrometry was performed (micrOTOF, Bruker Daltonics, Billerica, MA, USA). Specific rotations were determined with a high-sensitivity polarimeter (SEPA-300, Horiba, Kyoto, Japan). Microwave irradiation was carried out in a microwave reactor (μReactor Ex, Shikoku Instrumentation Co., Ltd., Kagawa, Japan). TLC analysis was performed on glass TLC plates (silica gel 60F₂₅₄, Merck, Darmstadt, Germany). Compounds were visualized either by exposure to UV light (254 nm) or by dipping in a solution of 10% H₂SO₄ in ethanol, in a solution of phosphomolybdic acid, H₃PO₄, and H₂SO₄, in H₂O, or in ninhydrin reagent, followed by heating. Column chromatography was performed with the solvent system (v/v) specified on silica gel BW-80S, BW-300, PSQ-60B (Fuji Silysia Chemical Ltd. Kasugai, Japan), or Wakosil HC-N (Wako Pure Chemical Industries, Ltd. Osaka, Japan). Gel permeation chromatography was performed with the solvent system (v/v) specified on Sephadex LH-20 (GE Healthcare UK Ltd. Little Chalfont, UK), or Bio-beads S-X1, S-X3 (Bio-Rad Laboratories, Inc. Hercules, CA, USA). Evaporation and concentration were carried out in vacuo.

3.2. Physical Data for All New Compounds

tert-Butyldiphenylsilyl 3-O-allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-2-azido-4,6-O-benzylidene-2-deoxy-β-d-glucopyranoside (4). To a mixture of phenyl 3-O-allyl-2,4,6-tri-O-benzyl-1-thio-β-d-galactopyranoside 2 (438 mg, 0.75 mmol), tert-butyldiphenylsilyl 2-azido-4,6-O-benzylidene-2-deoxy-β-d-glucopyranoside 3 (200 mg, 0.38 mmol), benzenesulfinyl piperidine (237 mg, 1.13 mmol), tri-tert-butylpyrimidine (373 mg, 1.50 mmol), and molecular sieves 4A (1.41 g) in propionitrile (12.5 mL) was added dropwise trifluoromethanesulfonic anhydride (140 μL, 0.83 mmol) at −80 °C under Ar, and stirred for 1 h at −80 °C. The reaction mixture was quenched with sat. NaHCO₃ aq., filtered through Celite, and diluted with EtOAc. The organic layer was separated, and the aqueous layer was extracted with EtOAc. The combined organic layer was successively washed with brine, dried over Na₂SO₄, and concentrated. The crude product was chromatographed on silica gel (PSQ-60B) with toluene–acetone (98:2) to give the title product 4 (338 mg, 89%). [α]_D −31.3° (c 1.1, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.70–7.15 (m, 30H, Ar), 5.93–5.85 (m, 1H, H₂C=CHCH₂), 5.39 (s, 1H, >CHPh), 5.31–5.27 (m, 1H, H₂C=CHCH₂), 5.16–5.13 (m, 1H, H₂C=CHCH₂), 4.96–4.89 (m, 2H, PhCH₂ × 2), 4.80 (d, 1H, J_gem = 10.7 Hz, PhCH₂), 4.63 (d, 1H, J_1,2 = 7.9 Hz, H-1^Gal), 4.57 (d, 1H, J_gem = 11.7 Hz, PhCH₂), 4.48 (d, 1H, J_1,2 = 7.8 Hz, H-1^GlcN), 4.26–4.19 (m, 2H, PhCH₂ × 2), 4.15–4.09 (m, 2H, H₂C=CHCH₂), 3.89 (dd, 1H, J_5,6b = 4.9 Hz, J_gem = 10.1 Hz, H-6a^GlcN), 3.78–3.69 (m, 4H, H-2^Gal, H-4^Gal, H-3^GlcN, H-4^GlcN), 3.59–3.51 (m, 3H, H-2^GlcN, H-6b^GlcN, H-6a^Gal), 3.36–3.31 (m, 2H, H-3^Gal, H-6b^Gal), 3.25 (dd, 1H, J_5,6a = J_5,6b = 6.5 Hz, H-5^Gal), 2.96–2.91 (m, 1H, H-5^GlcN), 1.16 (s, 9H, ^tBu); ¹³C-NMR (125 MHz, CDCl₃) δ 138.9, 138.8, 137.9, 137.3, 135.8, 134.9, 133.1, 132.5, 130.0, 129.8, 128.9, 128.4, 128.2, 128.2, 128.2, 128.1, 128.0, 127.9, 127.7, 127.6, 127.5, 127.4, 127.4, 126.0, 116.5, 102.7, 101.1, 97.2, 82.3, 79.9, 79.8, 78.7, 75.2, 74.4, 73.5, 73.1, 72.9, 71.7, 68.9, 68.8, 68.3, 66.2, 26.8, 19.1. HRMS (ESI) m/z: found [M + Na]⁺ 1026.4337, C₅₉H₆₅N₃O₁₀Si calcd. for [M + Na]⁺ 1026.4337.

tert-Butyldiphenylsilyl 3-O-allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-2-amino-4,6-O-benzylidene-2-deoxy-β-d-glucopyranoside (5). A mixture of 4 (576 mg, 0.57 mmol), powdered Zn (1.50 g, 23.0 mmol), and AcOH (0.66 ml, 11.5 mmol) in CH₂Cl₂ (14 mL) was stirred for 30 min at room temperature under Ar. The mixture was diluted with CHCl₃ and filtered through Celite. The filtrate was evaporated, and the residue was diluted with CHCl₃. The organic layer was successively washed with sat. NaHCO₃, water, and brine, dried over Na₂SO₄, and concentrated. The residue was chromatographed on silica gel with toluene–MeOH (95:5) to give the title product 5 (540 mg, 96%). [α]_D −21.8° (c 1.3, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.68–7.18 (m, 30H, Ar), 5.94–5.87 (m, 1H, H₂C=CHCH₂), 5.45 (s, 1H, >CHPh), 5.33–5.28 (m, 1H, H₂C=CHCH₂), 5.18–5.15 (m, 1H, H₂C=CHCH₂), 4.93–4.88 (m, 2H, PhCH₂ × 2), 4.78 (d, 1H, J_gem = 10.9 Hz, PhCH₂), 4.58 (d, 1H, J_gem = 11.6 Hz, PhCH₂), 4.49 (d, 1H, J_1,2 = 7.9 Hz, H-1^Gal), 4.43 (d, 1H, J_1,2 = 7.8 Hz, H-1^GlcN), 4.31–4.25 (m, 2H, PhCH₂ × 2), 4.17–4.08 (m, 2H, H₂C=CHCH₂), 3.96 (dd, 1H, J_5,6a = 4.9 Hz, J_gem = 10.4 Hz, H-6a^GlcN), 3.87–3.81 (m, 2H, H-2^Gal, H-4^Gal), 3.66–3.58 (m, 3H, H-4^GlcN, H-6b^GlcN, H-6a^GlcN), 3.54 (t, 1H, J_2,3 = J_3.4 = 9.2 Hz, H-3^GlcN), 3.41–3.37 (m, 3H, H-3^Gal, H-5^Gal, H-6b^Gal), 3.03–2.99 (m, 2H, H-2^GlcN, H-5^GlcN), 1.20 (s, 9H, ^tBu); ¹³C-NMR (125 MHz, CDCl₃) δ 138.9, 138.5, 137.9, 137.6, 135.8, 135.8, 134.8, 133.4, 132.9, 129.8, 129.7, 128.6, 128.4, 128.3, 128.3, 128.1, 128.1, 127.8, 127.7, 127.7, 127.5, 127.4, 127.3, 126.1, 116.7, 104.3, 100.8, 99.1, 83.5, 82.7, 79.9, 79.4, 75.7, 74.5, 73.5, 73.1, 73.0, 71.4, 68.4, 68.2, 66.8, 60.1, 27.0, 19.2. HRMS (ESI) m/z: found [M + Na]⁺ 1000.4432, C₅₉H₆₇NO₁₀Si calcd. for [M + Na]⁺ 1000.4432.

tert-Butyldiphenylsilyl 3-O-allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-4,6-O-benzylidene-2-deoxy-2-trichloroacetamido-β-d-glucopyranoside (6); To a solution of 5 (12.9 g, 13.2 mmol) in pyridine (132 mL) was added dropwise trichloroacetyl chloride (1.76 mL, 15.8 mmol) at 0 °C under Ar, and stirred at 0 °C for 1 h. The mixture was evaporated, and the residue was diluted with CHCl₃, successively washed with 2 M HCl, sat. NaHCO₃, and brine, dried over Na₂SO₄, and concentrated. The residue was chromatographed on silica gel with toluene–EtOAc (97:3) to give the title product 6 (14.8 g, quant.). [α]_D −9.8° (c 1.3, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.67–7.18 (m, 30H, Ar), 7.02 (d, 1H, J_2,NH = 7.0 Hz, NH), 5.95–5.87 (m, 1H, H₂C=CHCH₂), 5.45 (s, 1H, >CHPh), 5.34–5.30 (m, 1H, H₂C=CHCH₂), 5.24 (d, 1H, J_1,2 = 7.9 Hz, H-1^GlcN), 5.20–5.17 (m, 1H, H₂C=CHCH₂), 4.90 (d, 1H, J_gem = 11.6 Hz, PhCH₂), 4.82 (d, 1H, J_gem = 10.7 Hz, PhCH₂), 4.69 (d, 1H, PhCH₂), 4.57 (d, 1H, PhCH₂), 4.42 (d, 1H, J_1,2 = 7.9 Hz, H-1^Gal), 4.39–4.30 (m, 3H, PhCH₂ × 2, H-3^GlcN), 4.18–4.10 (m, 2H, H₂C=CHCH₂), 3.95 (dd, 1H, J_5,6a = 4.9 Hz, J_gem = 10.4 Hz, H-6a^GlcN), 3.82–3.79 (m, 2H, H-2^Gal, H-4^Gal), 3.65 (t, 1H, J_3,4 = J_4,5 = 9.2 Hz, H-4^GlcN), 3.61–3.54 (m, 2H, H-6b^GlcN, H-6a^Gal), 3.49 (dd, 1H, J_5,6b = 5.4 Hz, J_gem = 9.0 Hz, H-6b^Gal), 3.44–3.39 (m, 2H, H-2^GlcN, H-5^Gal), 3.31 (dd, 1H, J_2,3 = 9.8 Hz, J_3,4 = 2.9 Hz, H-3^Gal), 3.12–3.07 (m, 1H, H-5^GlcN), 1.06 (s, 9H, ^tBu); ¹³C-NMR (125 MHz, CDCl₃) δ 161.4, 138.9, 138.7, 137.8, 137.3, 135.9, 135.7, 134.8, 133.0, 132.5, 129.8, 129.8, 128.7, 128.5, 128.4, 128.1, 128.1, 128.0, 127.9, 127.9, 127.8, 127.5, 127.4, 127.4, 126.1, 116.8, 103.1, 100.8, 94.1, 92.2, 82.1, 79.9, 79.2, 77.6, 76.5, 75.8, 74.5, 73.6, 73.3, 73.2, 71.4, 68.3, 68.3, 65.9, 62.7, 26.9, 19.1. HRMS (ESI) m/z: found [M + Na]⁺ 1144.3368, C₆₁H₆₆Cl₃NO₁₁Si calcd. for [M + Na]⁺ 1144.3368.

tert-Butyldiphenylsilyl 3-O-allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-6-O-benzyl-2-deoxy-2-tri-chloroacetamido-β-d-glucopyranoside (7). To a mixture of 6 (814 mg, 0.73 mmol) and molecular sieves 4A (4.20 g) in CH₂Cl₂ (7.3 mL) was added triethylsilane (463 μL, 2.90 mmol) and trifluoromethane sulfonic acid (127 μL, 1.45 mmol) at −78 °C under Ar, and stirred for 1 h at −78 °C, and 1.5 h at −40 °C. The reaction mixture was quenched with triethylamine, filtered through Celite, and diluted with CHCl₃. The organic layer was successively washed with sat. NaHCO₃, water, and brine, dried over Na₂SO₄, and concentrated. The crude product was chromatographed on silica gel with toluene–EtOAc (89:11) to give the title product 7 (665 mg, 82%). [α]_D +1.5° (c 1.3, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.72–7.16 (m, 30H, Ar), 6.70 (d, 1H, J_2,NH = 7.1 Hz, NH), 5.93–5.86 (m, 1H, H₂C=CHCH₂), 5.32–5.29 (m, 1H, H₂C=CHCH₂), 5.19–5.17 (m, 1H, H₂C=CHCH₂), 5.11 (d, 1H, J_1,2 = 8.0 Hz, H-1^GlcN), 4.88 (d, 1H, J_gem = 11.6 Hz, PhCH₂), 4.82 (d, 1H, J_gem = 11.3 Hz, PhCH₂), 4.72 (d, 1H, PhCH₂), 4.53 (d, 1H, PhCH₂), 4.42–4.34 (m, 4H, PhCH₂ × 4), 4.23 (d, 1H, J_1,2 = 7.8 Hz, H-1^Gal), 4.19–4.12 (m, 2H, H₂C=CHCH₂), 4.03 (dd, 1H, J_2,3 = 10.1 Hz, J_3,4 = 8.4 Hz, H-3^GlcN), 3.80 (d, 1H, J_3,4 = 2.9 Hz, H-4^Gal), 3.75 (dd, 1H, J_2,3 = 9.8 Hz, H-2^Gal), 3.69 (s, 1H, OH), 3.58–3.47 (m, 6H, H-4^GlcN, H-5^Gal, H-6a^GlcN, H-6a^Gal, H-6b^GlcN, H-6b^Gal), 3.38–3.33 (m, 1H, H-2^GlcN), 3.29 (dd, 1 H, H-3^Gal), 3.17–3.13 (m, 1 H, H-5^GlcN), 1.06 (s, 9 H, ^tBu); ¹³C-NMR (125 MHz, CDCl₃) δ 161.5, 139.1, 138.6, 138.4, 137.6, 136.0, 135.8, 134.7, 133.1, 132.6, 129.7, 129.7, 128.4, 128.4, 128.2, 128.2, 127.9, 127.8, 127.8, 127.6, 127.5, 127.4, 127.3, 127.3, 116.8, 103.5, 93.7, 92.1, 81.7, 81.1, 79.5, 75.7, 75.1, 74.6, 73.6, 73.5, 73.4, 73.4, 71.7, 69.2, 69.1, 68.3, 61.3, 26.9, 19.1. HRMS (ESI) m/z: found [M + Na]⁺ 1146.3525, C₆₁H₆₈Cl₃NO₁₁Si calcd. for [M + Na]⁺ 1146.3525.

tert-Butyldiphenylsilyl 3-O-allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[3,4-di-O-acetyl-6-2-O-p-methoxybenzyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranoside (9). To a mixture of 7 (1.96 g, 1.74 mmol), 3,4-di-O-acetyl-2-O-p-methoxybenzyl-l-fucipyranosyl (N-phenyl)-2,2,2-trifluoroacetimidate 8 (1.87 g, 3.47 mmol), and molecular sieves AW-300 (5.22 g) in CPME/CH₂Cl₂ (1:1, 58.0 mL) was added TMSOTf (15.7 μL, 0.087 mmol) dropwise at −40 °C under Ar, and stirred for 1 h at −40 °C. The reaction mixture was quenched with sat. NaHCO₃, filtered through Celite, and diluted with CHCl₃. The organic layer was separated, and the aqueous layer was extracted with CHCl₃. The combined organic layer was successively washed with water and brine, dried over Na₂SO₄, and concentrated. The crude product was chromatographed on silica gel with hexane–acetone (80:20) to give the title product 9 (2.44 g, 95%). [α]_D −19.6° (c 1.3, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.72–7.11 (m, 32H, Ar), 6.93 (d, 1H, J_2,NH = 6.8 Hz, NH), 6.81–6.78 (m, 2H, Ar), 5.95–5.87 (m, 1H, H₂C=CHCH₂), 5.35–5.32 (m, 1H, H₂C=CHCH₂), 5.21–5.11 (m, 5H, H₂C=CHCH₂, H-1^GlcN, H-1^Fuc, H-3^Fuc, H-4^Fuc), 5.09–5.05 (m, 1H, H-5^Fuc), 4.88 (d, 1H, J_gem = 10.5 Hz, ArCH₂), 4.74–4.69 (m, 2H, ArCH₂ × 2), 4.55–4.49 (m, 3H, ArCH₂ × 3), 4.46–4.39 (m, 3H, ArCH₂ × 2, H-1^Gal), 4.34 (d, 1H, J_gem = 12.6 Hz, ArCH₂), 4.24–4.08 (m, 4H, H₂C=CHCH₂ × 2, ArCH₂, H-3^GlcN), 3.87–3.77 (m, 7H, OMe, H-4^GlcN, H-4^Gal, H-6a^Gal, H-2^Fuc), 3.73–3.68 (m, 2H, H-6a^GlcN, H-6b^Gal), 3.59 (dd, 1H, J_1,2 = 8.0 Hz, J_2,3 = 9.7 Hz, H-2^Gal), 3.34 (dd, 1H, J_5,6a = 4.9 Hz, J_5,6b = 8.9 Hz, H-5^Gal), 3.30–3.27 (m, 2H, H-2^GlcN, H-3^Gal), 3.09 (dd, 1H, J_5,6b = 1.5 Hz, J_gem = 11.7 Hz, H-6b^GlcN), 2.98–2.96 (m, 1H, H-5^GlcN), 2.11 (s, 3H, Ac), 1.97 (s, 3H, Ac), 1.06 (s, 9H, ^tBu), 0.78 (d, 3H, J_5,6 = 6.5 Hz, H-6^Fuc); ¹³C-NMR (125 MHz, CDCl₃) δ 170.3, 169.1, 160.9, 159.3, 138.9, 138.5, 138.5, 138.4, 135.8, 135.7, 134.9, 133.4, 132.6, 130.1, 129.7, 129.6, 128.8, 128.5, 128.5, 128.3, 128.3, 128.2, 128.1, 128.0, 127.6, 127.5, 127.4, 127.4, 127.3, 116.8, 113.7, 103.0, 97.1, 93.4, 92.2, 81.7, 79.9, 77.6, 76.0, 75.7, 75.0, 74.6, 73.5, 73.4, 73.3, 73.2, 73.1, 72.6, 72.3, 71.2, 70.8, 67.9, 67.0, 64.2, 55.2, 26.9, 20.9, 20.8, 19.2, 15.4. HRMS (ESI) m/z: found [M + Na]⁺ 1496.4890, C₇₉H₉₀Cl₃NO₁₈Si calcd. for [M + Na]⁺ 1496.4890.

tert-Butyldiphenylsilyl 3-O-allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[3,4-di-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranoside (10). A solution of 9 (29.3 mg, 19.9 μmol) in trifluoroacetic acid/CH₂Cl₂ (1:9, 0.80 mL) was stirred for 20 min at room temperature. The reaction mixture was diluted with toluene, and evaporated. The residue was dissolved with CHCl₃, successively washed with sat. NaHCO₃, water, and brine, dried over Na₂SO₄, and concentrated. The crude product was chromatographed on silica gel with hexane–EtOAC (80:20) to give the title product 10 (27.0 g, quant.). [α]_D −33.2° (c 1.1, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.69–7.15 (m, 30H, Ar), 6.98 (d, 1H, J_2,NH = 6.7 Hz, NH), 5.97–5.89 (m, 1H, H₂C=CHCH₂), 5.36 (dd, 1H, J_trans = 17.3 Hz, J_gem = 1.6 Hz, H₂C=CHCH₂), 5.23–5.19 (m, 2H, H₂C=CHCH₂, H-1^GlcN), 5.12–5.10 (m, 2H, H-1^Fuc, H-4^Fuc), 5.02–4.95 (m, 2H, H-5^Fuc, H-3^Fuc), 4.91 (d, 1H, J_gem = 10.4 Hz, PhCH₂), 4.75 (d, 1H, J_gem = 11.1 Hz, PhCH₂), 4.68 (d, 1H, PhCH₂), 4.53 (d, 1H, PhCH₂), 4.47–4.35 (m, 5H, PhCH₂ × 4, H-1^Gal), 4.22–4.11 (m, 3H, H-3^GlcN, H₂C=CHCH₂ × 2), 3.92 (t, 1H, J_3,4 = J_4,5 = 9.4 Hz, H-4^GlcN), 3.85–3.80 (m, 2H, H-2^Fuc, H-4^Gal), 3.70–3.56 (m, 4H, H-6a^Gal, H-6b^Gal, H-6a^GlcN, H-2^Gal), 3.34 (dd, 1H, J_5,6a = 4.9 Hz, J_5,6b = 8.7 Hz, H-5^Gal), 3.30 (dd, 1H, J_2,3 = 9.8 Hz, J_3,4 = 2.8 Hz, H-3^Gal), 3.22–3.19 (m, 2H, H-6b^GlcN, H-2^GlcN), 2.95–2.93 (m, 1H, H-5^GlcN), 2.13 (s, 3H, Ac), 2.05 (s, 3H, Ac), 1.75 (d, 1H, J_2,OH = 10.8 Hz, OH), 1.03 (s, 9H, ^tBu), 0.81 (d, 3H, J_5,6 = 6.5 Hz, H-6^Fuc); ¹³C-NMR (125 MHz, CDCl₃) δ 170.4, 170.2, 161.0, 138.8, 138.4, 138.2, 138.0, 135.9, 135.7, 134.8, 133.2, 132.5, 129.7, 129.7, 128.7, 128.6, 128.5, 128.3, 128.3, 128.2, 128.2, 127.7, 127.6, 127.5, 127.3, 116.9, 103.0, 97.5, 93.2, 92.1, 81.8, 80.1, 77.6, 76.2, 75.9, 74.6, 73.4, 73.3, 73.0, 72.9, 72.4, 71.8, 71.3, 67.9, 67.3, 67.2, 64.6, 63.4, 29.7, 26.9, 21.0, 20.7, 19.2, 15.4. HRMS (ESI) m/z: found [M + Na]⁺ 1376.4315, C₇₁H₈₂Cl₃NO₁₇Si calcd. for [M + Na]⁺ 1376.4315.

tert-Butyldiphenylsilyl 3-O-allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranoside (11). To a solution of 10 (9.32 g, 6.87 mmol) in pyridine (458 mL) was added acetic anhydride (458 mL) at 0 °C under Ar, and stirred for 7 h at room temperature. The reaction mixture was concentrated. The crude product was chromatographed on silica gel with hexane–EtOAc (67:33) to give the title product 11 (9.45 g, 98%). [α]_D −37.1° (c 1.2, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.69–7.14 (m, 30H, Ar), 6.99 (d, 1H, J_2,NH = 6.7 Hz, NH), 5.97–5.90 (m, 1H, H₂C=CHCH₂), 5.35 (dd, 1H, J_trans = 17.3 Hz, J_gem = 1.7 Hz, H₂C=CHCH₂), 5.22–5.11 (m, 7H, H₂C=CHCH₂, H-1^GlcN, H-1^Fuv, H-2^Fuc, H-3^Fuc, H-4^Fuc, H-5^Fuc), 4.91 (d, 1H, J_gem = 10.3 Hz, PhCH₂), 4.75 (d, 1H, J_gem = 11.0 Hz, PhCH₂), 4.67 (d, 1H, PhCH₂), 4.53–4.50 (m, 2H, PhCH₂ × 2), 4.46–4.43 (m, 2H, PhCH₂, H-1^Gal), 4.38 (d, 1H, J_gem = 12.5 Hz, PhCH₂), 4.31 (d, 1H, PhCH₂), 4.25 (t, 1H, J_2,3 = J_3,4 = 9.4 Hz, H-3^GlcN), 4.19–4.11 (m, 2H, H₂C=CHCH₂), 3.90 (t, 1H, J_4,5 = 9.4 Hz, H-4^GlcN), 3.84 (d, 1H, J_3,4 =2.5 Hz, H-4^Gal), 3.79–3.71 (m, 2H, H-6a^Gal, H-6b^Gal), 3.60 (dd, 1H, J_1,2 = 8.0 Hz, J_2,3 = 9.7 Hz, H-2^Gal), 3.37–3.34 (m, 1H, H-5^Gal), 3.31 (dd, 1H, H-3^Gal), 3.24–3.15 (m, 3H, H-2^GlcN, H-6a^GlcN, H-6b^GlcN), 2.99–2.97 (m, 1H, H-5^GlcN), 2.15 (s, 3H, Ac), 1.97 (s, 3H, Ac), 1.96 (s, 3H, Ac), 1.03 (s, 9 H, ^tBu), 0.82 (d, 3H, J_5,6 = 6.5 Hz, H-6^Fuc); ¹³C-NMR (125 MHz, CDCl₃) δ 170.5, 170.3, 169.3, 160.9, 138.7, 138.5, 138.4, 138.0, 135.9, 135.8, 134.9, 133.3, 132.7, 129.6, 128.7, 128.6, 128.5, 128.3, 128.3, 128.2, 127.6, 127.5, 127.4, 127.4, 127.3, 116.9, 103.1, 95.3, 93.1, 92.1, 81.8, 80.1, 76.2, 75.9, 74.7, 74.5, 73.3, 73.1, 73.1, 72.6, 72.5, 71.7, 71.2, 68.2, 68.1, 68.0, 66.8, 64.3, 63.6, 26.9, 20.8, 20.8, 20.7, 19.2, 15.3. HRMS (ESI) m/z: found [M + Na]⁺ 1418.4421, C₇₃H₈₄Cl₃NO₁₈Si calcd. for [M + Na]⁺ 1418.4421.

3-O-Allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-d-glucopyranose (12). To a solution of 11 (1.67 g, 1.19 mmol) in THF (11.9 mL) were added acetic acid (0.68 mL, 11.9 mmol) and 1 M tetra-n-butylammonium fluoride in THF (4.76 mL, 4.76 mmol) at 0 °C under Ar, and stirred for 1 d at room temperature. The reaction mixture was concentrated. The residue was diluted with EtOAc and water, and extracted with EtOAc. The combined organic layer was successively washed with sat. NaHCO₃, water, and brine, dried over Na₂SO₄, and concentrated. The crude product was purified by silica gel column chromatography with hexane–EtOAc (60:40) and gel permeation chromatography (LH-20, CHCl₃-MeOH (50:50)) to give the title product 12 (1.38 g, quant.). ¹H-NMR (500 MHz, CDCl₃) δ 7.41–7.20 (m, 20H, Ar), 6.76 (d, 1H, J_2,NH = 9.7 Hz, NH), 5.87–5.80 (m, 1H, H₂C=CHCH₂), 5.30–5.12 (m, 7H, H₂C=CHCH₂ × 2, H-1^GlcN, H-1^Fuc, H-2^Fuc, H-3^Fuc, H-4^Fuc), 4.97 (dd, 1H, J_4,5 = 12.8 Hz, J_5,6 = 6.5 Hz, H-5^Fuc), 4.87 (d, 1H, J_gem = 12.1 Hz, PhCH₂), 4.72 (d, 1H, J_gem = 11.4 Hz, PhCH₂), 4.63–4.47 (m, 6H, PhCH₂ × 5, H-1^Gal), 4.44 (d, 1H, J_gem = 11.6 Hz, PhCH₂), 4.34–4.28 (m, 1H, H-2^GlcN), 4.19–4.00 (m, 4H, H₂C=CHCH₂ × 2, H-3^GlcN, H-6a^GlcN), 3.96–3.91 (m, 1H, H-4^GlcN), 3.81–3.76 (m, 2H, H-4^Gal, H-6a^Gal), 3.70 (dd, 1H, J_5,6b = 7.4 Hz, J_gem = 9.3 Hz, H-6b^Gal), 3.64–3.46 (m, 3H, H-2^Gal, H-5^GlcN, H-6b^GlcN), 3.41–3.39 (m, 1H, H-5^Gal), 3.22 (dd, 1H, J_2,3 = 9.7 Hz, J_3,4 = 2.7 Hz, H-3^Gal), 3.09–3.08 (m, 1H, OH), 2.13 (s, 3H, Ac), 2.00 (s, 3H, Ac), 1.99 (s, 3H, Ac), 0.75 (d, 3H, H-6^Fuc); ¹³C-NMR (125 MHz, CDCl₃) δ 170.5, 170.3, 169.5, 161.3, 139.0, 138.6, 138.3, 137.5, 135.0, 134.7, 129.2, 128.8, 128.5, 128.5, 128.4, 128.3, 128.2, 128.2, 128.1, 127.9, 127.8, 127.8, 127.6, 127.5, 127.1, 116.9, 116.6, 103.8, 95.8, 92.7, 91.2, 81.9, 78.6, 77.6, 74.8, 74.3, 74.2, 73.7, 73.3, 73.2, 72.8, 72.3, 71.7, 71.7, 71.1, 68.7, 68.2, 68.1, 68.1, 67.4, 64.6, 64.5, 55.8, 20.8, 20.8, 20.7, 15.4, 15.4. HRMS (ESI) m/z: found [M + Na]⁺ 1180.3243, C₅₇H₆₆Cl₃NO₁₈ calcd. for [M + Na]⁺ 1180.3243.

3-O-Allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-d-glucopyranosyl (N-phenyl)-2,2,2-trifuoroacetimidate (13). A mixture of 12 (1.35 g, 1.16 mmol), (N-phenyl)-2,2,2-trifluoroacetoimidoyl chloride (482 mg, 2.32 mmol), and K₂CO₃ (802 mg, 5.80 mmol) in acetone (23.2 mL) was stirred for 1 h at room temperature. The reaction mixture was filtered through Celite, and concentrated. The crude product was purified by gel permeation chromatography [S-X3, toluene–EtOAc (75:25)] and silica gel column chromatography with hexane–EtOAc (71:29) to give the title product 13 (1.31 g, 85%). [α]_D −0.7° (c 1.4, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.38–7.08 (m, 23H, Ar), 6.75–6.71 (m, 3H, Ar, NH), 6.32 (br, 1H, H-1^GlcN), 5.88–5.80 (m, 1H, H₂C=CHCH₂), 5.31–5.21 (m, 5H, H₂C=CHCH₂, H-1^Fuc, H-2^Fuc, H-3^Fuc, H-4^Fuc), 5.14 (dd, 1H, J_trans = 10.5 Hz, J_gem = 1.4 Hz, H₂C=CHCH₂), 4.95 (dd, 1H, J_4,5 = 10.5 Hz, J_5,6 = 6.4 Hz, H-5^Fuc), 4.84 (d, 1H, J_gem = 11.9 Hz, PhCH₂), 4.73 (d, 1H, J_gem = 11.4 Hz, PhCH₂), 4.64–4.49 (m, 7H, PhCH₂ × 5, H-2^GlcN, H-1^Gal), 4.44 (d, 1H, J_gem = 11.6 Hz, PhCH₂), 4.16 (t, 1H, J_2,3 = J_3,4 = 9.2 Hz, H-3^GlcN), 4.10–4.05 (m, 3H, H₂C=CHCH₂ × 2, H-4^GlcN), 3.87–3.81 (m, 3H, H-4^Gal, H-6a^Gal, H-5^GlcN), 3.70 (dd, 1H, J_5,6b = 7.5 Hz, J_gem = 9.2 Hz, H-6b^Gal), 3.62–3.56 (m, 3H, H-2^Gal, H-6a^GlcN, H-6b^GlcN), 3.46–3.44 (m, 1H, H-5^Gal), 3.24 (dd, 1H, J_2,3 = 9.7 Hz, J_3,4 = 2.7 Hz, H-3^Gal), 2.14 (s, 3H, Ac), 2.02 (s, 3H, Ac), 2.00 (s, 3H, Ac), 0.78 (d, 3 H, H-6^Fuc); ¹³C-NMR (125 MHz, CDCl₃) δ 170.4, 120.2, 169.5, 161.2, 142.9, 138.7, 138.4, 138.2, 137.5, 134.9, 129.2, 129.0, 128.8, 128.5, 128.4, 128.3, 128.2, 128.1, 128.0, 127.8, 127.8, 127.7, 127.6, 127.3, 124.6, 119.3, 116.6, 103.8, 95.9, 92.4, 81.9, 78.5, 77.6, 74.9, 74.8, 74.2, 73.7, 73.7, 73.3, 73.2, 72.2, 71.9, 71.7, 71.6, 68.7, 68.1, 68.0, 66.7, 64.8, 54.7, 20.8, 20.8, 20.7, 15.4. HRMS (ESI) m/z: found [M + Na]⁺ 1351.3539, C₆₅H₇₀Cl₃F₃N₂O₁₈ calcd. for [M + Na]⁺ 1351.3539.

tert-Butyldiphenylsilyl 2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranoside (14). A mixture of Ir(COD)(PMe₂Ph)₂PF₆ (14.2 mg, 16.8 μmol) in THF (14.0 mL) was stirrede at room temperature for 15 min under H₂, and the atmosphere was replaced by Ar. To the mixture of activated Ir complex in THF was added a solution of 11 (781 mg, 0.56 mmol) in THF (14.0 mL) under Ar, and stirred for 30 min at room temperature. The reaction mixture was concentrated. The residue was dissolved with 90% acetone aq. (28.0 mL). To the solution were added HgCl₂ (380 mg, 1.40 mmol) and HgO (48.5 mg, 0.22 mmol), and stirred for 1 h at room temperature. The reaction mixture was diluted with CHCl₃ and water, and extracted with CHCl₃. The combined organic layer was successively washed with 10% KI aq., water, and brine, dried over Na₂SO₄, and concentrated. The crude product was purified by silica gel column chromatography with hexane–EtOAc (75:25) and gel permeation chromatography (LH-20, CHCl₃–MeOH (50:50)) to give the title product 14 (727 mg, 96%). [α]_D −41.9° (c 1.1, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.68–7.14 (m, 30H, Ar), 6.85 (d, 1H, J_2,NH = 7.2 Hz, NH), 5.24–5.18 (m, 4H, H-1^Fuc, H-2^Fuc, H-3^Fuc, H-4^Fuc), 5.10 (dd, 1H, J_4,5 = 12.9 Hz, J_5,6 = 6.4 Hz, H-5^Fuc), 5.05 (d, 1H, J_1,2 = 6.7 Hz, H-1^GlcN), 4.80 (d, 1H, J_gem = 11.1 Hz, PhCH₂), 4.76 (d, 1H, PhCH₂), 4.64 (d, 1H, J_gem = 11.2 Hz, PhCH₂), 4.58–4.52 (m, 3H, PhCH₂ × 3), 4.47 (d, 1H, J_1,2 = 7.8 Hz, H-1^Gal), 4.38 (d, 1H, J_gem = 12.5 Hz, PhCH₂), 4.32 (d, 1H, PhCH₂), 4.22 (t, 1H, J_2,3 = J_3,4 = 9.3 Hz, H-3^GlcN), 3.91 (t, 1H, J_4,5 = 9.3 Hz, H-4^GlcN), 3.83–3.79 (m, 3H, H-4^Gal, H-6a^Gal, H-6b^Gal), 3.51–3.43 (m, 2H, H-3^Gal, H-5^Gal), 3.36–3.32 (m, 2H, H-2^GlcN, H-2^Gal), 3.25 (dd, 1H, J_5,6a =2.5 Hz, J_gem = 11.0 Hz, H-6a^GlcN), 3.17 (dd, 1H, J_5,6b = 1.4 Hz, H-6b^GlcN), 2.98–2.96 (m, 1H, H-5^GlcN), 2.17 (s, 3H, Ac), 2.10 (d, 1H, J_3,OH = 6.8 Hz, OH), 1.99 (s, 3H, Ac), 1.96 (s, 3H, Ac), 1.16 (s, 9 H, ^tBu), 0.89 (d, 3H, H-6^Fuc); ¹³C-NMR (125 MHz, CDCl₃) δ 170.5, 170.2, 169.4, 161.1, 138.8, 138.4, 138.1, 137.9, 135.9, 135.8, 133.2, 132.6, 129.7, 129.7, 128.9, 128.6, 128.5, 128.3, 128.3, 128.3, 128.1, 128.1, 127.9, 127.6, 127.5, 127.5, 127.3, 102.8, 95.4, 93.6, 92.2, 81.2, 77.6, 75.8, 75.4, 75.3, 75.3, 74.6, 74.1, 73.4, 73.2, 73.2, 72.7, 71.7, 68.1, 68.1, 66.8, 64.3, 62.8, 26.9, 20.8, 20.8, 20.8, 19.2, 15.6. HRMS (ESI) m/z: found [M + Na]⁺ 1378.4108, C₇₀H₈₀Cl₃NO₁₈Si calcd. for [M + Na]⁺ 1378.4108.

tert-Butyldiphenylsilyl 3-O-allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranoside (15). To a mixture of the glycosyl donor 13 (343 mg, 0.26 mmol), the glycosyl acceptor 14 (234 mg, 0.17 mmol), and molecular sieves AW-300 (516 mg) in CH₂Cl₂ (5.7 mL) was added dropwise TMSOTf (3.1 μL, 17.2 μmol) at −40 °C under Ar, and stirred for 20 h at −40 °C. The reaction mixture was quenched with sat. NaHCO₃, filtered through Celite, and diluted with CHCl₃. The organic layer was separated, and the aqueous layer was extracted with CHCl₃. The combined organic layer was successively washed with water and brine, dried over Na₂SO₄, and concentrated. The crude product was purified by gel permeation chromatography [S-X1, toluene–EtOAc (75:25)] and silica gel column chromatography with toluene–EtOAc (89:11) to give the title product 15 (399 mg, 93%). [α]_D −58.2° (c 1.4, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.65–6.98 (m, 50H, Ar), 6.62–6.60 (m, 2H, NH × 2), 5.92–5.84 (m, 1H, H₂C=CHCH₂), 5.47 (d, 1H, J_1,2 = 7.2 Hz, H-1^GlcN), 5.32–4.99 (m, 13H, H-1^GlcN, H-1^Fuc × 2, H-2^Fuc × 2, H-3^Fuc × 2, H-4^Fuc × 2, H-5^Fuc × 2, H₂C=CHCH₂ × 2), 4.78–4.70 (m, 5H, PhCH₂ × 5), 4.64 (d, 1H, J_gem = 10.7 Hz, PhCH₂), 4.60–4.52 (m, 3H, PhCH₂ × 3), 4.50–4.40 (m, 6H, PhCH₂ × 5, H-1^Gal), 4.36–4.32 (m, 2H, PhCH₂, H-1^Gal), 4.29–4.17 (m, 3H, PhCH₂, H-3^GlcN × 2), 4.14–4.06 (m, 2H, H₂C=CHCH₂ × 2), 4.00 (t, 1H, J_3,4 = J_4,5 = 9.4 Hz, H-4^GlcN), 3.90 (d, 1H, J_3,4 = 2.3 Hz, H-4^Gal), 3.85–3.77 (m, 5H, H-4^Gal, H-4^GlcN, H-3^Gal, H-6a^Gal × 2), 3.70–3.69 (m, 2H, H-6b^Gal × 2), 3.61–3.57 (m, 3H, H-2^Gal × 2, H-6a^GlcN), 3.50–3.38 (m, 5H, H-6b^GlcN, H-2^GlcN, H-5^GlcN, H-5^Gal × 2), 3.26 (dd, 1H, J_2,3 = 9.7 Hz, J_3,4 = 2.6 Hz, H-3^Gal), 3.22 (dd, 1H, J_5,6a = 2.4 Hz, J_gem = 11.3 Hz, H-6a^GlcN), 3.12–3.10 (m, 1H, H-6b^GlcN), 2.98–2.90 (m, 2H, H-2^GlcN, H-5^GlcN), 2.16 (s, 3H, Ac), 2.13 (s, 3H, Ac), 2.08 (s, 3H, Ac), 2.02 (s, 3H, Ac), 1.97 (s, 3H, Ac), 1.93 (s, 3H, Ac), 1.00 (s, 9H, ^tBu), 0.79 (d, 3H, J_5,6 = 6.5 Hz, H-6^Fuc), 0.74 (d, 3H, J_5,6 = 6.5 Hz, H-6^Fuc); ¹³C-NMR (125 MHz, CDCl₃) δ 170.5, 170.3, 170.2, 169.4, 169.3, 161.0, 160.8, 139.2, 138.7, 138.5, 138.4, 138.3, 138.0, 137.5, 135.8, 135.7, 134.8, 133.2, 132.6, 129.7, 129.1, 129.0, 128.9, 128.7, 128.4, 128.3, 128.3, 128.2, 128.2, 128.1, 127.9, 127.8, 127.7, 127.6, 127.5, 127.4, 127.3, 116.7, 103.2, 103.2, 95.6, 95.5, 92.2, 92.1, 81.8, 81.2, 79.3, 77.6, 76.1, 75.6, 75.4, 75.2, 75.1, 74.9, 74.6, 74.5, 73.5, 73.3, 73.1, 73.0, 72.9, 72.8, 72.4, 71.7, 71.7, 71.4, 68.2, 68.2, 68.1, 68.0, 66.8, 66.7, 64.5, 64.3, 26.9, 20.9, 20.8, 20.8, 20.7, 19.1, 15.3. HRMS (ESI) m/z: found [M + Na]⁺ 2517.7349, C₁₂₇H₁₄₄Cl₆N₂O₃₅Si calcd. for [M + Na]⁺ 2517.7348.

3-O-Allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-d-glucopyranose (16). Compound 15 (453 mg, 0.18 mmol) was desilylated with 1 M TBAF/THF (0.72 mL, 0.72 mmol) and AcOH (0.10 mL, 1.81 mmol) in THF (3.6 mL) as described for 12. The crude product was purified by gel permeation chromatography [S-X1, toluene–EtOAc (75:25)] and silica gel column chromatography with toluene-EtOAc (67:33) to give the title product 16 (395 mg, 97%). Analysis of compound 16 was too difficult for anomeric isomer, so the product was analyzed in next step. HRMS (ESI) m/z: found [M + Na]⁺ 2279.6171, C₁₁₁H₁₂₆Cl₆N₂O₃₅ calcd for [M + Na]⁺ 2279.6170.

3-O-Allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-d-gluco-pyranosyl (N-phenyl)-2,2,2-trifuoroacetimidate (17). Compound 16 (97 mg, 42.9 μmol) was reacted with (N-phenyl)-2,2,2-trifluoroacetoimidoyl chloride (17.8 mg, 85.8 μmol) and K₂CO₃ (29.7 mg, 215 μmol) in acetone (1.7 mL) as described for 13. The crude product was purified by silica gel column chromatography with hexane–EtOAc (83:17) and gel permeation chromatography [S-X1, toluene–EtOAc (75:25)] to give the title product 17 (94.7 mg, 91%). [α]_D −35.2° (c 1.3, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.38–7.07 (m, 43H, Ar), 6.82 (d, 1H, J_2,NH = 9.4 Hz, NH), 6.71–6.69 (m, 2H, Ar), 6.26 (br, 1H, H-1^GlcN), 6.00 (d, 1H, J_2,NH = 4.7 Hz, NH), 5.88–5.81 (m, 1H, H₂C=CHCH₂), 5.30–5.14 (m, 10H, H₂C=CHCH₂ × 2, H-1^Fuc × 2, H-2^Fuc × 2, H-3^Fuc × 2, H-4^Fuc × 2), 5.07 (d, 1H, J_1,2 = 8.1 Hz, H-1^GlcN), 5.03–5.00 (m, 2H, H-5^Fuc, PhCH₂), 4.94 (dd, 1H, J_4,5 = 12.7 Hz, J_5,6 = 6.4 Hz, H-5^Fuc), 4.78 (d, 1H, J_gem = 11.6 Hz, PhCH₂), 4.71 (d, 1H, J_gem = 11.2 Hz, PhCH₂), 4.63 (d, 1H, J_gem = 11.9 Hz, PhCH₂), 4.60–4.37 (m, 13H, PhCH₂ ×11, H-1^Gal, H-2^GlcN), 4.32 (d, 1H, J_1,2 = 7.8 Hz, H-1^Gal), 4.20 (t, 1H, J_2,3 = J_3,4 = 9.2 Hz, H-3^GlcN), 4.07–4.01 (m, 3H, H₂C=CHCH₂ × 2, H-4^GlcN), 3.93–3.77 (m, 8H, H-4^GlcN, H-2^GlcN, H-6a^Gal, H-6a^Gal, H-4^Gal, H-5^GlcN, H-4^Gal, H-6b^Gal), 3.76–3.65 (m, 3H, H-3^Gal, H-3^GlcN, H-2^Gal), 3.62–3.52 (m, 5H, H-6a^GlcN, H-6b^GlcN, H-6a^GlcN, H-6b^Gal, H-2^Gal), 3.49–3.44 (m, 2H, H-5^Gal, H-6b^GlcN), 3.38 (dd, 1H, J_5,6a = 5.5 Hz, J_5,6b = 7.5 Hz, H-5^Gal), 3.21–3.18 (m, 2H, H-3^Gal, H-5^GlcN), 2.16 (s, 3H, Ac), 2.15 (s, 3H, Ac), 2.14 (s, 3H, Ac), 2.05 (s, 3H, Ac), 2.01 (s, 6H, Ac × 2), 0.85 (d, 3H, H-6^Fuc), 0.69 (d, 3H, J_5,6 = 6.4 Hz, H-6^Fuc); ¹³C-NMR (125 MHz, CDCl₃) δ 170.4, 170.3, 170.2, 170.1, 169.4, 160.9, 160.7, 142.7, 139.0, 138.8, 138.4, 138.3, 138.2, 138.1, 137.8, 137.6, 137.3, 134.9, 129.3, 129.0, 129.0, 128.8, 128.5, 128.4, 128.4, 128.3, 128.2, 128.2, 128.2, 128.1, 128.1, 127.9, 127.8, 127.7, 127.7, 127.6, 127.5, 127.3, 127.3, 125.2, 124.6, 119.2, 117.0, 116.6, 114.8, 103.4, 103.2, 99.4, 96.0, 95.6, 93.3, 92.6, 92.4, 81.8, 80.8, 78.5, 78.1, 77.6, 76.5, 75.1, 74.9, 74.7, 74.6, 74.4, 74.3, 74.1, 73.4, 73.3, 73.2, 72.9, 72.8, 72.4, 72.0, 71.6, 71.5, 71.4, 68.8, 68.5, 68.2, 68.1, 68.0, 67.9, 67.1, 66.7, 64.8, 64.5, 58.7, 54.6, 30.9, 29.6, 21.4, 20.9, 20.8, 20.8, 20.7, 20.7, 15.7, 15.2, 14.1. HRMS (ESI) m/z: found [M + Na]⁺ 2450.6466, C₁₁₉H₁₃₀Cl₆F₃N₃O₃₅ calcd. for [M + Na]⁺ 2450.6466.

tert-Butyldiphenylsilyl 2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranoside (18). Compound 15 (104 mg, 41.6 μmol) was deallylated by Ir(COD)(PMe₂Ph)₂PF₆ (1.1 mg, 1.25 μmol) in THF (1.0 mL × 2) and deprotected by HgCl₂ (28.2 mg, 104 μmol) and HgO (3.6 mg, 16.6 μmol) with 90% acetone aq. (2.1 mL) as described for 14. The crude product was purified by silica gel column chromatography with hexane–EtOAc (83:17) and gel permeation chromatography [S-X1, toluene–EtOAc (75:25)] to give the title product 18 (92.2 mg, 90%). [α]_D −46.2° (c 1.0, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.65–7.12 (m, 50H, Ar), 6.63 (d, 1H, J_2,NH = 7.3 Hz, NH), 6.27 (br, 1H, NH), 5.33–5.08 (m, 10H, H-1^GlcN, H-1^Fuc × 2, H-2^Fuc × 2, H-3^Fuc × 2, H-4^Fuc × 2, H-5^Fuc), 5.00 (dd, 1H, J_4,5 = 12.3 Hz, J_5,6 = 6.4 Hz, H-5^Fuc), 4.91 (br, 1H, H-1^GlcN), 4.84–4.82 (m, 2H, PhCH₂ × 2), 4.74 (d, 1H, J_gem = 10.9 Hz, PhCH₂), 4.67 (d, 1H, J_gem = 11.3 Hz, PhCH₂), 4.62–4.53 (m, 6H, PhCH₂ × 6), 4.51–4.46 (m, 3H, PhCH₂ × 3), 4.43–4.28 (m, 5H, H-1^Gal × 2, PhCH₂ × 3), 4.15–4.12 (m, 1H, H-3^GlcN), 4.04–4.00 (m, 2H, H-3^GlcN, H-4^GlcN), 3.90–3.87 (m, 2H, H-4^Gal, H-6a^Gal), 3.85–3.81 (m, 3H, H-6b^Gal, H-4^Gal, H-4^GlcN), 3.79–3.68 (m, 4H, H-3^Gal, H-6a^Gal, H-6b^Gal, H-2^GlcN), 3.64–3.60 (m, 2H, H-2^Gal, H-6a^GlcN), 3.52 (dd, 1H, J_5,6b = 2.0 Hz, J_gem = 10.7 Hz, H-6b^GlcN), 3.49–3.40 (m, 3H, H-5^Gal × 2, H-3^Gal), 3.36–3.32 (m, 2H, H-5^GlcN, H-2^Gal), 3.24–3.12 (m, 3H, H-6^GlcN × 2, H-2^GlcN), 2.92 (d, J_4,5 = 9.6 Hz, H-5^GlcN), 2.18 (s, 3H, Ac), 2.14 (s, 3H, Ac), 2.09 (s, 3H, Ac), 2.03 (s, 3H, Ac), 2.00 (d, 1H, J_3,OH = 5.2 Hz, OH), 1.98 (s, 3H, Ac), 1.93 (s, 3H, Ac), 1.00 (s, 9H, ^tBu), 0.84 (d, 3H, J_5,6 = 6.4 Hz, H-6^Fuc), 0.76 (d, 3H, H-6^Fuc); ¹³C-NMR (125 MHz, CDCl₃) δ 170.5, 170.3, 170.2, 170.1, 169.5, 169.3, 161.0, 160.7, 139.2, 138.6, 138.5, 138.4, 138.2, 138.1, 137.9, 137.8, 137.4, 135.8, 135.7, 133.1, 132.5, 129.7, 129.6, 129.1, 129.0, 129.0, 128.7, 128.4, 128.4, 128.3, 128.3, 128.2, 128.2, 128.1, 128.0, 128.0, 127.8, 127.8, 127.8, 127.7, 127.6, 127.5, 127.4, 127.4, 127.3, 125.3, 103.1, 99.0, 95.7, 95.5, 93.6, 92.4, 92.3, 81.5, 80.1, 77.6, 77.5, 76.2, 75.3, 75.2, 75.1, 75.1, 75.0, 74.9, 74.9, 74.7, 74.1, 73.9, 73.7, 73.5, 73.1, 73.1, 72.9, 72.9, 72.8, 71.6, 71.6, 68.2, 68.1, 68.1, 68.0, 67.9, 66.8, 66.6, 64.4, 64.3, 62.2, 59.3, 31.9, 29.6, 29.3, 26.8, 26.7, 22.6, 21.4, 20.9, 20.8, 20.8, 20.7, 20.7, 19.1, 18.8, 15.6, 15.4, 14.1. HRMS (ESI) m/z: found [M + Na]⁺ 2477.7035, C₁₂₄H₁₄₀Cl₆N₂O₃₅Si calcd. for [M + Na]⁺ 2477.7035.

tert-Butyldiphenylsilyl 3-O-allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranoside (19). To a mixture of the glycosyl donor 17 (276 mg, 113 μmol), the glycosyl acceptor 18 (210 mg, 85.0 μmol), and molecular sieves AW-300 (255 mg) in CH₂Cl₂ (2.8 mL) was added dropwise TMSOTf (3.0 μL, 17.0 μmol) at 0 °C under Ar, and stirred for 1 h at 0 °C. The reaction mixture was quenched with sat. NaHCO₃, filtered through Celite, and diluted with CHCl₃. The organic layer was separated, and the aqueous layer was extracted with CHCl₃. The combined organic layer was successively washed with brine, dried over Na₂SO₄, and concentrated. The crude product was purified by gel permeation chromatography [S-X1, toluene–EtOAc (75:25)] and silica gel column chromatography with toluene–EtOAc (83:17) to give the title product 19 (352 mg, 88%). [α]_D −50.5° (c 1.0, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.65–7.02 (m, 90H, Ar), 6.67 (d, 1H, J_2,NH = 7.1 Hz, NH), 6.27 (brs, 1H, NH), 5.97 (m, 2H, NH × 2), 5.86-5.78 (m, 1H, H₂C=CHCH₂), 5.43–4.87 (m, 26H, H-1^Fuc × 4, H-2^Fuc × 4, H-3^Fuc × 4, H-4^Fuc × 4, H-5^Fuc × 4, H-1^GlcN × 4, H₂C=CHCH₂ × 2), 4.84–4.27 (m, 37H, H-1^Gal × 4, H-2^GlcN, PhCH₂ × 32), 4.22–3.12 (m, 48H, H-2^Gal × 4, H-3^Gal × 4, H-4^Gal × 4, H-5^Gal × 4, H-6a^Gal × 4, H-6b^Gal × 4, H-2^GlcN × 3, H-3^GlcN × 4, H-4^GlcN × 4, H-5^GlcN × 3, H-6a^GlcN × 4, H-6b^GlcN × 4, H₂C=CHCH₂ × 2), 2.93 (d, 1H, J_4,5 = 9.6 Hz, H-5^GlcN), 2.21–1.88 (m, 36H, Ac × 12), 1.01 (s, 9H, ^tBu), 0.89–0.67 (m, 12H, H-6^Fuc × 4); ¹³C-NMR (125 MHz, CDCl₃) δ 171.1, 170.5, 170.4, 170.3, 170.2, 170.2, 169.6, 169.5, 169.4, 160.9, 160.7, 139.4, 139.2, 138.9, 138.6, 138.6, 138.5, 138.5, 138.4, 138.0, 137.9, 137.7, 137.6, 137.5, 136.0, 135.8, 135.0, 133.2, 132.6, 130.0, 129.8, 129.3, 129.2, 129.2, 129.1, 128.8, 128.7, 128.6, 128.5, 128.5, 128.4, 128.4, 128.3, 128.3, 128.2, 128.1, 128.1, 128.0, 127.9, 127.8, 127.8, 127.8, 127.7, 127.6, 127.6, 127.4, 125.4, 116.7, 103.5, 103.3, 103.1, 99.2, 99.1, 98.9, 96.0, 95.9, 95.7, 95.6, 93.7, 92.8, 92.6, 92.4, 81.9, 81.5, 81.4, 78.7, 77.6, 77.4, 76.6, 76.2, 75.6, 75.6, 75.4, 75.3, 75.3, 75.1, 75.0, 75.0, 74.8, 74.5, 74.1, 74.0, 73.9, 73.6, 73.3, 73.3, 73.3, 73.1, 73.0, 73.0, 72.9, 72.6, 72.8, 71.8, 71.7, 68.6, 68.5, 68.3, 68.2, 68.1, 67.0, 66.8, 64.6, 64.5, 64.4, 62.4, 60.4, 59.4, 58.9, 32.0, 29.7, 29.4, 27.0, 22.8, 21.5, 21.1, 21.0, 20.9, 20.9, 20.8, 20.8, 19.2, 15.8, 15.6, 15.4, 14.3, 14.2. HRMS (ESI) m/z: found [1/2M + Na]⁺ 2369.6540, C₂₃₅H₂₆₄Cl₁₂N₄O₆₉Si calcd for [1/2M + Na]⁺ 2369.6544.

3-O-Allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-d-glucopyranose (20). Compound 19 (324 mg, 68.9 μmol) was desilylated with 1 M TBAF/THF (0.28 mL, 0.28 mmol) and AcOH (39.0 μL, 0.69 mmol) in THF (1.4 mL) as described for 12. The crude product was purified by gel permeation chromatography [S-X1, toluene–EtOAc (75:25)] and silica gel column chromatography with toluene–EtOAc (71:29) to give the title product 20 (298 mg, 97%). [α]_D −42.5° (c 1.0, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.65–7.02 (m, 83H, Ar, NH × 3), 6.70 (d, 1H, J_2,NH = 9.9 Hz, NH), 5.86–5.79 (m, 1H, H₂C=CHCH₂), 5.63–3.11 (m, 112H, H-1^Gal × 4, H-2^Gal × 4, H-3^Gal × 4, H-4^Gal × 4, H-5^Gal × 4, H-6a^Gal × 4, H-6b^Gal × 4, H-1^GlcN × 4, H-2^GlcN × 4, H-3^GlcN × 4, H-4^GlcN × 4, H-5^GlcN × 4, H-6a^GlcN × 4, H-6b^GlcN × 4, H-1^Fuc × 4, H-2^Fuc × 4, H-3^Fuc × 4, H-4^Fuc × 4, H-5^Fuc × 4, H₂C=CHCH₂ × 2, H₂C=CHCH₂ × 2, PhCH₂ × 32), 2.17–1.99 (m, 36H, Ac × 12), 0.89–0.67 (m, 12H, H-6^Fuc × 4); ¹³C-NMR (125 MHz, CDCl₃) δ 170.4, 170.3, 170.3, 170.3, 170.2, 170.2, 169.5, 169.5, 169.4, 160.7, 160.3, 139.5, 139.5, 139.4, 138.9, 138.5, 138.4, 138.4, 138.4, 138.3, 138.2, 138.1, 137.8, 137.6, 137.5, 137.5, 137.4, 134.9, 129.1, 129.1, 129.0, 128.7, 128.6, 128.5, 128.4, 128.4, 128.4, 128.3, 128.3, 128.2, 128.2, 128.2, 128.1, 128.1, 128.1, 128.0, 127.8, 127.8, 127.7, 127.7, 127.7, 127.7, 127.6, 127.6, 127.6, 127.4, 127.3, 127.3, 127.3, 127.2, 125.3, 116.5, 103.5, 103.4, 103.1, 99.4, 99.3, 99.0, 96.0, 95.8, 95.6, 93.0, 92.9, 92.8, 92.8, 92.6, 91.2, 82.0, 81.8, 81.6, 78.3, 77.6, 76.6, 76.5, 75.7, 75.5, 75.4, 75.3, 75.1, 75.0, 74.8, 74.8, 74.7, 74.7, 74.6, 74.3, 74.2, 74.1, 74.0, 73.4, 73.3, 73.2, 73.2, 73.0, 72.8, 72.8, 72.4, 71.7, 71.6, 71.6, 70.8, 68.6, 68.5, 68.2, 68.1, 68.0, 67.2, 66.9, 66.8, 64.6, 64.4, 57.8, 57.6, 55.6, 29.7, 29.3, 29.3, 21.4, 20.9, 20.9, 20.8, 20.8, 15.8, 15.7, 15.6, 15.5, 15.2, 14.1. HRMS (ESI) m/z: found [1/2M + Na]⁺ 2250.5955, C₂₁₉H₂₄₆Cl₁₂N₄O₆₉ calcd for [1/2M + Na]⁺ 2250.5955.

3-O-Allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-d-glucopyranosyl (N-phenyl)-2,2,2-trifuoroacetimidate (21). Compound 20 (761 mg, 0.17 mmol) was reacted with (N-phenyl)-2,2,2-trifluoroacetoimidoyl chloride (55.0 μL, 0.34 mmol) and K₂CO₃ (117 mg, 0.85 mmol) in acetone (6.8 mL) as described for 13. The crude product was purified by silica gel column chromatography with hexane–EtOAc (83:17) to give the title product 21 (693 mg, 88%). Analysis of compound 21 was too difficult for a few isomers, so the product was analyzed in the next step. ¹H-NMR of product 21 could not be assigned because all peaks were shown as broad peaks in all range. ¹³C-NMR (125 MHz, CDCl₃) δ 170.4, 170.3, 170.2, 170.2, 170.2, 170.2, 169.5, 169.5, 160.8, 160.7, 160.4, 142.7, 139.4, 139.3, 139.1, 138.8, 138.5, 138.4, 138.4, 138.3, 138.3, 138.2, 138.1, 137.8, 137.6, 137.5, 137.5, 137.3, 134.9, 129.2, 129.1, 129.0, 129.0, 128.8, 128.6, 128.5, 128.4, 128.4, 128.3, 128.3, 128.2, 128.2, 128.1, 127.9, 127.8, 127.7, 127.6, 127.6, 127.6, 127.6, 127.4, 127.4, 127.3, 127.3, 127.2, 125.3, 124.7, 119.2, 117.0, 116.5, 114.8, 103.5, 103.4, 103.1, 99.3, 99.2, 99.0, 96.1, 95.8, 95.8, 95.6, 93.4, 92.9, 92.7, 92.4, 81.8, 81.6, 78.4, 77.6, 76.6, 76.5, 76.4, 75.6, 75.5, 75.4, 75.3, 75.2, 75.0, 74.8, 74.8, 74.7, 74.3, 74.1, 74.1, 74.0, 73.4, 73.3, 73.2, 73.0, 72.9, 72.9, 72.8, 72.8, 72.4, 72.0, 71.7, 71.6, 71.5, 68.5, 68.1, 68.1, 68.0, 67.9, 67.0, 66.9, 66.8, 66.6, 64.7, 64.5, 64.4, 58.3, 57.9, 54.7, 31.9, 30.9, 29.7, 29.3, 22.7, 21.4, 20.9, 20.9, 20.8, 20.8, 20.7, 15.8, 15.7, 15.6, 15.2, 14.1. HRMS (ESI) m/z: found [1/2M + Na]⁺ 2336.1103, C₂₂₇H₂₅₀Cl₁₂F₃N₅O₆₉ calcd for [1/2M + Na]⁺ 2329.1103.

N-(tert-Butoxycarbonyl)-3-aminopropyl 3-O-allyl-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranoside (22). To a mixture of the glycosyl donor 21 (144 mg, 31.0 μmol), N-Boc-3-amino-1-propanol (53.0 μL, 310 μmol), and molecular sieves AW-300 (93.0 mg) in CH₂Cl₂ (1.0 mL) was added dropwise 1% TMSOTf in CH₂Cl₂ solution (56.0 μL, 3.1 μmol) at 0 °C under Ar, and stirred for 1 h at 0 °C. The reaction mixture was quenched with sat. NaHCO₃, filtered by Celite, and diluted with CHCl₃. The organic layer was separated, and the aqueous layer was extracted with CHCl₃. The combined organic layer was successively washed with brine, dried over Na₂SO₄, and concentrated. The crude product was purified by gel permeation chromatography [S-X1, toluene–EtOAc (75:25)] and silica gel column chromatography with toluene–EtOAc (71:29) to give the title product 22 (113 mg, 79%). [α]_D −51.8° (c 1.0, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.65–7.02 (m, 80H, Ar), 6.67 (d, 1H, J_2,NH = 6.5 Hz, NH), 6.19 (brs, 1H, NH), 6.02–5.78 (m, 3H, NH × 2, H₂C=CHCH₂), 5.31–4.87 (m, 26H, H-1^Fuc × 4, H-2^Fuc × 4, H-3^Fuc × 4, H-4^Fuc × 4, H-5^Fuc × 4, H-1^GlcN × 4, H₂C=CHCH₂ × 2), 4.84–4.27 (m, 46H, H-1^Gal × 4, H-2^GlcN × 3, H-3^GlcN × 3, H-4^GlcN × 3, PhCH₂ × 32, NHCH₂CH₂CH₂O), 4.20–3.25 (m, 38H, H-2^Gal × 4, H-3^Gal × 3, H-4^Gal × 4, H-5^Gal × 4, H-6a^Gal × 4, H-6b^Gal × 4, H-3^GlcN, H-4^GlcN, H-5^GlcN × 3, H-6a^GlcN × 3, H-6b^GlcN × 3, H₂C=CHCH₂ × 2, NHCH₂CH₂CH₂O × 2), 3.27–3.01 (m, 7H, H-3^Gal, H-2^GlcN, H-5^GlcN, H-6^GlcN × 2, NHCH₂CH₂CH₂O × 2), 2.21–1.95 (m, 36H, Ac × 12), 1.71–1.59 (m, 2H, NHCH₂CH₂CH₂O × 2), 1.42 (s, 9H, ^tBu), 0.84–0.67 (m, 12H, H-6 ^Fuc × 4). ¹³C-NMR (125 MHz, CDCl₃) δ 170.4, 170.3, 170.3, 170.2, 169.5, 169.5, 169.4, 161.2, 160.8, 160.7, 160.6, 156.0, 139.3, 139.1, 139.0, 138.9, 138.5, 138.5, 138.5, 138.4, 138.4, 138.4, 138.3, 137.7, 137.5, 137.5, 135.0, 129.2, 129.1, 129.0, 128.8, 128.6, 128.5, 128.5, 128.4, 128.3, 128.3, 128.2, 128.2, 128.2, 128.2, 128.1, 128.1, 128.0, 127.8, 127.8, 127.8, 127.7, 127.7, 127.7, 127.6, 127.5, 127.4, 116.6, 103.5, 103.1, 99.1, 98.5, 95.9, 95.9, 95.7, 92.7, 92.5, 92.3, 81.9, 81.3, 81.2, 79.2, 78.7, 77.9, 77.6, 76.5, 76.2, 75.5, 75.3, 75.2, 75.1, 75.1, 75.0, 74.9, 74.9, 74.4, 73.9, 73.8, 73.5, 73.3, 73.2, 73.2, 73.1, 73.0, 72.9, 72.5, 71.8, 71.7, 71.6, 68.5, 68.4, 68.3, 68.2, 68.1, 68.1, 67.3, 66.8, 64.6, 64.5, 37.3, 29.7, 28.5, 20.9, 20.8, 20.8, 20.8, 20.8, 15.7, 15.7, 15.5, 15.3. HRMS (ESI) m/z: found [1/2M + Na]⁺ 2329.1507, C₂₂₇H₂₆₁Cl₁₂N₅O₇₁ calcd. for [1/2M + Na]⁺ 2329.1506.

N-(tert-Butoxycarbonyl)-3-aminopropyl 2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-6-O-benzyl-2-deoxy-2-trichloroacetamido-β-d-glucopyranoside (23). Compound 22 (231 mg, 50.0 μmol) was deallylated by Ir(COD)(PMe₂Ph)₂PF₆ (1.3 mg, 1.50 μmol) in THF (2.5 mL × 2) and deprotected by HgCl₂ (34.0 mg, 125 μmol) and HgO (4.3 mg, 20.0 μmol) with 90% acetone aq. (5.0 mL) as described for 14. The crude product was purified by silica gel column chromatography with hexane–EtOAc (75:25) to give the title product 23 (219 mg, 96%). [α]_D −59.0° (c 1.0, CHCl₃); ¹H-NMR (500 MHz, CDCl₃) δ 7.65–7.02 (m, 80H, Ar), 6.82 (d, 1H, J_2,NH = 7.0 Hz, NH), 6.22 (brs, 1H, NH), 5.78–5.63 (m, 2H, NH × 2), 5.31–4.88 (m, 23H, H-1^Fuc × 4, H-2^Fuc × 4, H-3^Fuc × 4, H-4^Fuc × 4, H-5^Fuc × 4, H-1^GlcN × 4), 4.87–4.27 (m, 38 H, H-1^Gal × 4, H-1^GlcN, PhCH₂ × 32, NHCH₂CH₂CH₂O), 4.20 (m, 1H, H-3^GlcN), 4.03–3.02 (m, 52H, H-2^Gal × 4, H-3^Gal × 4, H-4^Gal × 4, H-5^Gal × 4, H-6a^Gal × 4, H-6b^Gal × 4, H-2^GlcN × 4, H-3^GlcN × 3, H-4^GlcN × 4, H-5^GlcN × 4, H-6a^GlcN × 4, H-6b^GlcN × 4, NHCH₂CH₂CH₂O × 2, NHCH₂CH₂CH₂O × 2), 2.21–1.95 (m, 37H, Ac × 12, OH), 1.71–1.59 (m, 2H, NHCH₂CH₂CH₂O × 2), 1.42 (s, 9H, ^tBu), 0.90–0.73 (m, 12H, H-6^Fuc × 4); ¹³C-NMR (125 MHz, CDCl₃) δ 170.5, 170.3, 170.3, 170.2, 170.2, 169.6, 169.6, 169.5, 169.4, 167.8 161.3, 161.0, 160.7, 160.5, 156.0, 139.5, 139., 139.0, 138.9, 138.6, 138.5, 138.5, 138.4, 138.3, 138.3, 138.3, 137.9, 137.7, 137.7, 137.6, 137.5, 132.5, 130.9, 129.2, 129.2, 129.1, 129.0, 128.8, 128.8, 128.6, 128.6, 128.5, 128.5, 128.4, 128.4, 128.3, 128.3, 128.3, 128.2, 128.2, 128.1, 128.1, 127.8, 127.8, 127.8, 127.8, 127.7, 127.7, 127.6, 127.5, 127.4, 125.3, 103.3, 103.0, 99.6, 99.2, 98.6, 95.9, 95.8, 95.7, 92.8, 92.7, 92.4, 81.9, 81.5, 81.3, 79.6, 79.2, 77.9, 76.3, 76.1, 75.5, 75.3, 75.2, 75.2, 75.0, 74.9, 74.7, 74.2, 74.1, 73.8, 73.8, 73.3, 73.3, 73.2, 73.2, 73.1, 73.0, 72.9, 71.7, 71.6, 68.7, 68.6, 68.6, 68.2, 68.1, 67.3, 66.8, 64.6, 64.5, 38.8, 37.2, 30.4, 29.7, 29.0, 28.5, 23.8, 23.0, 21.5, 21.0, 20.9, 20.9, 20.9, 20.8, 20.8, 15.8, 15.6, 14.1, 14.1. HRMS (ESI) m/z: found [1/2M + Na]⁺ 2309.1352, C₂₂₄H₂₅₇Cl₁₂N₅O₇₁ calcd. for [1/2M + Na]⁺ 2309.1350.

N-(tert-Butoxycarbonyl)-3-aminopropyl 2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-2-acetamido-6-O-benzyl-2-deoxy-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-2-acetamido-6-O-benzyl-2-deoxy-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-2-acetamido-6-O-benzyl-2-deoxy-β-d-glucopyranosyl-(1→3)-2,4,6-tri-O-benzyl-β-d-galactopyranosyl-(1→3)-[2,3,4-tri-O-acetyl-α-l-fucopyranosyl-(1→4)]-2-acetamido-6-O-benzyl-2-deoxy-β-d-glucopyranoside (25). A mixture of 23 (60.0 mg, 13.1 μmol), powdered Zn (1.71 g, 26.2 mmol), and AcOH (1.89 mL, 32.8 mmol) in THF (1.3 mL) was stirred under microwave irradiation at 250 W for 1 h under Ar. The mixture was diluted with CHCl₃ and filtered through Celite. The filtrate was successively washed with sat. NaHCO₃, water, and brine, dried over Na₂SO₄, and concentrated. The residue was chromatographed on silica gel with CHCl₃–MeOH (92:8). The crude product was dissolved in DMF (1.3 mL), and stirred with Boc₂O (4.3 μL, 20.0 μmol) and Et₃N (5.4 μL, 39.0 μmol) at room temperature for 1 d. The reaction mixture was concentrated, and purified by gel permeation chromatography [S-X1, toluene–EtOAc (75:25)] and silica gel column chromatography with CHCl₃–acetone (80:20–67:33) to give the title product 25 (41.0 mg, 76% in 2steps). ¹H-NMR of the product 25 could not be assigned because all peaks were shown as broad peaks in all range. [α]_D −6.5° (c 1.0, CHCl₃); ¹³C-NMR (125 MHz, CDCl₃) δ 170.6, 170.4, 170.3, 170.2, 169.5, 169.4, 169.4, 156.0, 139.1, 139.0, 138.8, 138.7, 138.6, 138.6, 138.2, 137.9, 137.8, 137.7, 137.6, 135.9, 129.4, 129.2, 129.1, 129.0, 129.0, 129.0, 128.7, 128.6, 128.6, 128.5, 128.4, 128.3, 128.2, 128.2, 128.2, 128.1, 128.1, 128.0, 128.0, 127.9, 127.7, 127.7, 127.6, 127.6, 127.6, 127.4, 127.3, 127.1, 127.1, 125.3, 103.4, 103.3, 103.2, 100.9, 100.5, 99.0, 95.8, 95.5, 80.5, 79.8, 79.3, 77.6, 76.4, 75.4, 75.2, 75.0, 74.7, 74.6, 74.3, 73.3, 73.3, 73.2, 73.1, 71.8, 68.2, 67.6, 66.7, 64.4, 37.1, 33.7, 32.8, 31.9, 30.2, 30.1, 29.7, 29.5, 29.4, 28.5, 27.1, 23.4, 23.2, 23.2, 22.7, 21.5, 20.8, 20.8, 15.6, 15.5, 15.4, 15.3, 14.1. HRMS (ESI) m/z: found [1/2M + Na]⁺ 2105.3693, C₂₂₄H₂₆₉N₅O₇₁ calcd for [1/2M + Na]⁺ 2105.3688.

4. Conclusions

We have developed a convenient synthesis of the fourth repeated Le^a tandem repeat framework. Le^a trisaccharide was synthesized by β-selective galactosylation and α-selective fucosylation with high selectivity. Glycosyl acceptors and donors of Le^a derivatives were obtained readily in a few steps, and the Le^a tandem repeat derivatives, the hexasaccharide and dodecasaccharide, were constructed in high yields. This convergent synthetic strategy can efficiently produce oligosaccharides with repeating structures, which will make an important contribution to biological studies.