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DOI: 10.1055/s-0029-1217343
Chemical N-Glycosylation by Asparagine under Integrated Microfluidic/Batch Conditions
Publication History
Publication Date:
02 June 2009 (online)

Abstract
An integrated microfluidic/batch system was applied to the chemical N-glycosylation by the asparagine amide group, a key glycosyl bond-formation reaction in the synthesis of N-glycopeptides. By applying the advantageous features of microfluidic conditions, that is, efficient mixing and rapid heat transfer, the GlcNTrocβAsn and the Fucα(1-6)GlcNTrocβAsn fragments were efficiently prepared.
Key words
N-glycosylation - asparagine - microreactor - oligosaccharide - N-glycopeptide
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References and Notes
IMM micromixer: http://www.imm-main2.de/
10Comet X-01 micromixer: http://homepage3.nifty.com/techno-applications/ or e-mail: yukio-matsubara@nifty.com.
11
Procedure of N-Glycosylation
Using an Integrated Microfluidic/Batch System
A
solution of TMSOTf (33 µL, 180 µmol, 43 mM) in
CH2Cl2 (4.2 mL) was injected, in advance,
into the micromixer by a syringe pump at a flow rate of 1.0 mL/min.
Then a solution of donor 1a (1.0 g, 1.1
mmol, 260 mM) and acceptor 2 (110 mg, 360 µmol,
86 mM) dissolved in CH2Cl2 (4.2 mL) was injected
into the IMM micromixer by another syringe pump at a flow rate of
1.0 mL/min. The reaction was mixed at r.t. After the reaction
mixture was allowed to flow at r.t. for an additional 94 s through
a Teflon tube reactor (Φ = 1.0
mm, l = 1.0 m), the mixture was introduced
into a flask, and stirred for 12 h at this temperature. Then the
mixture was quenched by an aq NaHCO3 solution. The resulting
mixture was extracted with EtOAc, washed with brine, dried over Na2SO4,
filtered, and concentrated in vacuo to give the
crude
product. The residue was purified by column chromatography on silica
gel (from 25-33% EtOAc in hexane) to give N-glycoside 3a as
a white solid (376 mg, 85%). ESI-MS: m/z calcd
for C53H53Cl3N3O13 [M + H]+: 1044.3;
found: 1044.2. ¹H NMR (500 MHz, CDCl3): δ = 7.76
(d, J = 7.6
Hz, 1 H), 7.73 (d, J = 7.5
Hz, 1 H), 7.59 (d, J = 7.6
Hz, 1 H), 7.55 (d, J = 7.4
Hz, 1 H), 7.41-7.17 (m, 19 H), 6.80 (d, J = 8.5
Hz, 1 H), 5.91 (d, J = 9.2
Hz, 1 H), 5.88-5.80 (m, 1 H), 5.27 (dd, J = 17.2,
1.3 Hz, 1 H), 5.19 (dd, J = 10.5,
1.3 Hz, 1 H), 5.14-5.04 (m, 4 H), 4.89 (dd, J = 9.2, 9.2
Hz, 1 H), 4.75 (d, J = 12.1
Hz, 1 H), 4.69 (d, J = 12.0
Hz, 1 H), 4.68-4.58 (m, 3 H), 4.52-4.35 (m, 6
H), 4.15 (dd, J = 6.9,
6.9 Hz, 1 H), 3.65-3.61 (m, 3 H), 3.56-3.49 (m,
2 H), 2.86 (dd, J = 16.7,
3.8 Hz, 1 H), 2,68 (dd, J = 16.4,
4.2 Hz, 1 H).
Data for 3b
ESI-MS: m/z calcd for C63H67Cl3N3O19 [M + H]+:
1274.3; found: 1274.2. ¹H NMR (500 MHz, CDCl3): δ = 7.76
(d, J = 7.6
Hz, 1 H), 7.73 (d, J = 7.5
Hz, 1 H), 7.61 (d, J = 7.6 Hz,
1 H), 7.60 (d, J = 7.3
Hz, 1 H), 7.41-7.19 (m, 19 H), 6.89 (d, J = 8.2
Hz, 1 H), 5.94 (d, J = 8.6
Hz, 1 H), 5.86-5.79 (m, 1 H), 5.31-5.25 (m, 3
H), 5.18 (d, J = 11.6
Hz, 1 H), 5.13 (d, J = 12.2
Hz, 1 H), 5.06 (d, J = 12.4
Hz, 1 H), 4.99 (d, J = 3.5 Hz,
1 H), 4.90-4.87 (m, 2 H), 4.76 (d, J = 12.2
Hz, 1 H), 4.68 (dd, J = 12.1,
4.0 Hz, 2 H), 4.62-4.52 (m, 10 H), 4.37 (d, J = 11.8 Hz,
1 H), 4.23 (dd, J = 6.7,
6.7 Hz, 1 H), 3.80 (dd, J = 10.2,
3.5 Hz,1 H), 3.71 (dd, J = 11.8,
2.2 Hz, 1 H), 3.67-3.52 (m, 3 H), 2.85 (dd, J = 16.5,
3.6 Hz, 1 H), 2.69 (dd, J = 16.5,
3.9 Hz, 1 H), 2.09 (s, 3 H), 1.96 (s, 3 H), 1.06 (d, J = 6.5 Hz,
3 H).