Removal of some common glycosylation by-products during reaction work-up

Article abstract of DOI:10.1016/j.carres.2016.12.007

With the aim of improving the general glycosylation protocol to facilitate easy product isolation it was shown that amide by-products from glycosylation with trichloroacetimidate and N-phenyl trifluoroacetimidate donors could be removed during reaction work-up by washing with a basic aqueous solution. Excess glycosyl acceptor or lactol originating from glycosyl donor hydrolysis could equally be removed from the reaction mixture by derivatization with a basic tag and washing with an acidic solution during reaction work-up.

Full text of DOI:10.1016/j.carres.2016.12.007

Accepted Manuscript
Removal of some common glycosylation by-products during reaction work-up
Mads Heuckendorff, Henrik H. Jensen
PII:
S0008-6215(16)30550-X
10.1016/j.carres.2016.12.007
CAR 7309
DOI:
Reference:
To appear in: Carbohydrate Research
Received Date: 17 November 2016
Revised Date: 22 December 2016
Accepted Date: 22 December 2016
Please cite this article as: M. Heuckendorff, H.H. Jensen, Removal of some common glycosylation by-
products during reaction work-up, Carbohydrate Research (2017), doi: 10.1016/j.carres.2016.12.007.
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ACCEPTED MANUSCRIPT
Removal of some common glycosylation by-products
during reaction work-up
Mads Heuckendorff and Henrik H. Jensen*
Department of Chemistry, Aarhus University, Langelandsgade 140, 8000 Aarhus C, Denmark
hhj@chem.au.dk
Abstract
With the aim of improving the general glycosylation protocol to facilitate easy product isolation it
was shown that amide by-products from glycosylation with trichloroacetimidate and N-phenyl
trifluoroacetimidate donors could be removed during reaction work-up by washing with a basic
aqueous solution. Excess glycosyl acceptor or lactol originating from glycosyl donor hydrolysis
could equally be removed from the reaction mixture by derivatization with a basic tag and washing
with an acidic solution during reaction work-up.
1

ACCEPTED MANUSCRIPT
Keywords
Tagging; trichloroacetamide; non-chromatographic purification
Introduction
Chromatographic purification of reaction mixtures is not always sufficient for obtaining a clean
[
1]
reaction product. This is especially true for glycosylation reactions where many by-products are
[
2]
customarily observed and a time consuming screening of chromatography eluents are not always
the solution. In the case of having chemically benign impurities, the experimentalist, however
unsatisfactory, could choose to carry out the next step and hope the subsequent purification would
be easier and report the yield over two synthetic steps.
There are many protocols in the literature to facilitate easier purification of synthetic
[
3]
oligosaccharides. Solid phase oligosaccharide synthesis inspired by similar techniques in
[
4]
oligopeptide and oligonucleotide synthesis, was championed by the Seeberger group with
[
5],[6]
contribution from other groups
is undoubtedly an elegant solution to an otherwise challenging
task of easily separating the product from by-products. One pronounced benefit of solid phase
techniques in comparison to solution phase techniques is the possibility of using a large excess of
reagents and repeated treatment with fresh reagents to push reactions to completion at little other
expense than having to perform a simple filtration after the reaction to remove all compounds not
attached to the solid support. Other approaches used in carbohydrate chemistry to assist product
[7]
[8]
[9]
[10]
purification are fluorous liquid /solid phase separations, ionic tags, PEG tag,
lipophilic
In common for all these methods is the need for special equipment, reagents and
knowledge, which is expensive and/or not accessible to many average carbohydrate chemists.
[
11]
tags.
2

ACCEPTED MANUSCRIPT
Figure 1.
Recently, we have developed a simple protocol for anomeric deacylations using the cheap bulk
chemical 3-(dimethylamino)-1-propylamine (DMAPA, 1), in which the by-product
(
AcNH(CH ) N(CH ) ) could be removed by a simple acid wash procedure of the organic phase in
2 3 3 2
a separatory funnel during reaction work-up. We furthermore found this reagent to be ideal for
[
12]
removing excess of electrophilic reagents like e.g. benzoyl chloride and toluenesulfonyl chloride.
Inspired by the success of DMAPA (1) in our laboratory, we decided to investigate whether other
common by-products in carbohydrate chemistry could be easily eliminated from the crude reaction
mixture in a straightforward fashion during the common aqueous work-up. Given the nature of the
typical glycosylation the glycosyl donor leaving group or a derivative thereof will unavoidably be
present after the reaction. Other typical by-products originate from donor hydrolysis product
[
12]
(reducing sugar/lactol) and excess/unreacted acceptor. As our previous work with DMAPA (1),
our goal was to develop a protocol using inexpensive commercially available chemicals and simple
procedures that could be used to remove the above mentioned by-products.
Purification and Discussion
[13]
[14]
The trichloroacetimidate
and the N-phenyl trifluoroacetimidate
donors are among the most
popular glycosyl donor types in carbohydrate chemistry, due to their mild and catalytic activation.
Glycosylation reactions using imidate donors, however, generates lipophilic amide by-products (5
and 6) originating from the leaving group (Figure 1). From time to time, these amide by-products
3

ACCEPTED MANUSCRIPT
can be difficult to remove from the product mixture due to their physical properties. Furthermore,
1
especially trichloroacetamide impurities can easily be overlooked since their detection by H-NMR
is compromised owing to the lack of protons. There are currently no reported procedures for
removing these amide by-products by means other than column chromatography, which due to the
compound’s tailing tendency can be challenging.
Our first objective was to investigate if trichloroacetamide (5) and N-phenyl-trifluoroacetamide (6)
could be removed from organic solvents by basic extraction since the amide protons are slightly
[
15]
acidic due to the electron withdrawing trichloromethyl and trifluoromethyl groups.
there has been an interest in having different substituents on the phenyl group of N-phenyl-
Previously,
[
16]
trifluoroimidate leaving groups.
For that reason we also decided to investigate the basic
extraction of N-phenyl-trifluoroacetamide having electron withdrawing p-chloro, p-cyano and p-
nitro groups.
Four different aqueous solutions with varying alkalinity and demineralized water itself were
explored for their ability to extract (or wash out) the amides 5 and 6 from CH Cl (sat. NaHCO ,
2
2
3
1
0% Na CO , 10% Na PO and 1 M NaOH). As seen from Table 1, the base strength of the solution
2 3 3 4
is crucial for the effectiveness of the extractions. Only 1 M NaOH (one extraction) and 10% aq.
Na PO (five extractions) were efficient for extracting trichloroacetamide and N-phenyl-
3
4
trifluoroacetamide. In the case of N-(p-cyanophenyl)trifluoroacetamide and N-(p-
nitrophenyl)trifluoroacetamide three extractions were sufficient with the milder base 10% aq.
Na CO . (For the complete extraction table; see supporting information).
2
3
One could speculate whether base labile functionalities like the acetyl protecting group would be
stable under the conditions used to remove amides 5 and 6. Accordingly, extraction of amides from
a CH Cl solution also containing glucose pentaacetate (7, Figure 1) was successfully conducted
2
2
without losing the carbohydrate to degradation and/or the aqueous layer (For complete list see
supporting information). Furthermore, it was also investigated whether the amides could be
recovered from the aqueous phase by acidification and three times back extraction with CH Cl and
2
2
thereby potentially be recycled by converting amides to imidoyl chlorides that can be used to
[
17]
synthesize new glycosyl donor. As listed in
4

ACCEPTED MANUSCRIPT
Table 1 especially N-phenyl-trifluoroacetamide (6) was nearly fully recovered.
Table 1. Extractions of amides (200 mg) from CH Cl (20 mL) extracted by aqueous solution
2
2
(20 mL) over 30 seconds.
Entry
Amide
Aqueous solution
Extracted from organic phase
number of extractions)
Recovered from aq. phase
by back extraction with
(
CH Cl2
2
5

ACCEPTED MANUSCRIPT
1
2
3
5
5
6
7
8
9
1
1
1
H O
66% (5)
2
Saturated NaHCO3
47% (3)
42% (3)
96% (3)
99% (5)
>99% (1)
5% (3)
10% Na CO3
2
10% Na PO4
3
10% Na PO4
16%
22%
3
1M NaOH
H O
2
Saturated NaHCO3
7% (3)
10% Na CO3
34% (3)
94% (3)
>99% (5)
>99% (1)
2
0
1
2
10% Na PO4
3
10% Na PO4
85%
89%
3
1M NaOH
Having established a protocol for eliminating trichloroacetamide (5) and N-phenyl-
trifluoroacetamide (6) from a CH Cl layer by extraction/washing, we moved on to investigate the
2
2
possibility of removing sugar alcohols (excess glycosyl acceptors) and lactols (hydrolyzed glycosyl
donors) by aqueous work-up. First, the commercially available glycine derivative
dimethylaminoacetyl chloride hydrochloride (DMAACl (2), Figure 1) was explored as an acylating
reagent, which in itself has an amine tail available for protonation. The tagging procedure was
tested on different substrates with varying lipophilicity and nature of the OH functional group
(
primary, secondary and lactol). In presence of Et N and 4-dimethylaminopyridine (DMAP),
3
DMAACl (2) reacted fully with 7-11 as judged by TLC analysis within hours at ambient
temperature. Other tagging procedures were also tested; first reaction between alcohol or lactol with
either POCl or acryloyl chloride (3) followed by reaction of the installed electrophilic site with
3
either DMAPA (1) or N-methyl-piperazine (4) (Scheme 1).
After complete reaction of the amine as judged by TLC analysis, the reaction mixture was
transferred to a separatory funnel. First, dilution with diethyl ether to give good separation of the
6

ACCEPTED MANUSCRIPT
two resulting phases and then washing of the organic layer three times with an aqueous 1 M HCl
solution generally removed the tagged saccharides.
Scheme 1. Removal of sugar alcohol by derivatization by reagents POCl , 1, 2, 3 or 4 and
3
washing with hydrochloric acid.
O
HO
Cl
O
H
4 equiv. Et N,
3
4 equiv. Et N,
1) 4 equiv. POCl3
3
N
4
0.2 equiv. DMAP
CH Cl rt. 2 h
0.2 equiv. DMAP
CH Cl rt.
or acryloyl chloride (3), 2 h
2) 10 equiv. DMAPA (1)
or 1-methylpiperazine (4), 2 h
Cl
equiv.
2)
2
2,
2
2,
(
O
(linker)
O
O
N
O
N
N
O
H
O
or
O
in CH Cl solution
(linker)
2
2
N
N
in CH Cl solution
2
2
three extractions
with aq. 1 M HCl
three extractions
with aq. 1 M HCl
Cl
Cl
H
O
O
H
(linker)
O
O
N
N
N
O
H
O
or
O
(
linker)
in aqueous layer
N
H
Cl
N
in aqueous layer
Table 2. Extraction (3 times with 1 M aq. HCl) of sugar alcohols and lactols from an organic
layer by amine tagging.
7

ACCEPTED MANUSCRIPT
Extracted from
organic phase
Entry
1
Sugar alcohol or lactol
Tagging reagents
DMAACl (2)
Et N, DMAP
*
>99%
3
1
2
1
) POCl , Et N, DMAP
3 3
2
3
>99%
>99%
) 1-Methylpiperazine (4)
) Acryloyl chloride (3),
Et N, DMAP
3
2
) 1-Methylpiperazine (4)
DMAACl (2)
4
>99%
Et N, DMAP
3
1
2
1
) POCl , Et N, DMAP
3 3
5
6
>91%
>99%
) DMAPA (1)
) Acryloyl chloride (3),
Et N, DMAP
3
2
) DMAPA (1)
DMAACl (2)
7
>99%
Et N, DMAP
3
1
2
1
) POCl , Et N, DMAP
8
9
3
3
>96%
>99%
) DMAPA
) Acryloyl chloride (3)
Et N, DMAP
3
2
) DMAPA (1)
DMAACl (2)
1
0
>99%
Et N, DMAP
3
1
) Acryloyl chloride (3)
1
1
1
1
2
3
>99%
>99%
>99%
Et N, DMAP
3
2
) 1-Methylpiperazine (4)
DMAACl (2)
Et N, DMAP
3
1
) Acryloyl chloride (3)
Et N, DMAP
3
2
1
) 1-Methylpiperazine (4)
) Acryloyl chloride (3)
1
4
Et N, DMAP
3
>99%
2
) 1-Methylpiperazine (4)
8

ACCEPTED MANUSCRIPT
1
2
) Acryloyl chloride (3)
1
5
Et N, DMAP
3
94%
) 1-Methylpiperazine (4)
As seen from Table 2 the amine tagging by e.g. acylation and subsequent acidic aqueous work-up
offers effective approach for removing both primary, secondary sugar alcohols and lactols from the
organic phase, this also brings the opportunity to recycle unreacted acceptor by subsequent back
extraction and deacylation. When having primary alcohols or lactols, the best results were achieved
by using the secondary amine 1-methylpiperazine (4) since the primary amine DMAPA (1) in some
cases cleaved the linker on the alcohol or lactol. Glycine derivative DMAACl (2) possessing only
one amine group was effective in removing most sugar alcohols and acetylated lactols, but going to
lactols with four benzyl (13) or benzoyl groups (14) (compared to acceptors with 3 benzyl groups, 9
and 10), then DMAACl (2) was not effective enough to transfer all material to the aqueous phase.
The electrophilic reagent POCl worked well with sugar alcohols of all kinds, but POCl was not a
3
3
good choice as a linker in the case of some lactols. We ascribe this to glycosyl chloride formation,
which was observed in some cases by TLC analysis. Instead acryloyl chloride (3) in combination
with 1-methylpiperazine (4) was capable of transferring lactols with four benzyl or benzoyl groups
(13 and 14, respectively) into the aqueous phase (See Table 2, entry 14 and 15).
Next, it was decided to explore the tagging/washing efficiency of excess/by-product sugar alcohols
and lactols in a sequence of reactions involving a glycosylation reaction. First, glucose pentaacetate
(7) was easily converted to the imidate donor (15) over two steps using our previously published
1
2
one-pot procedure. The glucosyl imidate (15) was then activated under standard conditions with a
catalytic amount of TMSOTf at -78 °C in CH Cl in presence of the acceptor 9 (Scheme 2). TLC
2
2
analysis of the reaction mixture after 1 hour showed no formation of a major new spot, which would
indicate either that no glycosylation had taken place, or that acceptor and disaccharide were co-
polar in the used eluent system. The tagging reagent DMAACl (2) was then added and the reaction
was stirred for another 2 hours at ambient temperature before then reaction mixture eventually
1
13
underwent aqueous work-up. H and C-NMR spectra of the crude reaction outcome can be seen in
Figure 2 both before and after the acidic washing step. The reaction resulted in isolation of the
[
18]
disaccharide 16
in good purity and in 88% yield after the acidic washing step of the organic
layer. In this particular case, the presented technique is especially powerful since excess acceptor
9

ACCEPTED MANUSCRIPT
and disaccharide product would be difficult to separate by column chromatography due to their
similar level of polarity (see result of TLC analysis in Supporting Information).
Scheme 2. Reaction sequence of glucosyl imidate formation, glycosylation, tagging and
aqueous work-up.
1.5 equiv. OH
1
) 0.1 equiv TMSOTf
CH Cl2, 3 Å mol. sieves
-78 οC, 1 h
O
BnO
BnO
2
BnO
OMe
OAc
O
OAc
O
OAc
O
1
) DMAPA
9
AcO
AcO
AcO
AcO
AcO
AcO
O
OAc 2) Cl CCN, DBU(cat.)
3
2) 8 equiv. Et N
3
Cl
O
BnO
BnO
AcO
AcO
AcO
CH
2
Cl
2
O
CCl3 0.2 equiv. DMAP
O
H
7
15
CH
2
Cl
2
, rt. 2 h
16
BnO
N
OMe
NH
Cl
7
8%
6 equiv.
88%
3) 3 times extractions with aq. 1 M HCl
1
time extraction with 1 M NaOH
13
1
Figure 2. C-NMR (left) and H-NMR (right) of the crude reaction mixture shown in Scheme 1 before tagging and aqueous work-
up and after tagging and aqueous work-up. No chromatography was used at any point.
Conclusion
1
0

ACCEPTED MANUSCRIPT
In conclusion, we have described a new protocol for removing amide by-products from
glycosylation reactions with trichloroacetimidate- and N-phenyltrifluoroacetimidate donors. We
have furthermore shown that cheap and commercial amines can be used for tagging either directly
or via an electrophilic linker to lactols and sugar alcohols. Despite having large lipophilic groups,
the amine tagged sugar molecules can be removed from the organic layer during reaction work-up
by washing with a hydrochloric acid solution. Contrary to the solid phase approach the present
protocol is easily scalable and could in principle be used in general organic chemistry.
Experimental Section
General Remarks
All reagents were used as purchased without further purification. Dry solvents were taken from a
solvent purification system. Glassware used for water-free reactions were dried for 12 h at 120 °C
before use. Columns were packed with silica gel 60 (230–400 mesh) as the stationary phase. TLC
1
13
plates were visualized by 10% H SO in EtOH and heating until spots appeared. H-NMR and C-
2
4
NMR spectra were recorded on a 400/100 MHz spectrometer, respectively. Chemical shifts (δ) are
reported in ppm relative to the residual solvent signal. High-resolution mass spectral (HRMS) data
were obtained on an electrospray (ES) mass spectrometer analyzing time-of-flight.
Extraction studies
Extraction of amides
The amide (200 mg) was dissolved in CH Cl (20 mL) in a separatory funnel. The organic phase
2
2
was washed thrice (3x30 seconds) or once (1x30 seconds), with base (20 mL). The organic phase
was dried over MgSO , filtered and evaporated. The residue in the flask was weighed to evaluate
4
the effect of the extraction.
Recovery of amide
The amide (200 mg) was dissolved in CH Cl (20 mL) in a separatory funnel. The organic phase
2
2
was washed five times (5x30 seconds) or once (1x30 seconds), with base (20 mL). The basic
aqueous fractions were immediately added to a flask containing enough 1M HCl to neutralize the
base. The combined aqueous fraction was extracted thrice with CH Cl (20 mL, 3x30 seconds). The
2
2
1
1

ACCEPTED MANUSCRIPT
combined organic fractions were dried over MgSO , filtered and evaporated. The residue in the
4
flask was weighed to evaluate the effect of the extraction.
Extraction of amides from a solution containing β-D-glucose pentaacetate.
The amide (0.51 mmol) and β- -glucose pentaacetate (0.51 mmol) were dissolved in CH Cl (20
D
2
2
mL) in a separatory funnel. The organic phase was washed 5 times (5x30 seconds) or once (1x30
seconds), with base (20 mL). The organic phase was dried over MgSO , filtered and evaporated.
4
The residue in the flask was weighed to evaluate the effect of the extraction.
Tagging and extractions with dimethylaminoacetyl chloride (DMAACl)
The sugar alcohol (0.1 mmol) and DMAP (0.02mmol) were dissolved in CH Cl (2 mL), then
2
2
triethylamine (0.4 mmol) and DMAACl (0.4 mmol) were added. The reaction mixture was stirred
for 2 hours and then transferred to a separatory funnel using Et O (20 mL). The organic layer was
2
extracted thrice with 1 M HCl (20 mL). The organic layer was dried over MgSO , filtered and
4
evaporated. The residue in the flask was weighed to evaluate the effect of the tagging and
extraction.
Tagging and extractions with POCl and DMAPA/1-methylpiperazine
3
The sugar alcohol (0.1 mmol) and DMAP (0.02mmol) were dissolved in CH Cl (2 mL), then
2
2
triethylamine (0.4 mmol) and POCl (0.4 mmol) were added. The reaction mixture was stirred for 2
3
hours and then added the amine (1 mmol). The reaction mixture was stirred for additional 2 hours
and the mixture was then transferred to a separatory funnel using Et O (20 mL). The organic layer
2
was extracted thrice with 1 M HCl (20 mL). The organic layer was dried over MgSO , filtered and
4
evaporated. The residue in the flask was weighed to evaluate the effect of the tagging and
extraction.
Tagging and extractions with acryloyl chloride and DMAPA/1-methylpiperazine
The sugar alcohol (0.1 mmol) and DMAP (0.02mmol) were dissolved in CH Cl (2 mL), then
2
2
triethylamine (0.4 mmol) and acryloyl chloride (0.4 mmol) were added. The reaction mixture was
stirred for 2 hours and then added the amine (1 mmol). The reaction mixture was stirred for
additional 2 hours and the mixture was then transferred to a separatory funnel using Et O (20 mL).
2
The organic layer was extracted thrice with 1 M HCl (20 mL). The organic layer was dried over
1
2

ACCEPTED MANUSCRIPT
MgSO , filtered and evaporated. The residue in the flask was weighed to evaluate the effect of the
4
tagging and extraction.
One pot anomeric deacetylation and imidate formation.
A solution of glucose pentaacetate (0.5 mmol) and DMAPA (2.5 mmol) in CH Cl (2.5 mL) was
2
2
stirred at room temperature. After 3 hours CCl CN (5 mmol) and DBU (0.01 mmol) were added to
3
the reaction mixture. The reaction was stirred for another 45 minutes. The reaction was diluted with
CH Cl and washed once with 1 M HCl (aq.) and saturated NaHCO (aq.). The organic phase was
2
2
3
dried over Na SO , filtered and evaporated. The crude product was purified by elution trough a
2
4
short silica plug. (pentane/EtOAc 1:1) giving the product.
Glycosylation, tagging and extraction
A mixture of glycosyl donor (0.10 mmol), glycosyl acceptor (0.15 mmol), and freshly activated
molecular sieves (3 Å, 100 mg) in CH Cl (2 mL) was stirred under argon for ½ h. The solution was
2
2
cooled to – 78 °C using a dry ice/acetone bath. TMSOTf (0.1 mL of a 0.1 M solution of TMSOTf in
CH Cl ) were added. The reaction was stirred for 1 hour at – 78 °C. Upon completion the reaction
2
2
mixture was added Et N (0.8 mmol), DMAP (0.02 mmol) and DMAACl (0.6 mmol). The flask was
3
removed from the cooling bath and stirred for 2 hours at rt. The reactions mixture was then
transferred to a separatory funnel using Et O (20 mL). The organic layer was extracted thrice with 1
2
M HCl (20 mL), once with 1 M NaOH (20 ml) and finally with brine (20 mL). The organic layer
was dried over MgSO , filtered and evaporated, giving the glycosylation product.
4
N-phenyl-2,2,2-trifluoroacetimidate (6)
Aniline (5 mL, 0.055 mol) and Et N (7.7 mL, 0.055mol) was dissolved in dry CH Cl (20mL). The
3
2
2
mixture was cooled to 0 °C and then was TFAA (7.6 mL, 0.055 mol) slowly added. The reaction
was allowed to reach rt. and stirred for 2 hours. The CH Cl was evaporated and the mixture
2
2
dissolved in EtOAc and transferred to a separatory funnel. The organic phase was washed thrice
with 1M HCl solution, then saturated sodium bicarbonate solution and brine. The organic phase was
dried over MgSO and evaporated to dryness. The solid was recrystallized from pentane/Et O
4
2
giving white fluffy crystals. 7.02 g, 68%, R 0.32 (pentane/EtOAc 10:1) M 86.2-89.2 °C, lit. 87.7
f
p
[19] 1
°
C.
H NMR (400 MHz, CDCl ) δ 7.85 (s, 1H, NH), 7.45 (d, J 7.7 Hz, 2H), 7.28 (t, J 8.0 Hz,
3 H
1
3
2
H), 7.14 (t, J 7.4 Hz, 1H). C NMR (100 MHz, CDCl ) δ 155.3 (q, J 37.5 Hz, C=O), 135.2,
3 C C-F
1
3

ACCEPTED MANUSCRIPT
1
9
1
29.4, 126.5, 120.9, 115.9 (q, JC-F 288.4 Hz, CF ). F NMR (376 MHz, CDCl ) δ -75.7. HRMS
3 3 F
+
(
ES): calcd. for C H F NONa 212.0294; found 212.0288. Spectral values were in accordance with
8 6 3
those reported in ref. 19.
N-(4-chlorophenyl)-2,2,2-trifluoroacetimidate
4
-chloroaniline (5 g, 0.039 mol) and Et N (5.5 mL, 0.039 mol) was dissolved in dry CH Cl (20
3
2
2
mL). The mixture was cooled to 0°C and then was TFAA (5.5 mL, 0.039 mol) slowly added. The
reaction was allowed to reach rt. and stirred for 2 hours. The CH Cl was evaporated and the
2
2
mixture dissolved in EtOAc and transferred to a separatory funnel. The organic phase was washed
thrice with 1M HCl solution, then saturated sodium bicarbonate solution and brine. The organic
phase was dried with MgSO and evaporated to dryness. The solid was recrystallized from
4
pentane/EtOAc giving white fluffy crystals. 7.8 g, 88%, R 0.30 (pentane/EtOAc 10:1) M 123.5-
f
p
[
15] 1
1
7
1
25 °C, lit. 123-124 °C.
H NMR (400 MHz, CDCl ) δ 8.43 (s, 1H, NH), 7.77 (d, J 8.6 Hz, 1H),
3 H
1
3
.70 (d, J 8.6 Hz, 1H). C NMR (100 MHz, DMSO-d ) δ 154.6 (q, J 37.1 Hz, C=O), 135.4,
6
C
C-F
19
29.5, 128.9, 122.6, 115.8 (q, JC-F 288.6 Hz, CF ). F NMR (376 MHz, CDCl ) δ -75.7. HRMS
3
3
F
+
(
ES): calcd. for C H ClF NONa 245.9904; found 245.9887. Spectral values were in accordance
8 5 3
with those reported in ref. 20.
N-(4-cyanophenyl)-2,2,2-trifluoroacetimidate
4
-cyanoaniline (5 g, 0.042 mol) and Et N (5.9 mL, 0.042 mol) was dissolved in dry CH Cl (20
3
2
2
mL). The mixture was cooled to 0°C and then was TFAA (5.9 mL, 0.042 mol) slowly added. The
reaction was allowed to reach rt. and stirred for 2 hours. The CH Cl was evaporated and the
2
2
mixture was dissolved in EtOAc and transferred to a separatory funnel. The organic phase was
washed thrice with 1M HCl solution, then saturated sodium bicarbonate solution and brine. The
organic phase was dried over MgSO and evaporated to dryness. The solid was recrystallized from
4
pentane/Et O giving white fluffy crystals. 6.4 g, 70%, R 0.63 (pentane/EtOAc 10:1) M 165-168
2
f
p
[
21] 1
°
C, lit. 166-167 °C . H NMR (400 MHz, CDCl ) δ 8.47 (s, 1H, NH), 7.78 (d, J 8.9 Hz, 2H),
3 H
1
3
7
1
7
.70 (d, J 8.9 Hz, 2H). C NMR (100 MHz, DMSO-d ) δ 154.9 (q, J 37.6 Hz, C=O), 140.7,
6 C C-F
1
9
33.4, 121.1, 118.6(CN) 115.5 (q, JC-F 288.7 Hz, CF ), 107.7. F NMR (376 MHz, CDCl ) δ -
3
3
F
+
5.6. HRMS (ES): calcd. for C H F N OH 215.0427; found 215.0426. Spectral values were in
9
5
3
2
accordance with those reported in ref. 22.
N-(4-nitrophenyl)-2,2,2-trifluoroacetimidate
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4
-nitroaniline (5 g, 0.036 mol) and Et N (5.1 mL, 0.036mol) was dissolved in dry CH Cl (20mL).
3
2
2
The mixture was cooled to 0°C and then was TFAA (5.0 mL, 0.036 mol) slowly added. The
reaction was allowed to reach rt. and stirred for 2 hours. The CH Cl was evaporated and the
2
2
mixture dissolved in EtOAc and transferred to a separatory funnel. The organic phase was washed
trice with 1M HCl solution, then saturated sodium bicarbonate solution and brine. The organic
phase was dried over MgSO and evaporated to dryness. The solid was recrystallized from
4
pentane/Et O giving brown-yellow fluffy crystals. 7.43 g, 88%, R 0.38 (pentane/EtOAc 5:1) M
p
2
f
[
22] 1
1
1
1
48.5-152.5 °C, lit. 151.5-152 °C . H NMR (400 MHz, CDCl ) δ 8.30 (d, J 9.2 Hz, 2H), 8.26 (s,
3 H
1
3
H), 7.82 (d, J 9.2 Hz, 2H). C NMR (100 MHz, DMSO-d ) δ 155.02 (q, J 37.8 Hz, C=O),
6
C
C-F
1
9
44.04 , 142.43 , 124.85 , 121.05 , 115.48 (q, JC-F 288.6 Hz, CF ). F NMR (376 MHz, DMSO-d )
3
6
+
δ -74.0. HRMS (ES): calcd. for C H F N O H 235.0325; found 235.0327. Spectral values were in
F
8
5
3
2
3
accordance with those reported in ref. 23.
Methyl 6-O-(2,3,4,6-tetra-O-acetyl-β-
D
-glucopyranosyl)-2,3,4-tri-O-benzyl-α-
D
-glucopyranoside
(16)
RT
D
[24]
Yield: 70 mg, 88%, R : 0.47 (pentane/EtOAc 3:2), [α]
7.2 (c 1, CHCl ), lit. 8.7 (c 1, CHCl ),
3 3
f
1
H NMR (400 MHz, CDCl ): δ 7.30 – 7.12 (m, 15H), 5.11 – 5.05 (m, 1H), 5.01 – 4.92 (m, 2H),
3
H
4
–
.88 (d, J 10.9 Hz, 1H), 4.76 (d, J 10.8 Hz, 1H), 4.72 – 4.67 (m, 2H), 4.55 (d, J 12.1 Hz, 1H), 4.49
4.40 (m, 3H), 4.13 (dd, J 12.3, 4.7 Hz, 1H), 4.02 (dd, J 12.3, 2.4 Hz, 1H), 3.96 (dd, J 10.6, 1.5
Hz, 1H), 3.87 (t, J 9.3 Hz, 1H), 3.70 – 3.64 (m, 1H), 3.62 – 3.52 (m, 2H), 3.41 (dd, J 9.6, 3.5 Hz,
1
3
1
H), 3.36 – 3.30 (m, 1H), 3.26 (s, 3H), 1.94 (s, 3H), 1.92 (s, 3H), 1.89 (s, 3H), 1.85 (s, 3H). C
NMR (100 MHz, CDCl ): 170.8, 170.4, 169.4, 169.1, 138.8, 138.2, 138.1, 128.6, 128.6, 128.5,
3
1
7
8
28.3, 128.0, 128.0, 128.0, 127.8, 127.7, 100.8, 98.1, 82.0, 79.9, 77.7, 75.8, 75.0, 73.5, 73.1, 71.9,
+
1.4, 69.7, 68.4, 68.3, 62.0, 55.3, 20.8, 20.8, 20.7, 20.7. HRMS (ES): calcd. for C H O NH
4
4
2
50 15
12.3488; found 8123502. Spectral values were in accordance with those reported in ref. 18.
Associated Content
The Supporting Information is available free of charge on the publications website: General
1
13
procedures and H and C spectra of compounds.
Author Information
*Email: hhj@chem.au.dk
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5

ACCEPTED MANUSCRIPT
The Authors declare no competing financial interest.
Acknowledgments
We are grateful for financial support from The Villum Foundation (VKR023110).
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A simpel way to remove trichloroacetamide during work-up after glycosylation with
trichloroacetimidate glycosyl donors
Derivatization of lactols and excess acceptor with commercial reagent leads to their easy removal
without using chromatographic purification