Covalent linkage of N-methyl-6-oxyquinolinium betaine to trehalose

Article abstract of DOI:10.1016/j.carres.2011.10.022

The common route to link quinolinium and pyridinium fluorophores to biomolecules via bromoacetic acid has failed in labeling the disaccharide trehalose with N-methyl-6-oxyquinolinium betaine: the unexpected, extremely high instability of the N-carboxymeth

Full text of DOI:10.1016/j.carres.2011.10.022

Carbohydrate Research 346 (2011) 2960–2964
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Carbohydrate Research
journal homepage: www.elsevier.com/locate/carres
Note
Covalent linkage of N-methyl-6-oxyquinolinium betaine to trehalose
⇑
Falko Berndt, Mohsen Sajadi, Nikolaus P. Ernsting, Rainer Mahrwald
Department of Chemistry, Humboldt-Universität zu Berlin, Brook-Taylor-Straße 2, 12489 Berlin, Germany
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 11 August 2011
Received in revised form 7 October 2011
Accepted 12 October 2011
Available online 19 October 2011
The common route to link quinolinium and pyridinium fluorophores to biomolecules via bromoacetic
acid has failed in labeling the disaccharide trehalose with N-methyl-6-oxyquinolinium betaine: the unex-
pected, extremely high instability of the N-carboxymethyl ester was overcome by direct N-alkylation of
the quinoline derivative with trehalose triflate.
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
Trehalose
N-Methyl-6-oxyquinolinium betaine
N-Alkylation
N-Methyl-6-oxyquinolinium betaine (1) (Scheme 1) is a very
promising fluorophore for investigating solvation dynamics of bio-
molecules by time-dependent Stokes-shift spectroscopy.1 The
influence of the disaccharide trehalose 2 on the terahertz-far infra-
red spectrum of water has been investigated by this method.2 Tre-
halose was chosen because of its superior biological protection
function in organisms, compared to other saccharides.3 It prevents
denaturation of proteins and fusion of membranes under stress
conditions such as heat or desiccation.4 The trehalose–water inter-
face5 is expected to be crucial for the preservation efficiency. But
only the bulk THz-FIR spectrum of aqueous trehalose solutions
could be measured in previous experiments2,6 since 1 was freely
dispersed in the mixture. However, to monitor the interface itself,
the ‘molecular spectrometer’ has to be linked covalently to the
disaccharide. Herein, we report the synthesis of N-methyl-6-oxy-
quinolinum betaine labeled trehalose, which provides a challeng-
ing example for linking quinolinium fluorophores to biomolecules.
Little is known about the synthesis of N-alkyl-6-oxy-quinolini-
um betaines.7 In order to synthesize the required conjugate 3 the
‘classical’ route for the preparation of PNAs8 was continued for
two reasons: bromoacetic acid represents a convenient alkylation
reagent for N-heterocyclic compounds, and it has already been
widely employed in the synthesis of quinoline9 and 4-methylquin-
oline10 conjugates. Esterification of suitable protected trehalose 4
with bromoacetic acid followed by N-alkylation of 6-benzyloxy-
quinoline 5 should give access to the required conjugate 3
(Scheme 2).
mono- and ditritylated trehalose. Isolation of the monotritylated
trehalose was not feasible, and hence we developed a time- and
cost-saving one-pot procedure for the tritylation and benzylation
of trehalose. Subsequent detritylation led to protected trehalose
4 (34% over three steps) (Scheme 3). Esterification of bromoacetic
acid with trehalose 4 and subsequent quaternization of 6-benzyl-
oxyquinoline led to the protected conjugate
7 (53% yield,
Scheme 3). The alternative route (alkylation of benzyl-protected
quinoline 5 by bromoacetic acid, followed by esterification with
trehalose derivative 4) yielded the required conjugate in much
lower yields (<10%).
The remaining deprotection of trehalose ester 7 to give the re-
quired conjugate 3 turned out to be difficult. The very mild and
selective hydrogenation to cleave the benzyl ether causes side
reactions. Hydrogenation with Pd/C in aprotic solvents led to par-
tial reduction of the aromatic quinolinium system, whereas in
protic media rapid saponification and formation of trehalose was
observed. Even the use of milder hydrogenation catalysts such
as Raney Ni13 or Perlman’s Pd(OH)2/C14 could also not prevent
OH
O
HO
HO
O
OH
O
OH
Following this route, benzyl-protected trehalose 411 was syn-
thesized via a tritylation–benzylation–detritylation sequence of
trehalose.12 Tritylation of trehalose resulted in a mixture of
N
O
OH
HO
OH
2
1
⇑
Corresponding author. Tel.: +49 30 2093 8397.
E-mail address: rainer.mahrwald@chemie.hu-berlin.de (R. Mahrwald).
Scheme 1. N-Methyl-6-oxyquinolinium betaine (1) and trehalose (2).
0008-6215/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2011.10.022

F. Berndt et al. / Carbohydrate Research 346 (2011) 2960–2964
2961
O
The quinolinium salt 3 could be isolated in the form of its
hydrochloride salt only. All attempts to neutralize an aqueous
acidic solution (pH <3) of 3ꢀHCl with weak bases or just by dilution
with water failed. Rapid hydrolysis of the ester bond was observed.
The fast hydrolysis of 3ꢀHCl in protic solvents limits its use for
time-resolved fluorescence spectroscopy. To overcome this ex-
treme instability, we have explored a new linker-free strategy in
a direct synthesis of the improved conjugate 8 (Scheme 4).
Our first efforts were devoted to a protecting-group-free strat-
egy. To this end, trehalose was converted into the 6-deoxy-6-iodo
compound 9.16 Direct N-alkylation of 6-hydroxyquinoline 10 was
achieved under neat conditions at high temperature. Subsequent
neutralization yielded the trehalose conjugate 8 in 8% yield
(Scheme 5).
In order to improve the N-alkylation step in the synthesis of
conjugate 8, a primary 6-OH-group of trehalose was transformed
into a stronger leaving group.17 When used with the same
protecting group methodology of Scheme 3, we have converted
benzyl-protected trehalose 4 quantitatively into monotriflate 11.
Subsequent N-alkylation of 6-benzyloxyquinoline 5 by trehalose
monotriflate 11 was accomplished in refluxing acetonitrile. The
protected trehalose conjugate 12 was isolated in good yields
(78%, Scheme 6). O-Debenzylation was achieved by the use of TiCl4
to yield improved and stable conjugate 8 (82%).
Conjugate 8 (and the protonated form) is stable under measure-
ment conditions and shows no decomposition in acidic and slightly
basic aqueous solutions (2 < pH < 9). Therefore, it is suitable for
investigating the solvation dynamics of trehalose in water by fluo-
rescence upconversion techniques. In addition, the distance be-
tween trehalose and 1 is reduced compared to conjugate 3 and
the fluorophore is better located in the hydration shell of trehalose,
which is expected to improve the spatial resolution of the solvation
dynamics.
In summary, the polarity probe N-methyl-6-oxyquinolinuim
betaine 1 was successfully attached to trehalose. The unpredicted
instability of the ester bond of 3 was overcome by developing a
more facile and linker-free approach to the improved conjugate
7. This effective method for linking quinolinium fluorophores to
biomolecules will be enhanced in the near future to allow further
solvation dynamic studies by time-dependent Stokes-shift
spectroscopy.
OH
O
HO
HO
N
O
HO
O
O
O
HO
OH
3
OH
OBn
OBn
O
BnO
BnO
N
BnO
5
Br
O
OH
OBn
O
O
BnO
OH
OBn
4
Scheme 2. Retrosynthetic analysis of 3.
reduction of 7. This observation reveals the extremely strong elec-
trophilicity of the quinolinium system.
O-Debenzylation of trehalose ester 7 was finally achieved in the
presence of strong Lewis acids. The commonly used Lewis acids
iron(III) chloride, boron trichloride, and titanium tetrachloride15
successfully cleaved all benzyl ethers; however, TiCl4 delivered
the best yields.
OH
O
HO
HO
HO
O
OH
OH
O
HO
OH
2
(i) TrCl, DIPEA, 42%
(ii) BnBr, NaH, 85%
(iii) TsOH, 95%
(iv) bromoacetic acid, DCC, DMAP, 91%
OBn
O
Br
BnO
BnO
OH
O-
O
BnO
O
O
HO
HO
O
O
OH
O
OBn
BnO
OBn
N
OH
6
OBn
O
HO
5
OH
N
8
MeCN, reflux, 8 h
OBn
OBn
OBn
O
OBn
BnO
BnO
O
BnO
BnO
N
Br-
BnO
O
BnO
O
N
O
O
OH
5
O
O
OBn
BnO
OBn
BnO
OBn
OBn
7, 53%
4
Scheme 3. Synthesis of benzyl-protected trehalose 4 and coupling to 6-benzyloxy-
quinoline (5).
Scheme 4. Retrosynthetic analysis of compound 8.

2962
F. Berndt et al. / Carbohydrate Research 346 (2011) 2960–2964
OH
OH
OH
O
O
O
HO
HO
HO
N
HO
10
HO
HO
O
O
X
N
O
O
HO
OH
HO
OH
OH
OH
8, 8%
2: X = OH
9: X = I
NIS, PPh3, 52%
Scheme 5. Protecting group free synthesis of conjugate 8.
1H), 3.16 (dd, J = 9.9, 3.8 Hz, 1H), 2.89 (dd, J = 10.0, 9.3 Hz, 1H);
13C NMR (125 MHz, D2O): d 166.4, 157.1, 147.3, 133.6, 131.8,
128.2, 122.0, 119.9, 111.5, 93.2, 93.1, 72.5, 72.2, 72.0, 70.9, 70.8,
69.6, 69.4, 69.4, 64.9, 60.6, 58.0; HRESIMS (m/z): calcd for
[C23H30O13N]+: 528.1712; found: 528.1715.
OBn
O
BnO
BnO
OBn
BnO
O
+
OR
OBn
N
O
BnO
5
1.2. 2,3,4,20,30,40,60-Hepta-O-benzyl-
a,a-D-trehalose (4)
OBn
4: R = H
11: R = Tf
Tf2O, pyridine, 95%
OBn
MeCN, reflux, 7 h
OBn
A 1000-mL round-bottomed flask was charged with 20.00 g
(58.43 mmol) -trehalose, 19.9 mL (117 mmol, 2.0 equiv) of
a,a-D
DIPEA, and 450 mL of DMF. Afterward, 16.29 g (58.43 mmol,
1.0 equiv) of trityl chloride was added slowly and the resulting yel-
low solution was stirred for three days. Subsequently, the solution
was added dropwise over one hour to a suspension of 23.80 g
(584.28 mmol, 10.0 equiv) of NaH (60%) in 40 mL of dry DMF,
and the mixture was stirred for 30 min before it was cooled to
0 °C. To this suspension 62.5 mL (526 mmol, 9.0 equiv) of benzyl
bromide was added dropwise over 90 min and the mixture was
stirred for 30 h at room temperature. After addition of 200 mL of
satd aq NH4Cl solution the reaction mixture was extracted three
times with 300 mL of DCM. The combined organic phases were
washed with 250 mL of water and dried over MgSO4. Volatiles
were removed under reduced pressure and the remaining brown
residue was dissolved in 300 mL of DCM and 100 mL of MeOH.
After addition of 5.65 g (29.7 mmol, 0.5 equiv) of p-toluenesulfonic
acid (TsOH) the solution was stirred for 16 h. Then, additional
3.00 g (15.8 mmol, 0.3 equiv) of TsOH was added and the resulting
solution was stirred for further 3 h. The reaction mixture was di-
luted with 400 mL of satd aq NaHCO3 solution and extracted twice
with 300 mL of DCM. The combined organic phases were dried
over MgSO4 and the solvent was removed under reduced pressure.
The remaining oil was purified by flash chromatography on silica
gel (gradient: petroleum ether–acetone 9:1?8:2) yielding
19.40 g (19.95 mmol, 34% yield) of the title compound 4 as an or-
O
BnO
BnO
BnO
12, 78%
O
TfO-
N
O
OBn
OBn
BnO
(i) TiCl4
(ii) pH 7.0
OH
O
O
HO
HO
HO
8, 82%
O
N
O
HO
OH
OH
Scheme 6. Synthesis of conjugate 8.
1. Experimental
1.1. 6-O-(2-(6-Hydroxyquinolinium-1yl)acetyl)-
chloride (3ꢀHCl)
a,a
-D
-trehalose
ange oil. ½a 2D5
ꢁ
+93.7 (c 1.0 in CHCl3); 1H NMR (500 MHz, DMSO-d6):
d 7.38–7.05 (m, 35H), 5.28 (d, J = 3.4 Hz, 1H), 5.23 (d, J = 3.4 Hz,
1H), 4.88 (d, J = 1.3 Hz, 1H), 4.86 (d, J = 1.4 Hz, 1H), 4.80 (ps t,
J = 5.6, 5.6 Hz, 1H), 4.77–4.67 (m, 6H), 4.66–4.58 (m, 3H), 4.54–
4.38 (m, 3H), 4.07–3.99 (m, 1H), 3.93–3.79 (m, 3H), 3.61–3.46
(m, 7H); 13C NMR (125 MHz, DMSO-d6): d 138.7, 138.7, 138.4,
138.1, 138.0, 138.0, 137.9, 128.1–126.3 (35 CH), 92.0, 91.8, 80.8,
80.8, 79.0, 78.9, 77.3, 77.1, 74.4, 74.3, 74.0, 73.8, 72.3, 71.8, 71.7,
71.5, 70.1, 68.3, 62.8; HRESIMS (m/z): calcd for [C61H64O11Na]+:
995.4341; found: 995.4340.
A 100-mL argon-purged, dried Schlenk flask was charged with
0.536 g (0.40 mmol) of 11 and 30 mL dry DCM and cooled to
0 °C. Then, 0.88 mL (8.03 mmol, 20.0 equiv) of TiCl4 was added
dropwise and the resulting suspension was stirred for 3 h at 0 °C.
The reaction was quenched by addition of 30 mL of water and
the white precipitate was filtered off. Volatile compounds were re-
moved under reduced pressure, and the residue was purified by
flash chromatography on silica gel (gradient: DCM–MeOH–H2O–
formic acid 65:35:5:1?MeOH–H2O–formic acid 50:50:1) yielding
0.130 g (0.23 mmol, 57% yield) of the title compound 3ꢀHCl as an
1.3. 6-Benzyloxyquinoline (5)
amorphous orange solid. ½a D25
ꢁ
+77.5 (c 1.0 in MeOH); 1H NMR
To a suspension of 1.67 g (41.6 mmol, 1.2 equiv) of NaH (60%) in
10 mL of dry DMF at 0 °C was added dropwise a solution of 5.00 g
(34.6 mmol) of 6-hydroxyquinoline in 30 mL of dry DMF. The mix-
ture was allowed to warm to room temperature and stir for 30 min
before 4.95 mL (41.6 mmol, 1.2 equiv) of benzyl bromide was
added slowly. The reaction mixture was stirred for 16 h and then
(500 MHz, D2O): d 8.97 (dd, J = 5.8, 0.9 Hz, 1H), 8.94 (d, J = 8.5 Hz,
1H), 8.09 (d, J = 9.6 Hz, 1H), 7.92 (dd, J = 8.5, 5.8 Hz, 1H), 7.74 (dd,
J = 9.6, 2.8 Hz, 1H), 7.57 (d, J = 2.8 Hz, 1H), 5.90 (s, 2H), 4.86 (d,
J = 3.8 Hz, 1H), 4.74 (d, J = 3.8 Hz, 1H), 4.45 (dd, J = 12.2, 2.2 Hz,
1H), 4.26 (dd, J = 12.3, 4.6 Hz, 1H), 3.80–3.72 (m, 2H), 3.70–3.58
(m, 4H), 3.41 (dd, J = 9.9, 3.8 Hz, 1H), 3.31 (ps t, J = 9.4, 9.4 Hz,

F. Berndt et al. / Carbohydrate Research 346 (2011) 2960–2964
2963
the reaction was quenched by adding 100 mL of satd aq NH4Cl
solution. After extracting three times with 50 mL of DCM, the com-
bined organic phases were washed with 50 mL of water and dried
over MgSO4. Volatiles were removed under reduced pressure and
the remaining crude product was further purified by flash chroma-
tography on silica gel (petroleum ether–ethyl acetate 2:1) yielding
5.93 g (25.2 mmol, 73% yield) of the title compound 5 as a pale-yel-
low solid. 1H NMR (500 MHz, DMSO-d6): d 8.74 (dd, J = 4.2, 1.7 Hz,
1H), 8.24 (dd, J = 8.3, 1.1 Hz, 1H), 7.94 (m, 1H), 7.53 (m, 1H), 7.51
(m, 1H), 7.50–7.39 (m, 5H), 7.35 (m, 1H), 5.24 (s, 2H); 13C NMR
(125 MHz, DMSO-d6): d 156.1, 147.9, 143.7, 136.5, 134.6, 130.3,
128.8, 128.3, 127.8, 127.7, 122.1, 121.6, 106.9, 69.5; HRESIMS (m/
z): calcd for [C16H14ON]+: 236.1070, found: 236.1070.
74.3, 74.0, 73.9, 72.3, 71.8, 70.3, 70.2, 68.2, 68.0, 63.6, 57.4; HRE-
SIMS (m/z): calcd for [C79H78O13N]+: 1248.5468; found: 1248.5464.
1.6. 6-Deoxy-6-(6-oxyquinolinium-1yl)-a,a-D-trehalose (8)
1.6.1. Method A. Deprotection of 6-(6-Benzyloxyquinolinium-
1yl)-6-deoxy-2,3,4,20,30,40,60-hepta-O-benzyl-
a,a-D-trehalose
trifluoromethanesulfonate
A 250-mL argon-purged, dried Schlenk flask was charged with
1.50 g (1.18 mmol) of 12 and 80 mL of dry DCM and was cooled
to 0 °C. The dropwise addition of 3.10 mL (28.3 mmol, 24.0 equiv)
TiCl4 resulted in a suspension, which was stirred for 3 h at 0 °C.
The reaction was quenched by addition of 100 mL of water and
the white precipitate was filtered off. Volatile compounds were re-
moved under reduced pressure and the residue was purified by
flash chromatography on silica gel (gradient: DCM–MeOH–H2O
65:35:5?MeOH–H2O 3:2). The product was dissolved in 10 mL
of MeOH, and the solution was neutralized with Amberlite IRA-
410 (OHꢂ-form) and filtered. The solvent was removed under re-
duced pressure yielding 0.46 g (0.969 mmol, 82% yield) of the title
compound 8 as an amorphous orange solid.
1.4. 2,3,4,20,30,40,60-Hepta-O-benzyl-6-O-(2-bromoacetyl)-
a,a-D-
trehalose (6)
In a round-bottomed flask 7.41 g (7.61 mmol) of 4, 1.89 g
(9.15 mmol, 1.2 equiv) of DCC, 1.27 g (9.15 mmol, 1.2 equiv) of
bromoacetic acid, and 0.093 g (0.76 mmol, 0.1 equiv) of 4-DMAP
were dissolved in 120 mL of dry DCM and stirred for 16 h. An addi-
tional 0.157 g (0.76 mmol, 0.1 equiv) of DCC and 0.107 g
(0.77 mmol, 0.1 equiv) of bromoacetic acid were added to the solu-
tion. After 5 h the precipitate was filtered off and the remaining fil-
trate was concentrated under reduced pressure. The crude product
was purified by flash chromatography on silica gel (petroleum
ether–acetone 7:3) yielding 7.54 g (6.89 mmol, 91% yield) of the ti-
1.6.2. Method B. N-Alkylation of 6-deoxy-6-iodotrehalose
A round-bottomed flask containing 0.690 g (1.53 mmol) of 6-
deoxy-6-iodotrehalose (9) and 0.246 g (1.68 mmol, 1.10 equiv) of
6-hydroxyquinoline (10) was heated for 30 min at 190 °C. The mix-
ture was allowed to reach to room temperature and was taken up
in 5 mL of MeOH. After neutralization with Amberlite IRA-410
(OHꢂ-form) and filtration, the solvent was removed under reduced
pressure. Flash chromatography on silica yielded 0.069 g
tle compound 6. ½a D25
ꢁ
+95.3 (c 1.0 in CHCl3); 1H NMR (500 MHz,
DMSO-d6): d 7.40–7.12 (m, 35H), 5.29 (d, J = 3.4 Hz, 1H), 5.27 (d,
J = 3.4 Hz, 1H), 4.90 (d, J = 11.5 Hz, 1H), 4.88 (d, J = 11.9 Hz, 1H),
4.79–4.62 (m, 8H), 4.52 (d, J = 12.0 Hz, 1H), 4.50 (d, J = 12.9 Hz,
1H), 4.48 (d, J = 10.9 Hz, 1H), 4.43 (d, J = 12.0 Hz, 1H), 4.20–4.15
(m, 2H), 4.13–4.06 (m, 2H), 4.12–4.07 (m, 1H), 4.01 (ddd, J = 9.5,
3.4, 3.1 Hz, 1H), 3.88 (ps t, J = 9.3, 9.3 Hz, 1H), 3.86 (ps t, J = 9.4,
9.4 Hz, 1H), 3.60–3.51 (m, 6H); 13C NMR (125 MHz, DMSO-d6): d
166.9, 138.7, 138.5, 138.1, 137.9, 137.9, 137.8, 137.7, 128.4–127.0
(35CH), 92.1, 91.9, 80.8, 80.7, 78.9, 78.8, 77.3, 76.8, 74.5, 74.4,
74.0, 72.4, 71.8, 71.7, 70.3, 68.3, 68.3, 63.7, 26.9; HRESIMS (m/z):
calcd for [C63H65O12BrNa]+: 1115.3552, found: 1115.3556.
(0.15 mmol, 8%) of 8. ½a D25
ꢁ
+112.0 (c 0.5 in MeOH); 1H NMR
(500 MHz, D2O): d 8.95 (d, J = 5.5 Hz, 1H), 8.83 (d, J = 8.4 Hz, 1H),
8.21 (d, J = 9.6 Hz, 1H), 7.86 (dd, J = 8.2, 5.9 Hz, 1H), 7.68 (dd,
J = 9.6, 2.4 Hz, 1H), 7.46 (d, J = 2.4 Hz, 1H), 5.50 (d, J = 14.1 Hz,
1H), 4.99 (d, J = 3.7 Hz, 1H), 4.89 (dd, J = 14.5, 9.2 Hz, 1H), 4.14
(ps t, J = 9.2, 9.2 Hz, 1H), 4.05 (d, J = 3.6 Hz, 1H), 3.75 (ps t, J = 9.4,
9.4 Hz, 1H), 3.70–3.54 (m, 4H), 3.52 (ps t, J = 9.6, 9.6 Hz, 2H), 3.16
(ps t, J = 9.4, 9.4 Hz, 1H), 2.95 (dd, J = 9.9, 3.7 Hz, 1H); 13C NMR
(125 MHz, D2O): d 156.7, 146.8, 146.3, 133.0, 132.0, 127.6, 121.5,
120.2, 111.5, 93.6, 93.5, 72.6, 72.5, 72.1, 71.4, 70.7, 70.4, 70.0,
69.4, 60.4, 58.0; HRESIMS (m/z): calcd for [C21H28O11N]+:
470.1657; found: 470.1658.
1.5. 2,3,4,20,30,40,60-Hepta-O-benzyl-6-O-(2-(6-
benzyloxyquinolinium-1yl)acetyl)-a,a-D-trehalose bromide (7)
After dissolving 7.50 g (6.86 mmol) of 6-O-(2-bromoacetyl)-
2,3,4,20,30,40,60-hepta-O-benzyl-
-trehalose (6), and 2.42 g
a
,a-
D
1.7. 2,3,4,20,30,40,60-Hepta-O-benzyl-6-O-
(10.28 mmol, 1.5 equiv) of 6-benzyloxyquinoline (5) in 50 mL of
CH3CN, the resulting solution was stirred under reflux for 8 h.
Volatiles were removed under reduced pressure, and the crude
product was purified by flash chromatography on silica gel (DCM–
MeOH 9:1) yielding 4.82 g (3.63 mmol, 53% yield) of the title com-
trifluoromethanesulfonyl-a,a-D-trehalose (11)
A 50-mL argon-purged, dried Schlenk flask was charged with
1.47 g (1.51 mmol) of 4 and 6 mL of dry DCM before 0.15 mL
(1.81 mmol, 1.2 equiv) of dry pyridine was added. At ꢂ30 °C
0.28 mL (1.66 mmol, 1.1 equiv) of trifluoromethanesulfonic anhy-
dride was added dropwise and the suspension was stirred for 1 h
at 0 °C. The reaction was quenched by addition of 20 mL of DCM
and 10 mL of water. After phase separation the aq phase was ex-
tracted twice with 10 mL DCM. The combined organic phases were
washed with 10 mL of 10% H2SO4 and dried (MgSO4). The solvent
was removed under reduced pressure yielding 1.59 g (1.44 mmol,
95% yield) of the title compound 11 as a colorless oil, which was
pound 7. ½a 2D5
ꢁ
+83.6 (c 1.0 in CHCl3); 1H NMR (500 MHz, DMSO-d6):
d 9.36 (dd, J = 6.0, 1.0 Hz, 1H), 9.24 (d, J = 8.4 Hz, 1H), 8.37 (d,
J = 9.7 Hz, 1H), 8.22 (dd, J = 8.4, 5.8 Hz, 1H), 8.05 (d, J = 2.8 Hz,
1H), 7.94 (dd, J = 9.6, 2.8 Hz, 1H), 7.52–7.09 (m, 40H), 6.29 (d,
J = 17.7 Hz, 1H), 6.15 (d, J = 17.7 Hz, 1H), 5.28 (d, J = 11.7 Hz, 1H),
5.24 (d, J = 11.7 Hz, 1H), 5.19 (d, J = 3.9 Hz, 1H), 5.18 (d, J = 3.9 Hz,
1H), 4.86 (d, J = 11.2 Hz, 1H), 4.84 (d, J = 11.2 Hz, 1H), 4.74 (d,
J = 11.3 Hz, 1H), 4.71 (d, J = 11.3 Hz, 1H), 4.71–4.67 (m, 2H), 4.64
(s, 2H), 4.63 (d, J = 12.6 Hz, 1H), 4.55 (d, J = 11.0 Hz, 1H), 4.50–
4.39 (m, 3H), 4.20 (d, J = 11.1 Hz, 1H), 4.18 (dd, J = 13.0, 2.2 Hz,
1H), 4.09 (dd, J = 12.0, 3.4 Hz, 1H), 4.02–3.95 (m, 2H), 3.85–3.75
(m, 2H), 3.58–3.51 (m, 2H), 3.51–3.48 (m, 2H), 3.33–3.32 (m,
1H), 3.10 (ps t, J = 9.5, 9.5 Hz, 1H); 13C NMR (125 MHz, DMSO-
d6): d 165.4, 158.2, 148.2, 146.9, 138.5, 138.5, 138.0, 138.0, 137.9,
137.9, 137.8, 135.4, 133.9, 131.3, 128.4–127.2 (40CH), 122.5,
120.5, 109.1, 92.2, 92.0, 80.7, 80.5, 78.8, 78.8, 77.2, 76.5, 74.4,
used in the next step without further purification. ½a D25
ꢁ
+107.6 (c
1.0 in CHCl3); 1H NMR (500 MHz, CDCl3): d 7.57–7.03 (m, 35H),
5.25 (d, J = 3.6 Hz, 1H), 5.23 (d, J = 3.6 Hz, 1H), 5.08 (d, J = 10.9 Hz,
1H), 5.05 (d, J = 11.0 Hz, 1H), 4.96 (d, J = 10.8 Hz, 1H), 4.96 (d,
J = 10.9 Hz, 1H), 4.90 (d, J = 10.9 Hz, 1H), 4.87 (d, J = 10.8 Hz, 1H),
4.84 (d, J = 12.0 Hz, 1H), 4.72 (s, 2H), 4.65 (d, J = 11.9 Hz, 1H),
4.60 (d, J = 11.9 Hz, 1H), 4.58 (d, J = 10.6 Hz, 1H), 4.53 (d,
J = 10.7 Hz, 1H), 4.45 (d, J = 12.1 Hz, 1H), 4.28–4.21 (m, 2H),

2964
F. Berndt et al. / Carbohydrate Research 346 (2011) 2960–2964
4.21–4.16 (m, 1H), 4.12 (d, J = 9.4 Hz, 1H), 4.07 (ps t, J = 9.4, 9.4 Hz,
1H), 4.07 (ps t, J = 9.2, 9.2 Hz, 1H), 3.78–3.72 (m, 1H), 3.67 (dd,
J = 9.7, 3.6 Hz, 1H), 3.62–3.57 (m, 2H), 3.52 (ps t, J = 9.4, 9.4 Hz,
1H), 3.44 (dd, J = 10.7, 1.8 Hz, 1H); 13C NMR (125 MHz, CDCl3): d
138.6, 138.4, 138.1, 138.0, 137.8, 137.7, 137.5, 128.5–127.2
(35CH), 122.2, 119.7, 117.1, 114.2, 94.7, 94.1, 81.8, 81.4, 79.4,
79.2, 77.7, 76.3, 75.5, 75.5, 75.1, 75.1, 74.5, 73.5, 73.2, 72.6,
70.8, 68.3, 68.0; HRESIMS (m/z): calcd for [C62H67O13NF3S]+:
1122.4280, found: 1122.4279.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.carres.2011.10.022.
References
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This work was financed by the Deutsche Forschungsgemeins-
chaft (ER 154/9). We thank Daniel Wünsche for practical support.