Hexafluoroacetone as protecting and activating reagent. A new approach to N-linked glycopeptoids

Article abstract of DOI:10.1007/s00706-003-0147-5

A preparatively simple synthesis of N-linked glycopeptoids starting from iminodiacetic acid and glycosylamines, using hexafluoroacetone as protecting and activating reagent is described. Springer-Verlag 2004.

Full text of DOI:10.1007/s00706-003-0147-5

Monatshefte f€ur Chemie 135, 865–877 (2004)
DOI 10.1007/s00706-003-0147-5
Hexafluoroacetone as Protecting
and Activating Reagent. A New
Approach to N-linked Glycopeptoids
1
1
Klaus Burger1;ꢀ, Christoph Bottcher , Lothar Hennig ,
¨
and Samy A. Essawy2
1
Department of Organic Chemistry, University of Leipzig, D-04103 Leipzig, Germany
2
Chemistry Department, Faculty of Science, Benha University, Benha, Egypt
Received October 17, 2003; accepted November 24, 2003
Published online February 27, 2004 # Springer-Verlag 2004
Summary. A preparatively simple synthesis of N-linked glycopeptoids starting from iminodiacetic
acid and glycosylamines, using hexafluoroacetone as protecting and activating reagent is described.
Keywords. Iminodiacetic acid; Hexafluoroacetone; Dielectrophile; Glycosylamines; N-Glycosyla-
tion; Peptoids; Glycopeptoids.
Introduction
Peptide drugs are a fastly growing class of therapeutics [1]. However, proteolytic in-
stability, low lipophilicity, and lack of transport systems are the major drawbacks,
restricting their application considerably. In general, cell membranes resist passage
of most peptides. Consequently, peptides are rapidly degradated and excreted via
liver and kidney. The high conformational flexibility of peptides creates another
problem. Bioactive peptides often bind to different receptor sites causing undesired
side effects [2, 3]. Several strategies have been developed to overcome these prob-
lems, including the application of peptidomimetics [4, 5] and glycopeptides [6, 7]
as drug candidates.
The construction of peptoids [8] (Scheme 1), a new class of non-natural pep-
tides, added a new facette to peptide modification [9]. Peptoids are N-alkyl- and
N-aryl-substituted oligoglycines, having no chiral centers [10]. Other characteris-
tics are high conformational flexibility, high structural diversity, and improved
proteolytic stability. Comparison of the peptide chain with the peptoid chain
(Scheme 1) shows that the direction of the peptide bond has to be reversed
(retro-sequence) in order to provide the same relative arrangement of the side-chain
ꢀ
Corresponding author. E-mail: burger@organik.chemie.uni-leipzig.de

866
K. Burger et al.
Scheme 1
residues and carbonyl groups. Helical peptoid structures are favored when bulky
N-alkyl side-chains are present. Interestingly, these helices are not stabilized by
hydrogen bonds [1a, 11]. Noteworthy, the peptoid approach is amenable to solid-
phase synthesis [8a, 12] which provides libraries of high structural diversity.
Roy et al. were the first to combine the advantages of peptoid and glycoconju-
gate chemistry. They synthesized a series of N-linked [13] and O-linked glycopep-
toids [14] and used them as building blocks for the construction of HIV-1 protease
inhibitors [15] and TN-antigen glycopeptidomimetic clusters [16]. Recently, Kessler
et al. reported on a stereoselective synthesis of C-glycosylated peptoids [17].
The first example of this new class of ‘‘non-natural biooligomers’’, an aspar-
agine linked N-acetyl glucosaminide mimic, has been synthesized by Roy et al.
[13a] via a N-glycosylated tripeptoid. The latter was obtained in six steps starting
from tert-butyl bromoacetate and N-acetyl glucosaminide. The submonomer solid-
phase synthesis does not apply N-substituted glycine monomers. Instead, bromo-
acetyl building blocks are coupled to the growing peptoid chain and then subjected
to a nucleophilic displacement reaction of the bromide by a primary amine carry-
ing the side-chain substituent. Protection of the amino group is not necessary.
In contrast, in the Fmoc=OtBu strategy suitably protected N-substituted glycine
derivatives are coupled to polymer-bound NHR groups using PyBOP or PyBrOP
[8b].
Herein we describe a new, preparatively simple access to N-linked glycopep-
toids using hexafluoroacetone as protecting and activating reagent.
Results and Discussion
On application of hexafluoroacetone as protecting and activating agent, N-glyco-
sylated tripeptoids can be obtained in five steps with an overall yield of 12–23%,
starting from iminodiacetic acid (1, Scheme 2).
First, 1 is transformed into lactone 2 on reaction with hexafluoroacetone in
dimethylsulfoxide (DMSO) or dimethylformamide (DMF) at room temperature
[18]. This process includes simultaneous functionalization of the imino and one of
the carboxy groups. The second carboxy group remains unaffected. The lactone
moiety represents an activated ester. In a second step the exocyclic carboxy group
can be transformed into an acid chloride. Noteworthy, treatment of 2 with

Hexafluoroacetone as Protecting and Activating Reagent
867

868
K. Burger et al.
Fig. 1. 19F NMR spectra (CDCl3, 282 MHz) of compound 8d at 300 K
phosphorus pentachloride gives better yields than treatment with thionyl chloride. 3
represents a dielectrophile. Consequently, a sequence of two regioselective acylation
steps can be performed. On treatment with relatively weak nucleophiles (1 equiv.)
like glycosylamines in the presence of a tert-base, the more reactive acid chloride
moiety reacts regioselectively to give the N-glycosylated, ꢀ-carboxy activated gly-
cine derivative 4. NMR and TLC of the crude products indicate clearly that only the
ꢁ-anomers are formed. The ꢁ-anomers can be identified by a characteristic coupling
constant 3J1;2 ¼ 9.3 ꢁ 0.3 Hz. We applied compounds 4 as acyl transfer reagents for
amino acid esters and amides, providing dipeptoids and dipeptides in good yields,
respectively. Yields and reaction rate depend on the sterical demand of the nucleo-
philes (reaction rate: glycine (70–90%)>ꢀ-amino acid (68–76%)>N-alkylated gly-
cine derivatives (45–50%)). In the case of hexafluoroacetone protected amino acids
the cleavage of the lactone moiety is always coupled with the deprotection of the ꢀ-
amino group (4 ! 5). Therefore, elongation of the peptoid chain can be continued in
N-terminal position without the need of an extra deprotection step to give the N-
glycosylated tripeptoids 6 and peptide=peptoid hybrids 8, respectively. Compounds
1
5–8 show complex H NMR spectra due to the presence of mixtures of rotamers,
which can be observed in the case of secondary amides. In the trifluoroacetyl
protected N-glycosylated tripeptoid 8d three stereogenic centers are present giving
rise to eight rotational isomers, which indeed can be found in the 19F NMR spectrum
(Fig. 1). The structures of the newly synthesized compounds are unequivocally
confirmed by NMR and mass spectrometry.
The readily available N-glycosylated peptoid fragments are valuable building
blocks for the synthesis of glycopeptoids and glycopeptide mimetics in solution
and on solid phase. Peptoids represent a new class of lead structures for the therapy
of pathological events.
Experimental
Solvents were purified and dried prior to use. Reagents were used as purchased. TLC was performed
on Silica Gel 60 F254 (Merck) with detection by UV light or phosphomolybdic acid=ceric sulphate in
5% aqueous sulfuric acid followed by heating. Flash chromatography was performed using silica gel
(32–63mm) with solvent systems given in the text. Melting points (uncorrected) were determined on a
Boetius heating table. Optical rotation indices were measured using a Schmidt & Haensch Polartronic-
1
D polarimeter in a 5 cm cell. H (200 MHz, 300MHz, 400MHz, 600MHz), 13C (50 MHz, 75 MHz,
100 MHz, 150 MHz), and 19F (188 MHz, 282MHz) NMR spectra were recorded on Varian Gemini
200, Varian Gemini 300, Bruker DRX 400, and Bruker DRX 600 spectrometers. TMS was used as

Hexafluoroacetone as Protecting and Activating Reagent
869
1
reference for H and 13C NMR spectra (internal), and CF3COOH for 19F NMR spectra (external). IR
spectra were obtained with a FTIR spectrometer (Genesis ATI Mattson=Unicam). Mass spectra were
recorded on a Bruker Daltonics APEX II ESI-FT-ICR spectrometer or on a Finnigan ZAB-HSQ
spectrometer (FAB-matrix: 3-NBA).
Synthesis of N-Glycosylated Peptoids
Protocol 1: A solution of the acid chloride 3 (1 equiv.) in dichloromethane (DCM) (5 cm3 per 1 mmol)
was cooled to 0ꢂC. Over a period of 30min a freshly prepared solution of the corresponding
O-peracetylated ꢁ-D-glycosylamine (1 equiv.) and N-ethylmorpholine (NEM) (1 equiv.) in DCM
(5cm3 per 1 mmol) were added. After vigorous stirring for 1 h at 0ꢂC, DCM (20 cm3 per 1 mmol)
was added. The organic phase was extracted with diluted citric acid (10%) and sat. NaCl solution
(10 cm3 per 1 mmol). After drying with MgSO4, the organic solvent was evaporated and the residue
was purified by flash chromatography.
Protocol 2: A suspension of the corresponding amino acid hydrochloride (1.2 equiv.) in ethyl
acetate: 2-propanol (1:1, 5 cm3 per 1 mmol) was stirred with NEM (1.2 equiv.) for 15min. Then the
corresponding HFO (2,2-bis(trifluoromethyl)-1,3-oxazolidin-5-one, 1 equiv.) was added. The mixture
was stirred at room temperature until TLC analysis showed complete consumption of the HFO (1–4d).
After evaporation of the solvent, the residue was purified by flash chromatography.
Protocol 3: A solution of a N-terminal protected glycine derivative (1.2 equiv.) in DMF (5 cm3 per
1 mmol) was stirred for 3 min after addition of diisopropylethylamine (DIEA) (2.4 equiv.) and benzo-
triazole-1-yloxytrispyrrolidinophosphonium hexafluorophophate (PyBOP) (1.2 equiv.). Then the gly-
cosylated dipeptide ester (1.0 equiv.) was added and the reaction mixture was stirred for 18h at room
temperature. The solvent was evaporated in vacuo and the residue was redissolved in ethyl acetate
(30 cm3). The organic phase was extracted with diluted citric acid (10%, 10cm3), sat. NaHCO3
(10 cm3), and sat. NaCl solution (10 cm3), and finally dried with MgSO4. After removal of the organic
solvent in vacuo, the crude product was purified by flash chromatography.
N-(2,3,4,6-Tetra-O-acetyl-ꢁ-D-glucopyranosyl)-2-[5-oxo-2,2-bis(trifluoromethyl)-1,3-
oxazolidin-3-yl]acetamide (4a, C21H24F6N2O12)
Compound 3 (3.18 g, 10.6mmol) and Ac4-ꢁ-D-Glc-NH2 (3.68 g, 10.6mmol) were reacted due to
protocol 1. Purification by flash chromatography (ethyl acetate:petroleum ether¼ 1:1, Rf ¼ 0.32). Yield
5.32g (82%), colorless foam; [ꢀ]D ¼ þ21ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.3, DCM); 1H NMR (CDCl3, 300 MHz,
COSY): ꢂ ¼ 2.02, 2.03, 2.04, 2.07 (4s, 4OAc), 3.66 (d, 1H, J ¼ 16.8Hz, CH2Nasn), 3.73 (d, 1H,
J ¼ 16.8Hz, CH2Nasn), 3.76 (s, CH2Nasn), 3.81 (m, 5-H), 4.09 (dd, J ¼ 2.1, 12.6Hz, 6-Ha), 4.28
(dd, J ¼ 4.5, 12.6Hz, 6-Hb), 4.93 (dd, J ¼ 9.6, 9.6 Hz, 2-H), 5.07 (dd, J ¼ 9.3, 9.9 Hz, 4-H), 5.22
(dd, J ¼ 9.3, 9.3 Hz, 1-H), 5.32 (dd, J ¼ 9.6, 9.6 Hz, 3-H), 7.07 (d, J ¼ 9.0 Hz, 1-NH) ppm; 19F
NMR (CDCl3, 282 MHz): ꢂ ¼ 1.54 (m, 3F), 1.63 (m, 3F) ppm; 13C NMR (CDCl3, 50 MHz, HETCOR):
ꢂ ¼ 20.92, 21.02, 21.71 (4OAc), 49.88, 51.69 (2CH2Nasn), 62.31 (6-CH2), 68.80 (4-CH), 71.16 (2-CH),
73.15 (3-CH), 74.37 (5-CH), 78.72 (1-CH) 90.36 (sept, J ¼ 33Hz), 121.57 (q, J ¼ 291 Hz), 166.50,
168.08, 170.32, 170.58, 171.34, 171.52 ppm; IR (KBr): ꢃꢀ¼ 1849, 1753, 1711, 1522cmꢃ1; MS (FAB):
m=z ¼ 611.1 [M þ H]þ.
tert-Butyl N-{2-Oxo-2-[(2,3,4,6-tetra-O-acetyl-ꢁ-D-glucopyranosyl)amino]ethyl}glycyl-
glycinate (H-Nasn(Ac4-ꢁ-D-Glc)-Gly-OtBu, 5a=1, C24H37N3O13)
t
ꢀ
HCl H-Gly-O Bu (148mg, 0.88 mmol) and 4a (450 mg, 0.74mmol) were reacted due to protocol 2.
Reaction time: 18h. Purification by flash chromatography (DCM:MeOH¼ 10:1, Rf ¼ 0.40). Yield
300 mg (70%), colorless foam; [ꢀ]D ¼ þ7ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.3, DCM); 1H NMR (CDCl3,

870
K. Burger et al.
600 MHz, COSY): ꢂ ¼ 1.49 (s, 9H, CH3tBu), 2.02, 2.04, 2.05, 2.08 (4s, 4OAc), 3.27–3.38 (m,
2CH2Nasn), 3.83 (m, 5-H), 3.97 (dd, 1H, J ¼ 5.7, 18.0Hz, CH2Gly), 4.02 (dd, 1H, J ¼ 5.7, 18.0Hz,
CH2Gly), 4.10 (dd, J ¼ 1.8, 12.3Hz, 6-Ha), 4.30 (dd, J ¼ 4.5, 12.3Hz, 6-Hb), 4.98 (dd, J ¼ 9.6, 9.6 Hz,
2-H), 5.07 (dd, J ¼ 9.9, 9.6 Hz, 4-H), 5.25 (dd, J ¼ 9.3, 9.6 Hz, 1-H), 5.32 (dd, J ¼ 9.6, 9.6 Hz, 3-H),
7.12 (br.t, J ¼ 5.7 Hz, NHGly), 7.64 (d, J ¼ 9.3 Hz, 1-NH) ppm; 13C NMR (CDCl3, 50 MHz, HETCOR):
ꢂ ¼ 20.65 (2OAc), 20.78 (2OAc), 28.21 (CH3tBu), 41.47 (CH2Gly), 52.43, 52.57 (2CH2Nasn), 61.85
(6-CH2), 68.36 (4-CH), 71.02 (2-CH), 72.90 (3-CH), 73.81 (5-CH), 78.11 (1-CH), 82.54 (CtBu),
169.42, 169.68, 169.95, 170.76, 170.96, 171.24, 171.69 ppm; IR (KBr): ꢃꢀ¼ 3500–3300, 1750,
1662, 1534 cmꢃ1; HRMS (ESI): calcd. for C24H38N3O13 (Mþ Hþ) 576.23991, found 576.23939.
tert-Butyl N-{2-Oxo-2-[(2,3,4,6-tetra-O-acetyl-ꢁ-D-glucopyranosyl)amino]ethyl}glycyl-
phenylalaninate (H-Nasn(Ac4-ꢁ-D-Glc)-Phe-OtBu, 7a, C31H43N3O13)
t
ꢀ
HCl H-Phe-O Bu (410 mg, 1.59mmol) and 4a (809mg, 1.33 mmol) were reacted due to protocol 2.
Reaction time: 2d. Purification by flash chromatography (ethyl acetate, Rf ¼ 0.25). Yield 613mg
1
(69%), colorless foam; [ꢀ]D ¼ þ24ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.1, DCM); H NMR (CDCl3, 200MHz):
Phe
ꢂ ¼ 1.42 (s, 9H, CH3tBu), 2.00 (s, OAc), 2.02 (s, 2OAc), 2.05 (s, OAc), 3.00–3.27 (m, CH2
,
2CH2Nasn), 3.83 (ddd, J ¼ 2.2, 4.4, 9.9 Hz, 5-H), 4.08 (dd, J ¼ 2.1, 12.3Hz, 6-Ha), 4.30 (dd, J ¼ 4.5,
12.3Hz, 6-Hb), 4.78 (m, CHPhe), 4.92 (dd, J ¼ 9.6, 9.6Hz, 2-H), 5.07 (dd, J ¼ 9.6, 9.9Hz, 4-H), 5.25
(dd, J ¼ 9.6, 9.3 Hz, 1-H), 5.31 (dd, J ¼ 9.6, 9.3 Hz, 3-H), 6.96 (d, J ¼ 8.0 Hz, NHPhe), 7.14–7.32 (m,
5H, arom), 7.53 (d, J ¼ 9.6 Hz, 1-NH) ppm; 13C NMR (CDCl3, 50MHz): ꢂ ¼ 20.67, 20.75, 20.81
(4OAc), 28.09 (CH3tBu), 38.05 (CH2Phe), 52.36, 52.49 (2CH2Nasn), 53.43 (CHPhe), 61.80 (6-CH2),
68.27, 70.84, 72.88, 73.75 (2,3,4,5-CH), 78.00 (1-CH), 82.51 (CtBu), 127.10, 128.57, 129.51, 136.46
(arom), 169.66, 169.95, 170.07, 170.65, 170.93, 170.99, 171.68 ppm; IR (KBr): ꢃꢀ¼ 3500–3300, 1749,
1668, 1521 cmꢃ1; HRMS (ESI): calcd. for C31H44N3O13 (Mþ Hþ) 666.28687, found 666.28675.
tert-Butyl N-{2-Oxo-2-[(2,3,4,6-tetra-O-acetyl-ꢁ-D-glucopyranosyl)amino]ethyl}glycyl-
N-benzylglycinate (H-Nasn(Ac4-ꢁ-D-Glc)-Nphe-OtBu, 5a=2, C31H43N3O13)
H-Nphe-OtBu (240 mg, 1.09 mmol) and 4a (555 mg, 0.91 mmol) were reacted in ethyl acetate (5cm3)
due to protocol 2. Reaction time: 4d. Purification by flash chromatography (gradient elution: ethyl
acetate:petroleum ether¼ 2:1 ! 8:1, Rf ¼ 0.35). Yield 300 mg (50%), foam; [ꢀ]D ¼ þ2ꢂ cm3 gꢃ1 dmꢃ1
(c ¼ 1.1, DCM); 1H NMR (CDCl3, 400 MHz, 300K, COSY): rotational isomers, ratio: 3:2, ꢂ ¼ 1.42 (s,
tBu
tBu
CH3 min), 1.47 (s, CH3 maj), 1.91–2.08 (several s, OAc), 3.27–3.63 (CH2Nasn), 3.74 (d, J ¼ 18.4Hz,
Nphe
CH2
Nphe
Nphe
min),3.82(d, J ¼ 18.4Hz, CH2
min),3.83(m, 5-H),3.96(d,J ¼ 17.2Hz, CH2
maj),4.02(d,
Nphe
J ¼ 17.2Hz, CH2
maj), 4.06–4.11 (m, 6-Ha), 4.25–4.31 (m, 6-Hb), 4.51 (d, J ¼ 18.2 Hz, CH2Ph maj),
4.55 (d, J ¼ 18.2Hz, CH2Ph maj), 4.62 (d, J ¼ 14.8Hz, CH2Ph min), 4.68 (d, J ¼ 14.8Hz, CH2Ph min),
4.94–5.11 (m, 2,4-H), 5.23–5.34 (m, 1,3-H), 7.18–7.39 (m, H arom), 8.06 (d, J ¼ 9.9 Hz, 1-NH maj), 8.10
(d, J ¼ 9.9 Hz, 1-NH min) ppm;13C NMR (CDCl3, 100 MHz, 300K, HMQC):ꢂ ¼ 20.5–20.8 (OAc), 27.96
tBu
tBu
Nphe
Nphe
(CH3 min), 28.07 (CH3 maj), 47.97 (CH2
maj), 48.37 (CH2
min), 50.33 (CH2Nasn), 50.43
(CH2Ph min), 50.94 (CH2Ph maj), 52.64 (CH2Nasn), 61.74 (6-CH2), 68.19 (4-CH), 70.77 (2-CH), 72.93
(3-CH), 73.69 (5-CH), 77.80 (1-CH), 82.09 (CtBu maj), 82.94 (CtBu min), 126.72, 127.84, 128.10, 128.61,
128.76, 129.15, 135.42, 136.14, 167.66, 168.05, 169.55, 169.61, 170.30, 170.63, 171.02, 171.27,
172.15 ppm; IR (KBr): ꢃꢀ¼ 3500–3300, 1747, 1657cmꢃ1; HRMS (ESI): calcd. for C31H44N3O13
(Mþ Hþ) 666.28687, found 666.28664.
tert-Butyl N-Benzoylglycyl-N-{2-oxo-2-[(2,3,4,6-tetra-O-acetyl-ꢁ-D-glucopyranosyl)-
amino]ethyl}glycylglycinate (Bz-Gly-Nasn(Ac4-ꢁ-D-Glc)-Gly-OtBu, 6a, C33H44N4O15)
Bz-Gly-OH (65 mg, 0.36mmol) and 5a=1 (173mg, 0.30 mmol) were reacted due to protocol 3. Puri-
fication by flash chromatography (CHCl3:MeOH¼ 15:1, Rf ¼ 0.27). Yield 127 mg (57%), colorless

Hexafluoroacetone as Protecting and Activating Reagent
871
1
crystals; mp 102–103ꢂC; [ꢀ]D ¼ ꢃ36ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.2, DCM); H NMR (CDCl3, 600 MHz,
tBu
tBu
300 K, COSY): rotational isomers, ratio: 10:3; ꢂ ¼ 1.43 (s, CH3 maj), 1.45 (s, CH3 min), 1.97 (s,
OAc min), 1.98 (s, OAc min), 1.99 (s, OAc maj), 2.02 (s, OAc minþ maj), 2.03 (s, OAc maj), 2.04 (s,
Gly
Nasn
OAc minþ maj), 3.63 (dd, J ¼ 4.8, 18.0Hz, CH2 maj), 3.72 (d, J ¼ 16.2 Hz, CH2
maj), 3.80 (m,
Gly
5-H min), 3.82 (m, 5-H maj), 3.87 (dd, J ¼ 5.4, 18.0Hz, CH2
maj), 3.95 (dd, J ¼ 5.4, 18.0 Hz,
Gly
Gly
Nasn
CH2 min), 3.99 (dd, J ¼ 4.8, 18.0 Hz, CH2 min), 4.01 (d, J ¼ 18.0Hz, CH2
maj), 4.07–4.24
Nasn
Gly
Nasn
Gly
(m, 6-H2, CH2
J ¼ 18.0Hz, CH2
(4H), CH2
(2H), CH2
(1H), CH2
Gly
(1H)), 4.33 (d, 1H,
maj
min
min
maj
Nasn
maj), 4.46 (dd, 1H, J ¼ 6.0, 16.8Hz, CH2
maj), 5.00–5.10 (m, 2,4-H),
5.24–5.31 (m, 1,3-H), 7.10 (br.t, J ¼ 5.4 Hz, NHGly min), 7.34 (br.t, J ¼ 4.8 Hz, NHGly min), 7.41 (t,
J ¼ 7.8 Hz, Hmeta min), 7.44 (t, J ¼ 7.8 Hz, Hmeta maj), 7.48–7.52 (m, Hpara), 7.54 (br.t, J ¼ 5.4 Hz,
NHGly maj), 7.57 (br.t, J ¼ 5.4 Hz, NHGly maj), 7.81 (d, J ¼ 7.8 Hz, Hortho min), 7.85 (d, J ¼ 7.8 Hz,
Hortho maj), 8.82 (d, J ¼ 9.6Hz, 1-NH maj), 9.47 (d, J ¼ 9.3 Hz, 1-NH min) ppm; 13C NMR (CDCl3,
50MHz, 300 K): ꢂ ¼ 20.71, 20.86, 21.07 (OAc), 28.17 (CH3tBu), 41.71 (CH2
min), 42.11
Gly
Gly
(2CH2 maj), 42.31 (CH2 min), 53.00 (CH2
Gly
Nasn
Nasn
Nasn
maj), 53.10 (CH2
min), 53.28 (CH2
min), 54.81
Nasn
(CH2
maj), 61.82 (6-CH2 min), 62.07 (6-CH2 maj), 68.23 (4-CH min), 68.42(4-CHmaj), 70.75 (2-CH maj),
70.96 (2-CH min), 73.12 (3-CH maj), 73.46 (3-CH min), 73.76 (5-CH maj), 73.90 (5-CH min), 78.13
(1-CH maj), 78.27 (1-CH min), 82.60 (CtBu), 127.26, 128.74, 131.99, 133.48 (maj), 133.60 (min), 167.29,
167.62, 168.70, 168.86, 169.03, 169.15, 169.62, 169.69, 169.75, 169.95, 170.06, 170.14, 170.29, 170.53,
170.72,170.89, 171.49ppm;IR(KBr):ꢃꢀ¼ 3600–3300, 1754, 1652, 1535 cmꢃ1;MS(FAB):m=z ¼ 737.3
[Mþ H]þ.
N-(3,4,6-Tri-O-acetyl-2-acetamido-2-deoxy-ꢁ-D-glucopyranosyl)-2-[5-oxo-2,2-
bis(trifluoromethyl)-1,3-oxazolidin-3-yl]acetamide (4b, C21H25F6N3O11)
Compound 3 (1.89 g, 6.30 mmol) and Ac3-ꢁ-D-GlcNAc-NH2 (2.18 g, 6.30mmol) were reacted due to
protocol 1. Purification by flash chromatography (ethyl acetate:petroleum ether¼ 5:1, Rf ¼ 0.40). Yield
2.72g (71%), foam; [ꢀ]D ¼ þ11ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 2.5, DCM); 1H NMR (DMSO-d6, 400 MHz,
COSY): ꢂ ¼ 1.74 (s, NAc), 1.91, 1.97, 2.00 (3s, 3OAc), 3.69 (d, 1H, J ¼ 16.8Hz, CH2Nasn),
3.74 (d, 1H, J ¼ 16.8 Hz, CH2Nasn), 3.86 (ddd, J ¼ 2.1, 4.5, 9.9Hz, 5-H), 3.94 (m, 2-H), 3.97 (dd,
J ¼ 2.1, 12.6Hz, 6-Ha), 4.15 (s, CH2Nasn), 4.18 (dd, J ¼ 4.5, 12.6 Hz, 6-Hb), 4.83 (dd, J ¼ 9.6, 9.9Hz,
4-H), 5.12 (dd, J ¼ 9.9, 9.9 Hz, 3-H), 5.18 (dd, J ¼ 9.6, 9.3 Hz, 1-H), 7.95 (d, J ¼ 9.3 Hz, NHAc), 8.71
(d, J ¼ 9.3 Hz, 1-NH) ppm; 19F NMR (DMSO-d6, 282 MHz): ꢂ ¼ 2.21 (q, 3F, J ¼ 8.0 Hz), 2.50 (q, 3F,
J ¼ 8.0 Hz) ppm; 13C NMR (DMSO-d6, 100 MHz, HMQC): ꢂ ¼ 20.28, 20.31, 20.43 (3OAc), 22.33
(NHAc), 48.31, 48.76 (2CH2Nasn), 52.07 (2-CH), 61.74 (6-CH2), 68.36 (4-CH), 72.40 (5-CH), 73.12 (3-
CH), 78.02 (1-CH), 89.41 (sept, J ¼ 33Hz), 120.72 (q, J ¼ 292Hz), 167.47, 167.56, 169.26, 169.49,
169.53, 169.95 ppm; IR (KBr): ꢃꢀ¼ 3400, 1847, 1749, 1662, 1524 cmꢃ1; MS (FAB): m=z ¼ 610.1
[Mþ H]þ.
tert-Butyl N-{2-Oxo-2-[(3,4,6-tri-O-acetyl-2-acetamido-2-deoxy-ꢁ-D-glucopyranosyl)-
amino]ethyl}glycylglycinate (H-Nasn(Ac3-ꢁ-D-GlcNAc)-Gly-OtBu, 5b=1, C24H38N4O12)
t
ꢀ
HCl H-Gly-O Bu (152 mg, 0.91mmol) and 4b (436mg, 0.72 mmol) were reacted due to protocol 2.
Reaction time: 18h. Purification by flash chromatography (DCM:MeOH¼ 10:1, Rf ¼ 0.32). Yield
305 mg (74%), foam; [ꢀ]D ¼ ꢃ11ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.1, DCM); 1H NMR (CDCl3, 300MHz,
COSY): ꢂ ¼ 1.44 (s, 9H, CH3tBu), 1.90 (s, NAc), 2.01, 2.02, 2.05 (3s, 3OAc), 3.25 (d, 1H, J ¼ 16.7Hz,
CH2Nasn), 3.28 (s, CH2Nasn), 3.33 (d, 1H, J ¼ 16.7Hz, CH2Nasn), 3.79 (ddd, J ¼ 2.1, 4.5, 9.6 Hz, 5-H),
3.91 (dd, 1H, J ¼ 5.8, 17.8Hz, CH2Gly), 3.99 (dd, 1H, J ¼ 5.8, 17.8Hz, CH2Gly), 4.07 (dd, J ¼ 2.1,
12.6Hz, 6-Ha), 4.12 (m, 2-H), 4.24 (dd, J ¼ 4.5, 12.6Hz, 6-Hb), 5.04 (dd, J ¼ 9.3, 9.9 Hz, 4-H), 5.10
(dd, J ¼ 9.6, 9.3 Hz, 1-H), 5.13 (dd, J ¼ 9.9, 9.3Hz, 3-H), 6.73 (d, J ¼ 9.0 Hz, NHAc), 7.73 (br.t,
J ¼ 5.8 Hz, NHGly), 8.12 (d, J ¼ 9.0 Hz, 1-NH) ppm; 13C NMR (CDCl3, 75MHz, HETCOR):
ꢂ ¼ 20.67, 20.76, 20.81 (3OAc), 23.11 (NHAc), 28.15 (CH3tBu), 41.64 (CH2Gly), 52.63, 52.65

872
K. Burger et al.
(2CH2Nasn), 53.25 (2-CH), 61.96 (6-CH2), 68.24 (4-CH), 72.88 (3-CH), 73.52 (5-CH), 79.72 (1-CH),
82.81 (CtBu), 169.41, 169.47, 170.80, 171.30, 171.54, 172.20, 172.31 ppm; IR (KBr): ꢃꢀ¼ 3400, 1847,
1749, 1662, 1524 cmꢃ1; HRMS (ESI): calcd. for C24H39N4O12 (Mþ Hþ) 575.25590, found
575.25592.
tert-Butyl N-{2-Oxo-2-[(3,4,6-tri-O-acetyl-2-acetamido-2-deoxy-ꢁ-D-glucopyranosyl)-
amino]ethyl}glycylalaninate (H-Nasn(Ac3-ꢁ-D-GlcNAc)-Ala-OtBu, 7b, C25H40N4O12)
t
ꢀ
HCl H-Ala-O Bu (218 mg, 1.20 mmol) and 4b (609mg, 1.00mmol) were reacted due to protocol 2.
Reaction time: 2d. Purification by flash chromatography (DCM:MeOH¼ 15:1, Rf ¼ 0.35). Yield
1
450 mg (76%), foam; [ꢀ]D ¼ ꢃ14ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.3, DCM); H NMR (DMSO-d6, 400 MHz,
COSY): ꢂ ¼ 1.27 (d, J ¼ 7.2 Hz, CH3Ala), 1.40 (s, 9H, CH3tBu), 1.77 (s, NAc), 1.92, 1.97, 2.00
(3s, 3OAc), 3.07 (d, 1H, J ¼ 16.5 Hz, CH2Nasn), 3.08 (s, CH2Nasn), 3.15 (d, 1H, J ¼ 16.5 Hz,
CH2Nasn), 3.86 (m, 5-H), 3.91–3.97 (m, 2-H, 6-Ha), 4.15–4.20 (m, 6-Hb, CHAla), 4.83 (dd,
J ¼ 9.3, 9.6 Hz, 4-H), 5.15 (dd, J ¼ 9.6, 9.9 Hz, 1-H, 3-H), 7.99 (d, J ¼ 9.0 Hz, NHAc), 8.11 (d,
J ¼ 7.2 Hz, NHAla), 8.45 (d, J ¼ 9.0 Hz, 1-NH) ppm; 13C NMR (DMSO-d6, 100MHz, HMQC):
ꢂ ¼ 17.03 (CH3Ala), 20.29, 20.32, 20.42 (3OAc), 22.42 (NHAc), 27.52 (CH3tBu), 47.93 (CHAla),
51.43, 51.55 (2CH2Nasn), 51.94 (2-CH), 61.76 (6-CH2), 68.47 (4-CH), 72.12 (5-CH), 72.73 (3-CH),
78.07 (1-CH), 80.36 (CtBu), 169.22, 169.42, 169.93 (2C), 170.48, 171.56, 171.73ppm; IR (KBr):
ꢃꢀ¼ 3600–3300, 1745, 1665, 1534 cmꢃ1; HRMS (ESI): calcd. for C25H41N4O12 (Mþ Hþ)
589.27155, found 589.27140.
tert-Butyl N-{2-Oxo-2-[(3,4,6-tri-O-acetyl-2-acetamido-2-deoxy-ꢁ-D-glucopyranosyl)-
amino]ethyl}glycylsarcosinate (H-Nasn(Ac3-ꢁ-D-GlcNAc)-Nala-OtBu, 5b=2, C25H40N4O12)
t
ꢀ
HCl H-Sar-O Bu (191mg, 1.05 mmol) and 4b (536 mg, 0.88 mmol) were reacted due to protocol 2.
Reaction time: 4d. Purification by flash chromatography (DCM:MeOH¼ 15:1, Rf ¼ 0.21). Yield
235 mg (45%), foam; [ꢀ]D ¼ þ1ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 2.3, DCM); 1H NMR (CDCl3, 200MHz,
tBu
tBu
300 K, COSY): rotational isomers, ratio: 3:1; ꢂ ¼ 1.45 (s, CH3 maj), 1.46 (s, CH3 min), 1.90 (s,
NHAc), 2.02, 2.03, 2.07 (3s, 3OAc), 2.96 (s, NCH3 min), 2.97 (s, NCH3 maj), 3.15–3.56 (m, 2CH2Nasn),
3.77 (m, 5-H), 3.86–4.27 (m, CH2Nala, 6-H2, 2-H), 5.07–5.21 (m, 1,3,4-H), 6.14 (d, J ¼ 9.3 Hz,
NHAc), 8.13 (d, J ¼ 9.6 Hz, 1-NH min), 8.17 (d, J ¼ 9.6 Hz, 1-NH maj) ppm; 13C NMR (CDCl3,
50MHz, 300 K, HETCOR): ꢂ ¼ 20.73, 20.82, 20.88 (3OAc), 23.24 (NHAc), 28.18 (CH3tBu), 35.19
Nala
(CH3
Nala
min), 35.24 (CH3
maj), 50.40 (CH2Nala, CH2Nasn), 52.73 (CH2Nasn), 53.39 (2-CH), 61.99
(6-CH2), 68.17 (4-CH), 73.13 (3-CH), 73.75 (5-CH), 79.34 (1-CH), 82.09 (CtBu), 168.21, 169.44,
170.80, 170.99, 171.05, 171.33, 171.38, 171.47, 172.73, 172.81 ppm; IR (KBr): ꢃꢀ¼ 3600–3300,
1745, 1662, 1522 cmꢃ1; HRMS (ESI): calcd. for C25H41N4O12 (Mþ Hþ) 589.27155, found
589.27144.
tert-Butyl N-Trifluoroacetyl-N-benzylglycyl-N-{2-oxo-2-[(3,4,6-tri-O-acetyl-2-
acetamido-2-deoxy-ꢁ-D-glucopyranosyl)amino]ethyl}glycylsarcosinate
(TFA-Nphe-Nasn(Ac3-ꢁ-D-Glc-NAc)-Nala-OtBu, 6b, C36H48F3N5O14)
TFA-Nphe-OH (52 mg, 0.20 mmol) and 5b=2 (100 mg, 0.17mmol) were reacted due to protocol 3.
Purification by flash chromatography (DCM:MeOH¼ 10:1, Rf ¼ 0.32). Yield 78 mg (55%), colorless
solid; purity >95% (HPLC); mp 171–173ꢂC; [ꢀ]D ¼ ꢃ4ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.6, DCM); 19F NMR
(CDCl3, 282 MHz, 300 K): 2 complex signal groups, ratio: 1:4; ꢂ ¼ 8.68 (s), 9.50 (s), 9.54 (s), 9.61 (s)
ppm; IR (KBr): ꢃꢀ¼ 1745, 1668, 1541 cmꢃ1; HRMS (ESI): calcd. for C36H49F3N5O14 (Mþ Hþ)
854.30421, found 854.30332; MS (FAB): m=z ¼ 854.3 [M þ Na]þ, 832.3 [Mþ H]þ, 776.3

Hexafluoroacetone as Protecting and Activating Reagent
873
[M-C4H8 þ H]þ, 687.2 [M-(Sar-OtBu)]þ, 486.1 [M-(Ac3-GlcNAc-NH)]þ, 430.0 [M-(Ac3-GlcNAc-
NH)-C4H8)]þ, 330.0 [Ac3-GlcNAc]þ.
N-(2,3,4,6-Tetra-O-acetyl-ꢁ-D-galactopyranosyl)-2-[5-oxo-2,2-bis(trifluoromethyl)-
1,3-oxazolidin-3-yl]acetamide (4c, C21H24F6N2O12)
Compound 3 (4.79 g, 16.0mmol) and Ac4-ꢁ-D-Gal-NH2 (5.56 g, 16.0 mmol) were reacted due to
protocol 1. Purification by flash chromatography (ethyl acetate:petrolether¼ 1:1, Rf ¼ 0.38). Yield
7.63g (78%), foam; [ꢀ]D ¼ þ13ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 2.3, DCM); 1H NMR (CDCl3, 300MHz, COSY):
ꢂ ¼ 1.99, 2.03, 2.05, 2.17 (4s, 4OAc), 3.64 (d, 1H, J ¼ 16.8 Hz, CH2Nasn), 3.70 (d, 1H, J ¼ 16.8Hz,
CH2Nasn), 3.76 (s, CH2Nasn), 4.03–4.12 (m, 6-H2, 5-H), 5.05 (dd, J ¼ 9.6, 9.9Hz, 2-H), 5.13 (dd,
J ¼ 3.3, 10.2Hz, 3-H), 5.19 (dd, J ¼ 9.0, 9.3 Hz, 1-H), 5.41 (d, J ¼ 3.3 Hz, 4-H), 7.10 (d, J ¼ 9.0 Hz,
1-NH) ppm; 19F NMR (CDCl3, 282 MHz): ꢂ ¼ 1.49 (m, 3F), 1.56 (m, 3F) ppm; 13C NMR (CDCl3,
75MHz, HETCOR): ꢂ ¼ 20.86, 20.90, 20.96, 20.99 (4OAc), 49.71, 51.66 (2CH2Nasn), 61.40 (6-CH2),
67.47 (4-CH), 68.76 (2-CH), 71.06 (3-CH), 72.95 (5-CH), 78.87 (1-CH), 90.16 (sept, J ¼ 33Hz),
121.27 (q, J ¼ 291 Hz), 121.34 (q, J ¼ 291 Hz, CF3), 166.15, 167.64, 170.14, 170.42, 170.76,
171.56 ppm; IR (KBr): ꢃꢀ¼ 1850, 1752, 1707, 1523 cmꢃ1; MS (FAB): m=z ¼ 611.2 [Mþ H]þ.
tert-Butyl N-{2-Oxo-2-[(2,3,4,6-tetra-O-acetyl-ꢁ-D-galactopyranosyl)amino]ethyl}glycyl-
glycinate (H-Nasn(Ac4-ꢁ-D-Gal)-Gly-OtBu, 5c=1, C24H37N3O13)
t
ꢀ
HCl H-Gly-O Bu (330mg, 1.97mmol) and 4c (1000mg, 1.64 mmol) were reacted due to protocol 2.
Reaction time: 18h. Purification by flash chromatography (CH2Cl2:MeOH¼ 20:1, Rf ¼ 0.29). Yield
696 mg (74%), colorless foam; [ꢀ]D ¼ þ13ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.2, DCM); 1H NMR (CDCl3,
300 MHz, COSY): ꢂ ¼ 1.46 (s, 9H, CH3tBu), 1.97, 2.02, 2.09, 2.13 (4s, 4OAc), 3.26 (d, 1H, J ¼ 16.7Hz,
CH2Nasn), 3.30 (s, 2H, CH2Nasn), 3.35 (d, 1H, J ¼ 16.7Hz, CH2Nasn), 3.96 (d, J ¼ 5.7 Hz, CH2Gly),
3.98–4.11 (m, 5-H, 6-H2), 5.06–5.17 (m, 2,3-H), 5.22 (dd, J ¼ 9.0, 9.3 Hz, 1-H), 5.42 (d, J ¼ 2.3 Hz,
4-H), 7.15 (br.t, J ¼ 5.7 Hz, NHGly), 7.63 (d, J ¼ 9.0 Hz, 1-NH) ppm; 13C NMR (CDCl3, 75 MHz,
HETCOR): ꢂ ¼ 20.65, 20.70, 20.78, 20.90 (4OAc), 28.15 (CH3tBu), 41.72 (CH2Gly), 52.30, 52.40
(2CH2Nasn), 61.28 (6-CH2), 67.27 (4-CH), 68.70 (2-CH), 70.85 (3-CH), 72.51 (5-CH), 78.35
(1-CH), 82.49 (CtBu), 169.29, 169.85, 170.13, 170.54, 170.99, 171.58, 171.66 ppm; IR (KBr):
ꢃꢀ¼ 3600–3300, 1747, 1662, 1535 cmꢃ1; HRMS (ESI): calcd. for C24H38N3O13 (Mþ Hþ)
576.23991, found 576.23935.
tert-Butyl N-{2-Oxo-2-[(2,3,4,6-tetra-O-acetyl-ꢁ-D-galactopyranosyl)amino]ethyl}glycyl-
leucinate (H-Nasn(Ac4-ꢁ-D-Gal)-Leu-OtBu, 7c, C28H45N3O13)
t
ꢀ
HCl H-Leu-O Bu (440mg, 1.97 mmol) and 4c (1000mg, 1.64mmol) were reacted due to protocol 2.
Reaction time: 2 d. Purification by flash chromatography (DCM:MeOH¼ 23:1, Rf ¼ 0.30). Yield
710 mg (68%), foam; [ꢀ]D ¼ þ12ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.0, DCM); 1H NMR (CDCl3, 300 MHz,
COSY): ꢂ ¼ 0.96 (d, J ¼ 6.0 Hz, CH3Leu), 0.97 (d, J ¼ 6.3 Hz, CH3Leu), 1.48 (s, 9H, CH3tBu), 1.57–
1.68 (m, ꢁ-CH2Leu, ꢄ-CHLeu), 2.01, 2.05, 2.08, 2.16 (4s, 4OAc), 3.26 (d, 1H, J ¼ 15.9 Hz, CH2Nasn),
3.32 (s, 2H, CH2Nasn), 3.33 (d, 1H, J ¼ 15.9Hz, CH2Nasn), 4.04–4.14 (m, 6-H2, 5-H), 4.55 (m,
ꢀ-CHLeu), 5.10–5.15 (m, 2,3-H), 5.25 (dd, J ¼ 9.0, 9.3 Hz, 1-H), 5.45 (d, J ¼ 2.1 Hz, 4-H), 6.85 (d,
J ¼ 8.4 Hz, NHLeu), 7.63 (d, J ¼ 9.0 Hz, 1-NH) ppm; 13C NMR (CDCl3, 75 MHz, APT, HETCOR):
ꢂ ¼ 20.65, 20.70, 20.79, 20.87 (4OAc), 22.01 (CH3Leu), 22.98 (CH3Leu), 25.12 (ꢄ-CHLeu), 28.09
(CH3tBu), 41.55 (ꢁ-CH2Leu), 51.30 (ꢀ-CHLeu), 52.21, 52.29 (2CH2Nasn), 61.29 (6-CH2), 67.31
(4-CH), 68.52 (2-CH), 70.92 (3-CH), 72.56 (5-CH), 78.39 (1-CH), 82.19 (CtBu), 169.88, 170.12,
170.42, 170.58, 171.47, 171.74, 172.44ppm; IR (KBr): ꢃꢀ¼ 3500–3300, 1747, 1660, 1531 cmꢃ1
;
HRMS (ESI): calcd. for C28H46N3O13 (Mþ Hþ) 632.30251, found 632.30422.

874
K. Burger et al.
tert-Butyl N-{2-Oxo-2-[(2,3,4,6-tetra-O-acetyl-ꢁ-D-galactopyranosyl)amino]ethyl}glycyl-
N-isobutylglycinate (H-Nasn(Ac4-ꢁ-D-Gal)-Nleu-OtBu, 5c=2, C28H45N3O13)
H-Nleu-OtBu (504 mg, 2.69 mmol) and 4c (1.37 g, 2.24 mmol) were reacted in ethyl acetate (10 cm3)
due to protocol 2. Reaction time: 4 d. Purification by flash chromatography (gradient elution ethyl
acetate:petrolether¼ 3:1 ! 6:1, Rf ¼ 0.25). Yield 690mg (49%), foam; [ꢀ]D ¼ þ15ꢂ cm3 gꢃ1 dmꢃ1
(c ¼ 2.5, DCM); 1H NMR (CDCl3, 400 MHz, 300 K, COSY): rotational isomers, ratio: 1.9:1; ꢂ ¼ 0.90
Nleu
Nleu
Nleu
(d, J ¼ 6.6 Hz, CH3
min), 0.93 (d, J ¼ 6.4 Hz, CH3
min), 0.94 (d, J ¼ 6.6 Hz, CH3
maj), 0.95 (d,
Nleu
tBu
tBu
J ¼ 6.4 Hz, CH3
(several s, OAc), 3.04 (d, J ¼ 7.5 Hz, CH2 maj), 3.20–3.38 (m, CH2 min, 2CH2
maj), 1.47 (s, CH3 maj), 1.48 (s, CH3 min), 1.82–1.90 (m, CHiBu), 1.99–2.17
iBu
iBu
Nasn
Nasn
min, CH2
maj), 3.81 (d, 1H, J ¼ 18.5Hz,
maj), 4.01–4.15 (m, 5-H, 6-H2),
maj),
Nasn
Nasn
3.40 (d, 1H, J ¼ 16.3Hz, CH2
maj), 3.50 (d, 1H, J ¼ 16.3 Hz, CH2
Nleu Nleu
Nleu
CH2
min), 3.89 (d, 1H, J ¼ 18.5Hz, CH2
min), 3.96 (s, CH2
5.12 (dd, J ¼ 2.8, 10.0Hz, 3-H), 5.17 (dd, J ¼ 10.0, 9.3 Hz, 2-H), 5.25 (dd, J ¼ 9.3, 9.6 Hz, 1-H min),
5.26 (dd, J ¼ 9.3, 9.6 Hz, 1-H maj), 5.44 (d, J ¼ 2.8 Hz, 4-H), 8.08 (d, J ¼ 9.6 Hz, 1-NH) ppm; 13C NMR
Nleu
Nleu
Nleu
(CDCl3, 100 MHz, 300 K, HMQC): ꢂ ¼ 20.10 (CH3
maj), 20.13 (CH3
maj), 20.20 (2CH3
min),
maj), 50.18
min), 50.49, 52.91 (2CH2Nasn), 55.33 (CH2iBu), 61.33 (6-CH2), 67.26 (4-CH), 68.61 (2-CH),
tBu
tBu
Nleu
20.61, 20.67, 20.73, 20.81 (4OAc), 28.07 (CH3 min), 28.14 (CH3 maj), 49.01 (CH2
Nleu
(CH2
71.12 (3-CH), 72.47 (5-CH), 78.10 (1-CH), 81.96 (CtBu maj), 82.01 (CtBu min), 168.10, 169.88, 170.15,
170.41, 170.51, 171.28, 172.36ppm; IR (KBr): ꢃꢀ¼ 3500–3300, 1746, 1650 cmꢃ1; HRMS (ESI): calcd.
for C28H46N3O13 (Mþ Hþ) 632.30251, found 632.30340.
tert-Butyl N-Benzoylglycyl-N-{2-oxo-2-[(2,3,4,6-tetra-O-acetyl-ꢁ-D-galactopyranosyl)-
amino]ethyl}glycylleucinate (Bz-Gly-Nasn(Ac4-ꢁ-D-Gal)-Leu-OtBu, 8c, C37H52N4O15)
Bz-Gly-OH (116mg, 0.65 mmol) and 7c (341mg, 0.54 mmol) were reacted due to protocol 3. Purifica-
tion by flash chromatography (CHCl3:MeOH¼ 10:1, Rf ¼ 0.24). Yield 269 mg (62%), colorless crys-
1
tals; mp 107–108ꢂC; [ꢀ]D ¼ ꢃ3ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 2.0, DCM); H NMR (CDCl3, 600MHz, 300 K,
tBu
COSY): rotational isomers, ratio: 3:1; ꢂ ¼ 0.90–0.97 (m, CH3Leu), 1.46 (s, CH3
maj), 1.47 (s,
CH3 min), 1.53–1.73 (m, ꢁ-CH2Leu, ꢄ-CHLeu), 1.92–2.13 (several s, OAc), 3.82 (d, 1H, J ¼ 16.3Hz,
tBu
Nasn
Hz, CH2
Nasn
Nasn
(3H), 2CH2
maj), 3.99–4.35 (m, CH2
min, CH2Gly, 5-H, 6-H2), 4.42 (m,
maj
ꢀ-CHLeu maj), 4.51 (m, ꢀ-CHLeu min), 5.12 (dd, J ¼ 3.0, 10.1Hz, 3-H), 5.16 (dd, J ¼ 9.0, 10.1Hz, 2-
H), 5.26 (dd, J ¼ 9.0, 10.1Hz, 1-H maj), 5.30 (dd, J ¼ 9.0, 10.1Hz, 1-H min), 5.42 (d, J ¼ 3.0 Hz,
4-H min), 5.43 (d, J ¼ 3.0 Hz, 4-H maj), 6.86 (d, J ¼ 7.2 Hz, NHLeu min), 7.26 (partially covered by
the chloroform signal, NHGly min), 7.38 (br.t, J ¼ 4.8 Hz, NHGly maj), 7.41–7.44 (m, Hmeta), 7.49–7.52
(m, Hpara), 7.60 (d, J ¼ 7.2 Hz, NHLeu maj), 7.84 (d, J ¼ 7.3, Hortho min), 7.86 (d, J ¼ 7.3 Hz, Hortho maj),
8.28 (d, J ¼ 9.0Hz, 1-NH maj), 9.08 (d, J ¼ 9.0 Hz, 1-NH min) ppm; 13C NMR (CDCl3, 150 MHz,
Leu
Leu
Leu
Leu
300 K, HMQC): ꢂ ¼ 20.6–21.0 (OAc), 22.02 (CH3
maj), 22.15 (CH3
min), 22.74 (CH3
maj),
22.82 (CH3 min), 24.87 (ꢄ-CHLeu min), 25.12 (ꢄ-CHLeu maj), 28.08 (CH3tBu), 41.13 (ꢁ-CH2 maj),
Leu
Leu
Gly
Gly
41.60 (ꢁ-CH2
CHLeu maj), 52.72 (CH2
min), 41.65 (CH2
Nasn
min), 41.73 (CH2
Nasn
maj), 51.87 (ꢀ-CHLeu min), 52.13 (ꢀ-
Nasn
maj), 52.87 (CH2
min), 54.05 (CH2
maj þ min), 61.22 (6-CH2), 67.18
(4-CH), 68.46 (2-CH maj), 68.81 (2-CH min), 71.18 (3-CH maj), 71.33 (3-CH min), 72.49 (5-
CH maj), 72.66 (5-CH min), 78.48 (1-CH maj), 78.72 (1-CH min), 82.27 (CtBu maj þ min), 127.25, 128.67,
131.92, 133.62 (min), 133.72 (maj), 167.4–171.9 ppm; IR (KBr): ꢃꢀ¼ 3600–3300, 1753, 1654, 1536cmꢃ1
;
MS (FAB): m=z ¼ 793.3 [Mþ H]þ, 737.3 [M-C4H8 þ H]þ, 606.3 [M-Leu-OtBu]þ, 576.2 [M-(Bz-Gly)-
(C4H8) þ 2H]þ, 446.1 [M-(Ac4-Gal-NH)]þ, 390.3 [M-(Ac4-Gal-NH)-(C4H8)]þ, 331.0 [Ac4-Gal]þ.
N-[2,3,6-Tri-O-acetyl-4-O-(2,3,4,6-tetra-O-acetyl-ꢁ-D-glucopyranosyl)-
ꢁ-D-glucopyranosyl]-2-[5-oxo-2,2-bis(trifluoromethyl)-1,3-oxazolidin-
3-yl]acetamide (4d, C33H40F6N2O20)
Compound 3 (740mg, 2.47 mmol) and Ac4-ꢁ-D-Glc(1! 4)-Ac3-ꢁ-D-Glc-NH2 (1.57 g, 2.47 mmol)
were reacted due to protocol 1. Purification by flash chromatography (ethyl acetate:petroleum

Hexafluoroacetone as Protecting and Activating Reagent
875
ether ¼ 2:1, Rf ¼ 0.39). Yield 1.82g (82%), foam; [ꢀ]D ¼ þ8ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 2.0, DCM); 1H NMR
(CDCl3, 400 MHz, COSY): ꢂ ¼ 1.98, 2.01 (2s, 2OAc), 2.03 (s, 3OAc), 2.09, 2.12 (2s, 2OAc), 3.62–3.72
(m, 2CH2Nasn, 5-H, 50-H), 3.77 (dd, J ¼ 9.0, 9.0 Hz, 4-H), 4.05 (dd, J ¼ 1.8, 12.4Hz, 60-Ha), 4.12 (dd,
J ¼ 4.1, 12.1Hz, 6-Ha), 4.36 (dd, J ¼ 4.5, 12.1 Hz, 60-Hb), 4.49 (dd, J ¼ 1.0, 12.1Hz, 6-Hb), 4.51 (d,
J ¼ 8.0 Hz, 10-H), 4.84 (dd, J ¼ 9.6, 9.6 Hz, 2-H), 4.92 (dd, J ¼ 9.0, 8.2 Hz, 20-H), 5.06 (dd, J ¼ 9.5,
9.7 Hz, 40-H), 5.14 (dd, J ¼ 9.6, 9.3 Hz, 1-H, 30-H), 5.30 (dd, J ¼ 9.1, 9.4Hz, 3-H), 6.96 (d, J ¼ 9.0 Hz,
1-NH) ppm; 19F NMR (CDCl3, 282 MHz): ꢂ ¼ 0.21 (m, 3F), 0.40 (m, 3F) ppm; 13C NMR (CDCl3,
75MHz, HETCOR): ꢂ ¼ 20.45 (2OAc), 20.50 (3OAc), 20.63, 20.79 (2OAc), 49.29, 51.26 (2CH2Nasn),
61.28, 61.68 (6,60-CH2), 67.89 (40-CH), 70.85 (2-CH), 71.57 (20-CH), 71.99, 72.00 (50,3-CH), 72.88
(30-CH), 74.74, 76.10 (4,5-CH), 78.09 (1-CH), 89.72 (sept, J ¼ 33Hz), 100.65 (10-CH), 120.84 (q,
J ¼ 291 Hz), 120.94 (q, J ¼ 289 Hz), 165.62, 167.20, 168.97, 169.29, 169.38, 170.18 (2C), 170.45,
171.10; IR (KBr): ꢃꢀ¼ 1848, 1755, 1521 cmꢃ1; MS (FAB): m=z ¼ 899.5 [Mþ H]þ.
tert-Butyl N-{2-Oxo-2-[(2,3,6-tri-O-acetyl-4-O-(2,3,4,6-tetra-O-acetyl-
ꢁ-D-glucopyranosyl)-ꢁ-D-glucopyranosyl)amino]ethyl}glycylglycinate
(H-Nasn(Ac4-ꢁ-D-Glc-(1 ! 4)-Ac3-ꢁ-D-Glc)-Gly-OtBu, 5d=1, C36H53N3O21)
t
ꢀ
HCl H-Gly-O Bu (66 mg, 0.40 mmol) and 4d (297mg, 0.33 mmol) were reacted due to protocol 2.
Reaction time: 18h. Purification by flash chromatography (DCM:MeOH¼ 10:1, Rf ¼ 0.28). Yield
258 mg (90%), foam; [ꢀ]D ¼ ꢃ7ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.3, DCM); 1H NMR (CDCl3, 400 MHz, COSY):
ꢂ ¼ 1.49 (s, 9H, CH3tBu), 1.99, 2.01 (2s, 2OAc), 2.03 (s, 3OAc), 2.09, 2.12 (2s, 2OAc), 3.29–3.34 (m,
2CH2Nasn), 3.66 (ddd, J ¼ 2.3, 4.4, 9.8 Hz, 50-H), 3.74–3.78 (m, 4,5-H), 3.97 (d, J ¼ 5.6 Hz, CH2Gly),
4.05 (dd, J ¼ 2.3, 12.5Hz, 60-Ha), 4.13 (dd, J ¼ 3.4, 11.8Hz, 6-Ha), 4.36 (dd, J ¼ 4.4, 12.5 Hz, 60-Hb),
4.48 (dd, J ¼ 1.2, 11.8Hz, 6-Hb), 4.51 (d, J ¼ 8.0 Hz, 10-H), 4.88 (dd, J ¼ 9.6, 9.6 Hz, 2-H), 4.92 (dd,
J ¼ 9.3, 8.0 Hz, 20-H), 5.07 (dd, J ¼ 9.6, 9.6Hz, 40-H), 5.15 (dd, J ¼ 9.3, 9.3Hz, 30-H), 5.19 (dd,
J ¼ 9.3, 9.3 Hz, 1-H), 5.29 (dd, J ¼ 9.3, 9.3 Hz, 3-H), 7.14 (br.t, J ¼ 5.6 Hz, NHGly), 7.55 (d, J ¼ 9.3 Hz,
1-NH) ppm; 13C NMR (CDCl3, 100 MHz, HMQC): ꢂ ¼ 20.59 (2OAc), 20.62 (2OAc), 20.73, 20.77,
20.93 (3OAc), 28.13 (CH3tBu), 41.78 (CH2Gly), 51.99, 52.19 (2CH2Nasn), 61.78, 61.84 (6,60-CH2), 68.00
(40-CH), 71.15 (2-CH), 71.68 (20-CH), 72.11, 72.24 (3,50-CH), 72.98 (30-CH), 74.73, 76.15 (4,5-CH),
78.00 (1-CH), 82.78 (CtBu), 100.65 (10-CH), 169.25, 169.41, 169.48, 169.67, 170.40, 170.65, 170.68,
171.30, 171.63, 171.91 ppm; IR (KBr): ꢃꢀ¼ 3600–3300, 1749, 1655, 1540 cmꢃ1; HRMS (ESI): calcd.
for C36H54N3O21 (Mþ Hþ) 864.32443, found 864.32419.
tert-Butyl N-{2-Oxo-2-[(2,3,6-tri-O-acetyl-4-O-(2,3,4,6-tetra-O-acetyl-
ꢁ-D-glucopyranosyl)-ꢁ-D-glucopyranosyl)amino]ethyl}glycylprolinate
(H-Nasn(Ac4-ꢁ-D-Glc-(1 ! 4)-Ac3-ꢁ-D-Glc)-Pro-OtBu, 7d, C39H57N3O21)
t
ꢀ
HCl H-Pro-O Bu (247mg, 1.19 mmol) and 4d (890mg, 0.99mmol) were reacted due to protocol 2.
Reaction time: 3 d. Purification by flash chromatography (DCM:MeOH¼ 15:1, Rf ¼ 0.29). Yield
600 mg (67%), foam; [ꢀ]D ¼ ꢃ27ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.9, DCM); 1H NMR (CDCl3, 300 MHz,
300 K, COSY): rotational isomers, ratio: 5:1, ꢂ (major rotamer)¼ 1.44 (s, 9H, CH3tBu), 1.96, 1.99,
2.00 (3s, 3OAc), 2.01 (s, 2OAc), 2.08, 2.11 (2s, 2OAc), 1.85–2.20 (m, ꢁ,ꢄ-CH2Pro), 3.24 (d, 1H,
J ¼ 17.1Hz, CH2Nasn), 3.32 (s, 2H, CH2Nasn), 3.33 (d, 1H, J ¼ 17.1Hz, CH2Nasn), 3.36–3.51 (m, ꢂ-
Pro
CH2 ), 3.64 (m, 50-H), 3.71–3.78 (m, 4,5-H), 4.01–4.11 (m, 60,6-Ha), 4.32–4.44 (m, 60,6-Hb, ꢀ-
CHPro), 4.49 (d, J ¼ 8.1 Hz, 10-H), 4.84–4.93 (m, 2,20-H), 5.04 (dd, J ¼ 9.3, 9.6 Hz, 40-H), 5.13 (dd,
J ¼ 9.0, 9.3 Hz, 30-H), 5.20 (dd, J ¼ 9.3, 9.6 Hz, 1-H), 5.25 (dd, J ¼ 9.3, 9.6 Hz, 3-H), 7.83 (d,
J ¼ 9.6 Hz, 1-NH) ppm; 13C NMR (CDCl3, 75 MHz, 300 K): ꢂ (major rotamer)¼ 20.5–21.0 (7OAc),
24.64 (ꢄ-CH2Pro), 28.05 (CH3tBu), 29.07 (ꢁ-CH2Pro), 45.96 (ꢂ-CH2Pro), 50.97, 52.40 (2CH2Nasn), 59.82
(ꢀ-CHPro), 61.78, 61.93 (6,60-CH), 68.02, 71.10, 71.69, 72.11, 72.47, 73.05, 74.69, 76.22
(2,3,4,5,20,30,40,50-CH), 77.77 (1-CH), 81.95 (CtBu), 100.69 (10-CH), 169.14, 169.17, 169.44, 169.62,

876
K. Burger et al.
170.35, 170.43, 170.63, 170.71, 171.43, 172.41ppm; IR (KBr): ꢃꢀ¼ 3600–3300, 1751, 1645cmꢃ1
HRMS (ESI): calcd. for C39H58N3O21 (Mþ Hþ) 904.35573, found 904.35375.
;
tert-Butyl N-Trifluoroacetyl-N-isobutylglycyl-N-{2-oxo-2-[(2,3,6-tri-O-acetyl-4-O-(2,3,4,6-
tetra-O-acetyl-ꢁ-D-glucopyranosyl)-ꢁ-D-glucopyranosyl)amino]ethyl}glycylprolinate
(TFA-Nleu-Nasn(Ac4-ꢁ-D-Glc-(1 ! 4)-Ac3-ꢁ-D-Glc)-Pro-OtBu,
8d, C47H67F3N4O23)
TFA-Nleu-OH (45 mg, 0.20 mmol) and 7d (150mg, 0.17mmol) were reacted due to protocol 3.
Purification by chromatography (ethyl acetate:CHCl3:MeOH ¼ 5:5:1, Rf ¼ 0.30). Yield 80 mg (42%),
colorless solid; purity >95% (HPLC); mp 121–122ꢂC; [ꢀ]D ¼ ꢃ30ꢂ cm3 gꢃ1 dmꢃ1 (c ¼ 1.2, DCM);
19F NMR (CDCl3, 282 MHz, 300 K): 2 complex signal groups, ratio: 1:4; ꢂ ¼ 4.16 (s), 4.20 (s), 4.24
(s), 4.33 (s), 5.57 (s), 5.65 (s), 5.73 (s), 5.81 (s) ppm; HRMS (ESI): calcd. for C47H68F3N4O23
(Mþ Hþ) 1113.42210, found 1113.42221; MS (FAB): m=z ¼ 1135.4 [Mþ Na]þ, 1113.4 [Mþ H]þ,
1079.3 [M-C4H8 þ Na]þ, 942.4 [M-(Pro-OtBu)]þ, 478.3 [M-(Ac7-Cellobiosyl-NH)]þ, 422.2 [M-(Ac7-
Cellobiosyl-NH)-C4H8]þ, 331.2 [Ac4-Glc]þ.
Acknowledgement
The authors thank Deutsche Forschungsgemeinschaft and Fonds der Chemischen Industrie for financial
support. S.A.E. thanks DAAD for a grant.
References
[1] (a) Sewald N, Jakubke H-D (2002) Peptides: Chemistry and Biology. Wiley-VCH, Weinheim;
(b) Giannis A, Kolter T (1993) Angew Chem Int Ed Engl 32: 1244
€
[2] Olson GL, Bolin DR, Pat Bonner M, Bos M, Cook CM, Fry DC, Graves BJ, Hatada M, Hill DE,
Kahn M, Madison VS, Rusiecki VK, Sarabu R, Sepinwall J, Vincent GP, Voss ME (1993) J Med
Chem 36: 3039
[3] Morgan BA, Gainor JA (1989) Ann Rep Med Chem 24: 243
[4] Fisher JF, Harrison AW, Bundy GL, Wilkinson KF, Rush BD, Ruwart MJ (1991) J Med Chem 34:
3140
ꢁ
[5] Polt R, Szabo L, Treiberg J, Li Y, Hruby VJ (1992) J Am Chem Soc 114: 10249
[6] Hirschmann R, Nicolaou KC, Pietrancio S, Leahy EM, Salvino J, Arison B, Cichy Ma, Spoors
PG, Shakespeare WC, Sprengeler PA, Hamley P, Smith III AB, Reisine T, Rayor K, Maechler L,
Donaldson C, Vale W, Freidinger RM, Cascieri MR, Strader CD (1993) J Am Chem Soc 115:
12550
[7] Lit. [1a] pp 78 and references cited therein
[8] (a) Zuckermann RN, Kerr JM, Kent SBH, Moos WH (1992) J Am Chem Soc 114: 10646; (b)
Simon RJ, Kania RS, Zuckermann RN, Huebner VD, Jewell DA, Banville S, Ng S, Wang L,
Rosenberg S, Marlowe CK, Spellmeyer DC, Tan R, Frankel AD, Santi DV, Cohen FE, Bartlett PA
(1992) Proc Natl Acad Sci USA 89: 9367
[9] (a) Kessler H (1993) Angew Chem Int Ed Engl 32: 543; (b) Horwell DC (1995) TIBTECH 13:
€
132; (c) Mohle K, Hofmann H-J (1996) J Mol Model 2: 307
[10] Miller SM, Simon RJ, Ng S, Zuckermann RN, Kerr JM, Moos WH (1994) Bioorg Med Chem
Lett 4: 2657
[11] Armand P, Kishenbaum K, Goldsmith RA, Farr-Jones S, Barron KTV, Dill KA, Mierke DF,
Cohen FE, Zuckermann RN, Bradley EK (1998) Proc Natl Acad Sci USA 95: 4309
[12] Gante J (1994) Angew Chem Int Ed Engl 33: 1699

Hexafluoroacetone as Protecting and Activating Reagent
877
[13] (a) Saha UK, Roy R (1995) Tetrahedron Lett 36: 3635; (b) Roy R, Saha UK (1996) Chem
Commun 201
[14] (a) Kim JM, Roy R (1997) Carbohydrate Res 298: 173; (b) Hu Y-J, Roy R (1999) Tetrahedron
Lett 40: 3305
[15] Saha UK, Roy R (1997) Tetrahedron Lett 38: 7697
[16] Kim JM, Roy R (1997) Tetrahedron Lett 38: 3487
[17] Dechantsreiter MA, Burkhart F, Kessler H (1998) Tetrahedron Lett 39: 253
[18] Weygand F, Burger K, Engelhardt K (1966) Chem Ber 99: 1461