Synthesis of protected hydroxyethylamine transition-state analogs of N-Ac-muramyl-L-alanine

Article abstract of DOI:10.1080/00397910802044280

A novel N-acetyl-α-D-glucosamine derivative containing an epoxide moiety 5 has been synthesized and converted into a series of protected hydroxyethylamine transition-state analogs of N-Ac-muramyl-L-ala peptide 6a-e using a microwave-accelerated reaction. Copyright Taylor & Francis Group, LLC.

Full text of DOI:10.1080/00397910802044280

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Synthesis of Protected
Hydroxyethylamine Transition-
State Analogs of N-Ac-Muramyl-
L-alanine
Andrej Babič a & Slavko Pečar a b
a Faculty of Pharmacy, University of Ljubljana ,
Ljubljana, Slovenia
b Institute Jožef Stefan , Ljubljana, Slovenia
Published online: 09 Sep 2008.
To cite this article: Andrej Babič & Slavko Pečar (2008) Synthesis of Protected
Hydroxyethylamine Transition-State Analogs of N-Ac-Muramyl-L-alanine, Synthetic
Communications: An International Journal for Rapid Communication of Synthetic
Organic Chemistry, 38:18, 3052-3061
To link to this article: http://dx.doi.org/10.1080/00397910802044280
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Synthetic Communications1, 38: 3052–3061, 2008
Copyright # Taylor & Francis Group, LLC
ISSN: 0039-7911 print/1532-2432 online
DOI: 10.1080/00397910802044280
Synthesis of Protected Hydroxyethylamine
Transition-State Analogs of
N-Ac-Muramyl-L-alanine
1,2
ˇ1
ˇ
Andrej Babic and Slavko Pecar
1Faculty of Pharmacy, University of Ljubljana, Ljubljana, Slovenia
2Institute Jozˇef Stefan, Ljubljana, Slovenia
Abstract: A novel N-acetyl-a-D-glucosamine derivative containing an epoxide
moiety 5 has been synthesized and converted into a series of protected hydro-
xyethylamine transition-state analogs of N-Ac-muramyl-L-ala peptide 6a–e using
a microwave-accelerated reaction.
Keywords: N-Ac-muramyl-L-ala,
synthesis, transition-state analogs
hydroxyethylamine,
microwave-assisted
INTRODUCTION
Hydroxyethylamine (HEA) transition-state analogs are a well-established
class of compounds with significant relevance in medicinal chemistry.[1–3]
Our research is directed toward new HEA transition-state inhibitors of
Mur ligases. The latter constitute a group of enzymes catalyzing cytoplas-
mic steps of bacterial peptidoglycan biosynthesis in which several amino
acids are attached to the growing peptidoglycan precursor.[4] The first
ligase in line is MurC ligase, which attaches L-alanine to UDP-muramic
acid.[5] Thus, the tetrahedral transition state could be mimicked by an
HEA analog. The most straightforward synthetic route toward this type
of HEA analog involves a reaction between an epoxide and an appropri-
ate amino acid, however, most reported procedures suffer from several
drawbacks such as low yields, high excesses of reagents, and prolonged
Received December 17, 2007.
Address correspondence to Andrej Babic, Faculty of Pharmacy, University of
ˇ
Ljubljana, Askerceva 7, 1000 Ljubljana, Slovenia. E-mail: andrej.babic@ffa.
uni-ij.si
ꢀ
ˇ
3052

Analogs of N-Ac-Muramyl-L-alanine
3053
reaction times.[6–8] For that reason, we have developed a reliable and
high-yield synthetic method involving a direct amino acid opening of
epoxide for the synthesis of HEA derivatives.[9,10] The reaction is cata-
lyzed by calcium trifluorosulfonate and is microwave accelerated, there-
fore, it is fast and appropriate for parallel synthesis. Herein we describe
the application of this reaction in the synthesis of HEA transition-state
analogs of N-Ac-muramyl-L-Ala peptide.
RESULTS AND DISCUSSION
To synthesize the desired compounds, we used the reaction pathway pre-
sented in Scheme 1. The starting material was a-D-glucosamine hydro-
chloride 1, which was selectively and quantitatively N-acetylated with
acetanhydride in methanol at ambient temperature.[11] Fischer glycosyla-
tion of 2 with benzyl alcohol gave predominantly the a-anomer of 3. It is
noteworthy that high a=b ratios were only obtainable if prolonged reac-
tion times in refluxing benzene were used. To leave the 30-hydroxy group
of the system available for further synthetic steps, the 40- and 60-hydroxy
groups were blocked with benzaldehyde to yield compound 4 in a
good yield, according to known procedures.[12] At this point, a common
way of introducing the epoxide moiety to the 30-hydroxy group would
be a two-step procedure involving Williamson ether synthesis with
3-bromopropene with subsequent oxidation of the double bond. How-
ever, a direct, one-step approach with epichlorohydrin was reasonably
Scheme 1. Reagent and conditions: (a) Na=MeOH (b) (Ac)2O, rt, 1 h, quant.;
(c) BnOH, PTSA, benzene, reflux, 24 h, 63%; (d) Ph-CHO, (EtO)3CH, PTSA,
DMF, dioxane, rt, 20 h, 76%; (e) NaH, epichlorhydrine, 15-crown-5, dioxane,
rt, 30 h, 81%.

ˇ ˇ
A. Babic and S. Pecar
3054
Scheme 2. Reagent and conditions: (a) NH2CHR1COOR2, Ca(OTf)2, dioxane,
MW, 120 ꢀC, 20 min, 45–60%.
successful. The reaction was carried out in dioxane with sodium hydride
as the base and 15-crown-5 as catalyst.[13] The appropriate crown ether
had to be used to increase the reaction rate at ambient temperature
because raised reaction temperatures lead to several side products and
much lower yields. Compound 5 was obtained as a mixture of two dia-
stereoisomers (dr ¼ 50:50) as no stereoselectivity of the reaction was
observed, even at room temperature.
As Scheme 2 indicates, compound 5 was reacted with various carboxy
protected L-amino acids to give compounds 6a–e (Table 1). The reaction
was carried out in a sealed reactor using microwave acceleration. Calcium
trifluorosulfonate (calcium triflate) is crucial for good yields, especially
considering the relatively high steric hindrance of epoxide 5. Using this
catalyst also left the protecting groups intact under the reaction con-
ditions selected. NMR indicated that optically active carboxy protected
L-amino acids reacted with both diastereoisomers without preference.
No by-products, resulting from the nucleophilic attack of the more steri-
cally hindered electrophilic carbon atom of the oxirane ring, were
detected. A small excess (1.2 eq.) of carboxy protected amino acids was
used to simplify purification. Regardless, some crude reaction products
proved troublesome to purify by flash chromatography.
Table 1. Reaction products 6a–e and reaction yields
Compound
R1
R2
Yield (%)
6a
6b
6c
6d
6e
H
CH3
CH(CH3)2
CH2CH(CH3)2
CH3
Bn
Bn
Bn
Bn
57
60
58
57
45
t-Bu

Analogs of N-Ac-Muramyl-L-alanine
3055
In conclusion, we have synthesized a series of protected HEA analogs
using a direct reaction between the epoxide 5 and several protected amino
acids. Use of calcium trifluorosulfonate and microwave acceleration
enabled the use of a small excess of amino acids and ensured short reac-
tion times with good yields.
EXPERIMENTAL
Chemicals from Acros and SigmaAldrich were used without further puri-
fication. Analytical thin-layer chromatography (TLC) was performed on
Merck silica gel (60 F 254) plates (0.25 mm), and components were visua-
lized with ultraviolet light and dyed with 20% sulphuric acid in ethanol,
2,4-dinitrophenalhydrazine, and ninhydrin. Microwave-accelerated reac-
tions were carried out using a monomode CEM microwave reactor. Flash
chromatography was carried out using Merck silica gel (40–63 mm).
Melting points were determined on a Reichert hot-stage microscope
1
and are uncorrected. H, 13C, DEPT-135, gradient homonuclear corre-
lation spectroscopy (COSY), and gradient heteronuclear single quantum
coherence spectroscopy (HSQC) NMR spectra were recorded on a
Bruker Avance DPX300 spectrometer in CDCl3 or dimethyl sulfoxide
(DMSO)-d6 solution with TMS as the internal standard. IR spectra were
obtained on a Perkin–Elmer 1600 Fourier transform infrared spec-
troscopy (FTIR) spectrometer. Microanalyses were performed on a
Perkin-Elmer C, H, N analyzer 240 C. Mass spectra were obtained using
1 VG-Analytical Autospec Q and Q-TOF Premier mass spectrometers.
2-Deoxy-2-acetylamido-D-glucopyranoside (2)
Compound 2 was synthesized according to Ref. 11. Amorphous solid:
30.1 g, 98%. a=b ratio ¼ 85=15. Mp 203–205 ꢀC. IR (KBr, cmꢁ1): 3326,
1
2910, 1627, 1550, 1428, 1290, 1126, 1054, 563, 726. H NMR (DMSO-
d6, 300 MHz) (a-anomer): d (ppm) ¼ 1.82 (s, 3H, COCH3), 3.06–3.17
(m, 1H, H-4), 3.31–4.69 (m, 5H, H-2, H-3, H-5, H-6, H-60), 4.41 (t,
1H, OH-6, J ¼ 6.3 Hz), 4.75 (d, 1H, OH-4, J ¼ 6.3 Hz), 4.92 (d, 1H,
OH-3, J ¼ 6.0 Hz), 4.97 (t, 1H, H-1, J ¼ 4.1 Hz), 6.39 (d, 1H, OH-1,
J ¼ 5.0 Hz), 7.68 (d, 1H, NH, J ¼ 7.9 Hz). MS (FAB), m=z ¼ 222
[MH]þ . The spectral data are in accordance with the reference material
purchased from Acros.
Benzyl-2-deoxy-2-acetylamido-a-D-glucopyranoside (3)
N-Acetyl-D-glucosamine (2) (25 g, 113 mmol) and 1.9 g (10 mmol) of
p-toluenesulfonic acid monohydrate were suspended in 300 ml of benzene.

ˇ ˇ
A. Babic and S. Pecar
3056
Benzyl alcohol (180 ml) was added, and the reaction mixture was
refluxed. The water produced in the reaction was removed with a
Dean–Stark apparatus. After 24 h, the solvents were removed under
reduced pressure. Ether–petrolether mixture (1:1) (700 ml) was added to
the oily residue and stirred vigorously for 3 h. The amorphous precipitate
was filtered off, and the crude product was recrystallized from ethanol to
yield colorless crystals: 22.2 g, 63%. Mp 182–185 ꢀC (ref. Mp 183–184 ꢀC).
1
IR (KBr, cmꢁ1): 3285, 2931, 1630, 1552, 1376, 1027, 734, 695. H NMR
(DMSO-d6, 300 MHz): d (ppm) ¼ 1.84 (s, 3H, COCH3), 3.09–3.20 (m,
1H, H-4), 3.41–4.75 (m, 5H, H-2, H-3, H-5, H-6, H-6’), 4.33 (t, 1H,
OH-6, J ¼ 5.7 Hz), 4.42 (d, 1H, CH2Ph, Jgem ¼ 12.6 Hz), 4.67 (d,
1H, CH2Ph, Jgem ¼ 12.6 Hz), 4.70–4.72 (m, 2H, H-1, OH-4), 5.0 (d,
1H, OH-3, J ¼ 5.6 Hz), 7.35–7.37 (m, 5H, Ph-H), 7.79 (d, 1H, NH,
J ¼ 8.1 Hz). MS (FAB), m=z ¼ 312 [MH]þ . The spectral data are in
accordance with Ref. 12.
Benzyl-4,6-O-benzylidene-2-deoxy-2-acetylamido-a-D-glucopyranoside (4)
Colorless solid: 14.6 g, 76%. Mp 262–263 ꢀC (ref. M. p. 262 ꢀC). IR (KBr,
cmꢁ1): 3300, 3066, 2966, 2868, 1650, 1552, 1372, 1013, 928, 727, 592.
1H NMR (CDCl3, 300 MHz): d (ppm) ¼ 1.99 (s, 1H, CH3CO), 2.96 (d,
1H, OH-3, J ¼ 3.3 Hz), 3.60 (app t, 1H, H-4, J ¼ 9.2 Hz), 3.70–3.79
(m, 1H, H-60), 3.86 (dd, 1H, J ¼ 4.6 Hz, J ¼ 9.5 Hz, H-5), 3.96 (ddd,
1H, H-3, J ¼ 3.0 Hz, J ¼ 9.2 Hz, J ¼ 9.2 Hz), 4.20–4.28 (m, 2H, H-2,
H-6), 4.50 (d, 1H, CH2Ph, Jgem ¼ 11.8 Hz), 4.78 (d, 1H, CH2Ph,
Jgem ¼ 11.8 Hz), 4.94 (d, 1H, H-1, J ¼ 3.8 Hz), 5.57 (s, 1H, CH-Ph),
5.79 (d, 1H, NH, J ¼ 8.7 Hz), 7.26–7.52 (m, 10H, H-Ar). MS (FAB),
m=z ¼ 400 [MH]þ . The spectral data are in accordance with Ref. 12.
Benzyl-4,6-O-benzylidene-2-deoxy-2-acetylamido-3-O-(oxiran-2-yl)
methyl-a-D-glucopyranoside (5)
Benzyl-4,6-O-benzylidene-2-deoxy-2-acetylamido-a-D-glucopyranoside (4)
(1.0 g, 2.51 mmol) was dried with benzene co-evaporation under reduced
pressure. The obtained white solid was mixed with catalytic amounts of
tetrabutylammonium iodide and suspended in 0.5 g of 15-crown-5 and
30 ml of dry dioxane. The suspension was flushed with argon, and then
480 mg (20 mmol) of sodium hydride was added and left to react for
0.5 h. When the evolution of hydrogen stopped, 2.31 g (25 mmol) of epi-
chlorohydrin was added to the reaction mixture and stirred at ambient
temperature under argon. After 30 h, TLC analysis indicated the reaction
was complete, and the reaction mixture was evaporated to dryness under

Analogs of N-Ac-Muramyl-L-alanine
3057
reduced pressure. The white solid was suspended in dichloromethane
(30 ml) and cooled on an ice-water bath. Saturated sodium hydrogencar-
bonate solution (20 ml) was added slowly to quench the unreacted
sodium hydride. The water phase was extracted with 3 ꢂ 30 ml of dichlor-
omethane. The organic phase was washed with brine and dried over
sodium sulfate. The crude product was recrystallized from ethanol to
yield a colorless solid: 931 mg, 81%, two diastereoisomersꢃ, mp 220–
ꢀ
224 and 228–232ꢃ C. IR (KBr, cmꢁ1): 3298, 2859, 1652, 1557, 1373,
1
726, 696. H NMR (CDCl3, 300 MHz): d (ppm) ¼ 1.98 (2.00)ꢃ (s þ s,
3H, CH3CO), 2.54 (2.72)ꢃ (dd þ dd, 1H, CH2a, J ¼ 2.7 Hz, J ¼ 5.1 Hz,
(J ¼ 4.2 Hz, J ¼ 5.2 Hz)ꢃ), 2.76–2.80 (m, 1H, CH2b), 3.05–3.12 (m, 1H,
CH2CH), 3.34 (4.21)ꢃ (dd þ dd, 1H, CH2c, J ¼ 7.3 Hz, J ¼ 12.4 Hz),
3.61–3.98 (m, 4H, H-4, H-5, H-6’, H-3), 3.92 (4.06)ꢃ (dd þ dd, 1H,
CH2d, J ¼ 2.5 Hz, J ¼ 13.2 Hz), 4.10–4.31 (m, 2H, H-2, H-6), 4.50
(4.72)ꢃ (dd þ dd, 2H, CH2Ph, Jgem ¼ 11.8 Hz, Dt ¼ 74 Hz), 4.98 (5.06)ꢃ
(d þ d, 1H, H-1, J ¼ 3.8 Hz), 5.57 (s, 1H, CHPh), 5.89 (6.09)ꢃ (d þ d,
1H, NH, J ¼ 8.3 Hz), 7.36–7.48 (m, 10H, Ph-H). 13C NMR (CDCl3,
75 MHz): d (ppm) ¼ 23.6, 44.1, 44.9, 51.7, 52.1, 53.1, 53.7, 63.4, 69.3,
70.2 70.4, 70.6, 74.4, 78.6, 82.8, 83.1, 97.9, 98.1, 101.7, 126.3, 126.4,
128.5, 128.6, 128.7, 128.9, 129.0, 129.4, 171.4. MS (FAB), m=z ¼ 456
[MH]þ . Microanalysis calcd. for C25H29N1O7: (%): C, 65.92; H, 6.42;
N, 3.07. Found: C, 66.01; H, 6.67; N, 2.84.
General Procedure and Characterization of Products 6a–e
Entries 6a–d: The carboxy protected amino acid salt (2.0 mmol) was sus-
pended in 70 mL of dichloromethane and washed with 0.5 M sodium
hydroxide solution (3 ꢂ 10 mL), water (5 mL), and brine (2 ꢂ 10 mL).
The organic phase was dried with sodium sulphate and evaporated under
reduced pressure and temperature not exceeding 30 ꢀC. A colourless oil
was obtained and dried in vacuo.
Entry 6e: The tert-butyl ester of L-alanine was prepared according to
known procedures.[14]
The free amino acid ester (0.66 mmol), epoxide 5 (0.55 mmol), and
calcium triflate (0.14 mmol) were suspended in 6 mL of anhydrous diox-
ane. The reaction mixture was heated and stirred for 20 min at 120 ꢀC and
2–3 bar in a sealed 10 ml reactor under monomode microwave irradiation
of 50 W. After cooling with compressed air to ambient temperature, the
solvent was evaporated under reduced pressure. The colorless crude
products 6a–e were purified with flash chromatography using dichloro-
methane–methanol mixtures. When necessary, the chromatographic
separation was improved using gradient elution.

ˇ ˇ
A. Babic and S. Pecar
3058
(6a) Benzyl 2-((R,S)-3-(1-O-benzyl-4,6-O-benzylidene-2-deoxy-2-acetyla-
mido-3-a-D-glucopyranosidyloxy)-2-hydroxypropylamino)acetate
Yield: 57%, two diastereoisomersꢃ, mp 180–186 ꢀC. IR (KBr, cmꢁ1):
3437, 2925, 1660, 1449, 1377, 1089. 1H NMR (CDCl3, 300 MHz): d
(ppm) ¼ 1.95 (s, 3H, CH3CO), 2.55 (2.62)ꢃ (dd þ dd, 1H, CH2aNH,
Jgem ¼ 122 Hz, J ¼ 7.3 Hz), 2.63–2.69 (m, 1H, CH2bNH), 3.37 (3.39)ꢃ
(s þ s, 2H, CH2COO), 3.48 (dd, 1H, CH2cO, J ¼ 7.1, 10.1 Hz), 3.60–
3.91 (m, 6H, CHOH, CH2dO, H-3, H-4, H-5, H-60), 4.18–4.25 (m, 2H,
H-2, H-6), 4.51 (4.71)ꢃ (d þ d, 1H, OCH2Ph, Jgem ¼ 11.8 Hz), 4.97
(4.98)ꢃ (d þ d, 1H, H-1, J ¼ 3.9 Hz), 5.14 (s, 2H, COOCH2Ph), 5.54
(s, 1H, CHPh), 6.05 (6.21)ꢃ (d þ d, 1H, NHCO, J ¼ 8.6 Hz), 7.26–7.48
(m, 15H, PhH). 13C NMR (CDCl3, 75 MHz): d (ppm) ¼ 23.70, 51.27,
53.49, 63.49, 66.98, 69.31, 69.95, 70.52, 77.64, 79.05, 82.56, 98.03,
101.83, 126.39–129.45, 135.94, 137.30, 137.59, 170.71, 172.74. LRMS
(FAB), m=z ¼ 621 [MH]þ. HRMS (ESI), calcd. for C34H41N2O9 m=z:
621.2812 [MH]þ, found: 621.2823. Microanalysis calcd. for
C34H40N2O9 ꢂ 1.5H2O (%): C, 63.98; H, 6.63; N, 4.39. Found: C,
63.54; H, 6.79; N, 4.77.
(6b) (S)-Benzyl-2-((R,S)-3-(1-O-benzyl-4,6-O-benzylidene-2-
deoxy-2-acetylamido-3-a-D-glucopyranosidyloxy)-2-hydroxypropylamino)-
propanoate
Yield: 60%, two diastereoisomersꢃ, mp 198–205 ꢀC. IR (KBr, cmꢁ1):
3294, 2924, 2855, 1741, 1658, 1551, 1452, 1376 1090, 734,
896. 1H NMR (CDCl3, 300 MHz): d (ppm) ¼ 1.26 (d, 3H, CH3,
J ¼ 6.9 Hz), 1.95 (s, 3H, CH3CO), 2.36 (2.65)ꢃ (dd þ dd, 1H, CH2aNH,
Jgem ¼ 12 Hz, J ¼ 8.1 Hz), 2.53 (2.70)ꢃ (dd þ dd, 1H, CH2bNH,
Jgem ¼ 12 Hz, J ¼ 4.2 Hz), 3.27–3.36 (m, 1H, CHCOO), 3.46 (dd, 1H,
CH2cO, J ¼ 8.3, 11.7 Hz), 3.58–3.92 (m, 6H, CHOH, CH2dO, H-3,
H-4, H-5, H-6’), 4.16–4.28 (m, 2H, H-2, H-6), 4.48 (4.72)ꢃ (d þ d,
1H, OCH2Ph, Jgem ¼ 11.9 Hz), 4.97 (5.01)ꢃ (d þ d, 1H, H-1, J ¼ 3.9
Hz), 5.13 (5.14)ꢃ (s þ s, 2H, COOCH2Ph), 5.53 (s, 1H, CHPh), 6.04
(6.31)ꢃ (d þ d, 1H, NHCO, J ¼ 8.4 Hz), 7.27–7.50 (m, 15H, PhH). 13C
NMR (CDCl3, 75 MHz): d (ppm) ¼ 19.39, 23.66, 49.95, 53.42, 57.02,
63.53, 66.90, 69.33, 69.83, 70.52, 77.68, 78.76, 82.62, 98.04, 101.79,
126.41–129.43, 136.20, 137.44, 137.59, 170.72, 175.66. MS (FAB),
m=z ¼ 635 [MH]þ. HRMS (ESI), calcd. for C35H43N2O9 m=z:
635.2969 [MH]þ, found: 635.2963. Microanalysis calcd. for
C35H42N2O9 ꢂ H2O(%): C, 64.44; H, 6.80; N, 4.29. Found: C, 64.73;
H, 6.78; N, 3.95.

Analogs of N-Ac-Muramyl-L-alanine
3059
(6c) (S)-Benzyl-2-((R,S)-3-(1-O-benzyl-4,6-O-benzylidene-2-
deoxy-2-acetylamido-3-a-D-glucopyranosidyloxy)-2-hydroxypropylamino)-
3-methylbutanoate
Yield: 58%, two diastereoisomersꢃ, mp 160–169 ꢀC. IR (KBr, cmꢁ1):
1
3294, 2926, 2856, 1731, 1656, 1551, 1450, 1375, 1087, 696. H NMR
(CDCl3, 300 MHz): d (ppm) ¼ 0.88–0.92 (m, 7H, (CH3)2CH), 1.96 (s,
3H, CH3CO), 1.85–2.05 [m, 1H, (CH3)2CH], 2.36 (2.65)ꢃ (dd þ dd, 1H,
CH2aNH, Jgem ¼ 12 Hz, J ¼ 8.1 Hz), 2.51 (2.70)ꢃ (dd þ dd, 1H,
CH2bNH, Jgem ¼ 12 Hz, J ¼ 4.2 Hz), 3.06 (m, 1H, CHCOO), 3.49 (dd,
1H, CH2cO, J ¼ 8.3, 11.7 Hz), 3.47–3.92 (m, 6H, CHOH, CH2dO, H-3,
H-4, H-5, H-60), 4.16–4.28 (m, 2H, H-2, H-6), 4.49 (4.72)ꢃ (d þ d, 1H,
OCH2Ph, Jgem ¼ 11.9 Hz), 4.97 (5.01)ꢃ (d þ d, 1H, H-1, J ¼ 3.9 Hz),
5.13 (5.14)ꢃ (s þ s, 2H, COOCH2Ph), 5.54 (5.55)ꢃ (s þ s, 1H, CHPh),
6.05 (6.40)ꢃ (d þ d, 1H, NHCO, J ¼ 8.4 Hz), 7.27–7.50 (m. 15H, PhH).
13C NMR (CDCl3, 75 MHz): d (ppm) ¼ 18.74, 19.73, 23.64, 31.93,
50.71, 53.48, 63.51, 66.79, 67.51, 68.21, 69.33, 70.52, 77.63, 78.74,
82.70, 98.01, 101.76, 126.38–129.37, 136.09, 137.36, 137.70, 170.60,
175.09. MS (FAB), m=z ¼ 663 [MH]þ. HRMS (ESI), calcd. for
C37H47N2O9 m=z: 663.3282 [MH]þ; found: 663.3276. Microanalysis
calcd. for C37H46N2O9 ꢂ 1/2H2O (%): C, 66.19; H, 7.01; N, 4.17. Found:
C, 65.94; H, 6.85; N, 3.98.
(6d) (S)-Benzyl-2-((R,S)-3-(1-O-benzyl-4,6-O-benzylidene-2-deoxy-
2-acetylamido-3-a-D-glucopyranosidyloxy)-2-hydroxypropylamino)-4-
methylpentanoate
Yield: 57%, two diastereoisomersꢃ, mp : 169–175 ꢀC. IR (KBr, cmꢁ1):
3289, 2925, 1737, 1654, 1551, 1454, 1375, 1089, 734, 696. 1H NMR
(CDCl3, 300 MHz): d (ppm) ¼ 0.86 (0.90)ꢃ [d þ d, 6H, (CH3)2, J ¼ 8.1
Hz], 1.40–1.50 [m, 2H, (CH3)2CHCH2], 1.60–1.75 [m, 1H, (CH3)2CH],
1.97 (s, 3H, CH3CO), 2.28 (2.60)ꢃ (dd þ dd, 1H, CH2aNH, Jgem ¼ 12
Hz, J ¼ 8.1 Hz), 2.48 (2.72)ꢃ (dd þ dd, 1H, CH2bNH, Jgem ¼ 12 Hz,
J ¼ 4.2 Hz), 3.26 (q, 1H, CHCOO, J ¼ 7.8 Hz), 3.41–3.93 (m, 5H, 7H,
CHOH, CH2cO, CH2dO, H-3, H-4, H-5, H-60), 4.15–4.28 (m, 2H, H-2,
H-6), 4.51 (4.74)ꢃ (d þ d, 1H, OCH2Ph, Jgem ¼ 11.9 Hz), 5,00 (5.06)ꢃ
(d þ d, 1H, H-1, J ¼ 3.7 Hz), 5.15 (s, 2H, COOCH2Ph), 5.57 (s, 1H,
CHPh), 6.00 (6.31)ꢃ (d þ d, 1H, NHCO, J ¼ 7.8 Hz), 7.25–7.53 (m.
15H, PhH). 13C NMR (CDCl3, 75 MHz): d (ppm) ¼ 22.49, 22.72,
23.69, 24.82, 41.19, 49.83, 53.70, 59.62, 63.52, 66.35, 66.50, 68.82,
70.53, 77.44, 78.75, 82.68, 98.02, 101.77, 126.40–129.02, 137.02, 137.22,
170.25, 175.65. MS (FAB), m=z ¼ 677 [MH]þ. HRMS (ESI), calcd. for

ˇ ˇ
A. Babic and S. Pecar
3060
C38H49N2O9 m=z: 677.3438 [MH]þ; found: 677.3448. Microanalysis
calcd. for C38H48N2O9 (%): C, 67.44; H, 7.15; N, 4.14. Found: C,
67.13; H, 7.31; N, 4.19.
(6e) (S)-tert-Butyl-2-((R,S)-3-(1-O-benzyl-4,6-O-benzylidene-2-
deoxy-2-acetylamido-3-a-D-glucopyranosidyloxy)-2-hydroxypropylamino)-
propanoate
Yield: 45%, two diastereoisomersꢃ, mp 167–173 ꢀC. IR (KBr, cmꢁ1):
3423, 1639, 1372, 1090, 694. 1H NMR (CDCl3, 300 MHz):
d
(ppm) ¼ 1.26 (d, 3H, CH3, J ¼ 6.9 Hz), 1.47 (s, 9H, C(CH3)3), 1.99 [s,
3H, CH3CO], 2.36 (2.65)ꢃ (dd þ dd, 1H, CH2aNH, Jgem ¼ 12 Hz,
J ¼ 8.1 Hz), 2.54 (2.70)ꢃ (dd þ dd, 1H, CH2bNH, Jgem ¼ 12 Hz, J ¼ 4.2
Hz), 3.27–3.36 (m, 1H, CHCOO), 3.46 (dd, 1H, CH2cO, J ¼ 7.8,
11.5 Hz), 3.65–3.96 (m, 6H, CHOH, CH2dO, H-3, H-4, H-5, H-60),
4.16–4.27 (m, 2H, H-2, H-6), 4.51 (4.74)ꢃ (d þ d, 1H, OCH2Ph,
Jgem ¼ 11.9 Hz), 4.99 (5.03)ꢃ (d þ d, 1H, H-1, J ¼ 3.9 Hz), 5.59 (s, 1H,
CHPh), 6.02 (6.35)ꢃ (d þ d, 1H, NHCO, J ¼ 8.4 Hz), 7.27–7.50 (m.
10H, PhH). 13C NMR (CDCl3, 75 MHz): d (ppm) ¼ 19.50, 23.63,
28.46, 53.78, 57.92, 63.52, 67.48, 69.32, 70.51, 77.60, 78.70, 81.39,
82.68, 98.00, 101.77, 126.39–129.39, 137.35, 137.59, 170.70, 175.18. MS
(FAB), m=z ¼ 601 [MH]þ. HRMS (FAB), calcd. for C32H45N2O9 m=z:
601.31350 [MH]þ; found: 601.31251. Microanalysis calcd. for
C32H44N2O9 (%): C, 63.98, H, 7.38; N, 4.66. Found: C, 63.44; H, 7.32;
N, 4.66.
ACKNOWLEDGMENT
This work was supported by the European Union FP6 Integrated Project
EUR-INTAFAR (Project No. LSHM-CT-2004–512138) under the
theme Life Sciences, Genomics, and Biotechnology for Health and by
the Ministry of Higher Education, Science, and Technology of the
Republic of Slovenia. The authors thank Dr. Roger Pain for critical read-
ing of the manuscript.
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