Synthesis and evaluation of paromomycin derivatives modified at C(4′)

Article abstract of DOI:10.1002/hlca.200890167

The 2-amino-2-deoxy-α-D-glucopyranosyl moiety (ring I) of paromomycin was replaced by a 2,4-diamino-2,4-dideoxy-α-D-glucopyranosyl, 2,4-diamino-2,4-dideoxy-α-D-galactopyranosyl, 2-amino-2-deoxy-α-D- galactopyranosyl, or 3,4,5-trideoxy-4-aza-α-D-erythro-heptoseptanosyl moiety to investigate the effect of the substituent at C(4′) on the interaction with ribosomal RNA. The triflate 6 was prepared from the key intermediate pentaazido 3′,6′-dibenzyl ether 5, and the hexosulose 10 was obtained by oxidation of 5 with Dess - Martin's periodinane. Stereoselective reduction of 10 with NaBH₄ gave the alcohol 11 that was transformed into the triflate 12. The epimeric hexaazides 7 and 13 were obtained by treating the triflates 6 and 12, respectively, with tetrabutylammonium azide. Periodate cleavage of glycol 2 yielded the dialdehyde 24 that was reductively aminated with aniline and benzylamine to give the 3,4,5-trideoxy-4-aza-α-D-erythro-heptoseptanosides 25 and 26, respectively. Standard azide reduction and debenzylation yielded 9 (2,4-diamino-2,4-dideoxy-α-D-galactopyranosyl ring I), 13 (2-amino-2-deoxy-α-D-galactopyranosyl ring I), 17 (2,4-diamino-2,4- dideoxy-α-D-glucopyranosyl ring I), and 27 and 28 (3,4,5-trideoxy-4-aza- α-D-erythro-heptoseptanosyl ring I). The derivatives 9 and 13 possessing a D-galacto-configured ring I were less active than the corresponding D-gluco-analogues 17 and paromomycin (1), respectively. The C(4′)- aminodeoxy derivative 17 (D-gluco ring I) and the known 4′- deoxyparomomycin (23), prepared by a new route, displayed slightly lower antibacterial activities than paromomycin (1). Cell-wall permeability is not responsible for the unexpectedly low activity for 17, as shown by cell-free translation assays. The results evidence that the orientation of the substituent at C(4′) is more important than its nature for drug binding and activity.

Full text of DOI:10.1002/hlca.200890167

Helvetica Chimica Acta – Vol. 91 (2008)
1533
Synthesis and Evaluation of Paromomycin Derivatives Modified
at C(4’)
by Rashmi Pathaka), De´borah Perez-Fernandeza), Rahul Nandurdikara), Sarath K. Kalapalab),
Erik C. Bçttgerb), and Andrea Vasella*a)
a) Laboratorium für Organische Chemie, ETH Zürich, Wolfgang-Pauli-Strasse 10, CH-8093 Zürich
b) Institut für Medizinische Mikrobiologie, Universität Zürich, Gloriastrasse 30/32, CH-8006 Zürich
The 2-amino-2-deoxy-a-d-glucopyranosyl moiety (ring I) of paromomycin was replaced by a 2,4-
diamino-2,4-dideoxy-a-d-glucopyranosyl, 2,4-diamino-2,4-dideoxy-a-d-galactopyranosyl, 2-amino-2-de-
oxy-a-d-galactopyranosyl, or 3,4,5-trideoxy-4-aza-a-d-erythro-heptoseptanosyl moiety to investigate the
effect of the substituent at C(4’) on the interaction with ribosomal RNA. The triflate 6 was prepared from
the key intermediate pentaazido 3’,6’-dibenzyl ether 5, and the hexosulose 10 was obtained by oxidation
of 5 with Dess – Martinꢀs periodinane. Stereoselective reduction of 10 with NaBH4 gave the alcohol 11
that was transformed into the triflate 12. The epimeric hexaazides 7 and 13 were obtained by treating the
triflates 6 and 12, respectively, with tetrabutylammonium azide. Periodate cleavage of glycol 2 yielded the
dialdehyde 24 that was reductively aminated with aniline and benzylamine to give the 3,4,5-trideoxy-4-
aza-a-d-erythro-heptoseptanosides 25 and 26, respectively. Standard azide reduction and debenzylation
yielded 9 (2,4-diamino-2,4-dideoxy-a-d-galactopyranosyl ring I), 13 (2-amino-2-deoxy-a-d-galactopy-
ranosyl ring I), 17 (2,4-diamino-2,4-dideoxy-a-d-glucopyranosyl ring I), and 27 and 28 (3,4,5-trideoxy-4-
aza-a-d-erythro-heptoseptanosyl ring I). The derivatives 9 and 13 possessing a d-galacto-configured ring
I were less active than the corresponding d-gluco-analogues 17 and paromomycin (1), respectively. The
C(4’)-aminodeoxy derivative 17 (d-gluco ring I) and the known 4’-deoxyparomomycin (23), prepared by
a new route, displayed slightly lower antibacterial activities than paromomycin (1). Cell-wall
permeability is not responsible for the unexpectedly low activity for 17, as shown by cell-free translation
assays. The results evidence that the orientation of the substituent at C(4’) is more important than its
nature for drug binding and activity.
Introduction. – The crystal structures of paromomycin [1][2] and of a few other
aminoglycoside antibiotics in complex with the decoding A site of bacterial 16SrRNA
show a very similar relative orientation of rings I and II that is responsible for the key
interactions that result in drug binding [3 – 10]; for paromomycin, the interaction of
ring I appears to be most important. The base at position 1408 of 16SrRNA is a critical
determinant of aminoglycoside selectivity. Drug-susceptible bacterial ribosomes are
characterised by an adenine, while drug resistance of eukaryotic cytoplasmic ribosomes
is conferred by a guanine at the homologous position [11]. Considering that
modifications of ring I may selectively affect the interaction with the bacterial (in
contradistinction to the eukaryotic) decoding site, we had replaced HOꢀC(6’) of
paromomycin with either H or F [12]. However, these modifications reduced the
activity both against M. smegmatis wild-type ribosomes characterized by A1408 and
against A1408G (eukaryotic type) mutants. The 3’,6’-anhydro derivative, where ring I
adopts an inverted chair conformation, lost any activity.
ꢁ 2008 Verlag Helvetica Chimica Acta AG, Zürich

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Helvetica Chimica Acta – Vol. 91 (2008)
It appeared of interest to investigate the effect on the biological activity of the
somewhat less incisive inversion of the configuration of C(4’) and of the introduction of
other substituents. We planned to prepare galacto-configured paromomycin analogues,
and to replace the C(4’)ꢀOH by a C(4’)ꢀNH2 group. The crystal structure of
paromomycin in complex with the bacterial 16SrRNA decoding site suggests that
C(4’)ꢀOH interacts with the phosphate group of A1493, an interaction that might be
strengthened by replacing the OH by a ammonium group. For the sake of a better
comparison, we also wished to evaluate the activity and selectivity in our test system of
the known 4’-deoxyparomomycin (23) [13], with the intention of preparing this
compound by a different synthetic route. Finally, we planned to also synthesize the
(3’,4’-imido)-heptoseptanosyl 4’-azaparomomycin derivatives 27 and 28, this time
opting for a novel, if again more incisive, modification of ring I.
Results and Discussion. – The known pentaazido-paromomycin 2 [12] was prepared
from paromomycin sulfate (1) by diazo transfer [14 – 16] from TfN3 in the presence of
CuSO4 · 5 H2O and t-BuOK (Scheme 1). Benzylidenation of 2 gave 75% of 3 [12] that
was O-benzylated with BnBr in the presence of NaH in THF to yield 55% of the
hexabenzyl ether 4. Regioselective reductive cleavage of the 1,3-dioxane ring of 4 with
NaCNBH3 in the presence of 0.7n HCl [17][18] gave the 6’-O-benzyl ether 5
(93%). This common intermediate was transformed into the triflate 6 (62%) and
further, by treatment with Bu4NN3 [19], to the galacto-configured hexaazide 7 (79%).
Staudinger reduction of 7 and hydrolytic workup gave the hexaamine 8 (71%).
Hydrogenolysis of the BnO groups in the presence of Pd(OH)2 in AcOH/H2O 2 :3
yielded the 4’-epi-4’-amino-4’-deoxyparomomycin hexaacetate 9 (88%). Remarkably,
replacing AcOH/H2O 2 :3 by AcOH/H2O/D2O 2 :2 :1 improved this hydrogenoly-
sis.
Oxidation of the alcohol 5 with Dess – Martinꢀs periodinane gave the hexos-4-
uloside 10 (81%; Scheme 2) that was reduced with NaBH4 to the pentaazide 11 (77%)
possessing a galacto-configured ring I. Staudinger reduction and hydrolysis of 11 to the
pentaamine 12 (65%), followed by hydrogenolysis, gave the 4’-epi-paromomycin
pentaacetate trihydrate 13 (82%).
To prepare the 4’-amino-4’-deoxyparomomycin analogue 17, we transformed the
pentaazide 11 into the triflate 14 (75%; Scheme 3). Displacement of the TfO by an N3
group yielded the hexaazide 15 (63%), possessing a gluco-configured ring I, that was
transformed via 16 into the hexaammonium acetate 17 (70%), similarly as described
above for the preparation of 9 and 13.
The synthesis of 4’-deoxyparomomycin (23) was initiated by cleaving the dioxane
ring of 4 with TsOH · H2O in CH2Cl2/MeOH 4 :1 to give diol 18 (42%, 59% conversion;
Scheme 4). Staudinger reaction and hydrolysis, followed by (benzyloxy)carbonylation of
the amino groups led to the carbamate 19 (80% from 18) that was transformed into the
6’-O-pivaloyl ester 20 (83%). The xanthate 21 was obtained in good yield from 20, but
standard Barton–McCombie deoxygenation conditions (Bu3SnH and AIBN in boiling
toluene [20]) did not transform 21 to the desired deoxy derivative 22. The deoxy
derivative was, however, obtained in 90% yield by treating 21 with H3PO2, AIBN, and
Et3N in 1,4-dioxane [21]. Reductive deprotection and aminolysis of 22 by treatment with
Na in liquid NH3 yielded 68% of 4’-deoxyparomomycin (23).

Helvetica Chimica Acta – Vol. 91 (2008)
1535
Scheme 1
a) TfN3, t-BuOK, t-BuOH/H2O 2 :1, 268; 49%. b) PhCHO, HCO2H, ꢀ 5 to 08; 75%. c) NaH, BnBr,
Bu4NI, THF, 0 to 268; 55%. d) NaCNBH3, 0.7m HCl in Et2O, 4-Š mol. sieves, THF, 08; 93%. e) Tf2O,
pyridine, CH2Cl2, ꢀ 108; 62%. f) Bu4NN3, MeCN, 268; 79%. g) 0.1m aq. NaOH, 1m PMe3 in THF, THF,
508; 71%. h) H2, Pd(OH)2, AcOH/H2O/D2O 2 :2 :1, 6 bar, 268; 78%.

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Helvetica Chimica Acta – Vol. 91 (2008)
Scheme 2
a) Dess – Martin periodinane, CH2Cl2, 268; 81%. b) NaBH4, THF/H2O 1:1, 268; 77%. c) 0.1m aq. NaOH,
1m PMe3 in THF, THF, 508; 65%. d) H2, Pd(OH)2, AcOH/H2O/D2O 2 :2 :1, 6 bar, 268; 82%.
Scheme 3
a) Tf2O, pyridine, CH2Cl2, ꢀ 10 to 208; 75%. b) Bu4NN3, MeCN, 268; 63%. c) 0.1m aq. NaOH, 1m PMe3
in THF, THF, 508; 80%. d) H2, Pd(OH)2, AcOH/H2O/D2O 2 :2 :1, 6 bar, 268; 88%.

Helvetica Chimica Acta – Vol. 91 (2008)
1537
Scheme 4
a) TsOH · H2O, CH2Cl2/MeOH 4 :1, r.t.; 42% yield, 59% conversion. b) 0.1m aq. NaOH, 1m PMe3 in THF,
THF, 508. c) ZCl (Z ¼ (benzyloxy)carbonyl), Na2CO3, MeOH, r.t.; 80% from 18. d) PivCl, pyridine,
CH2Cl2, r.t.; 83%. e) NaH, CS2, 1H-imidazole (cat.), MeI, THF, 08 to r.t.; 84%. f) H3PO2, 2,2’-
azobis(isobutyronitrile) (AIBN), Et3N, dioxane, reflux; 90%. g) Na, liq. NH3, THF, ꢀ 78 to r.t.; 68%.
Periodate cleavage of the pentaazide 2 was expected to proceed regioselectively,
considering the preferred 4C1 conformation of ring IV [22]. Indeed, treatment of 2 with
1.4 equiv. of H5IO6 gave the dialdehyde 24 that was reductively aminated with aniline,
or with benzylamine [23][24], to provide the 4-phenylated and 4-benzylated 3,4,5-
trideoxy-4-aza-a-d-erythro-heptoseptanosides 25 (32%; Scheme 5) and 26 (36% 2),
respectively. Staudinger reaction/hydrolysis of 25 and 26 gave the hexaamines 27 (66%)
and 28 (68%), respectively.
A comparison of the 1H-NMR spectra of the benzylated azides and the O-
benzylated or debenzylated free amines showed that the ribofuranose ring of the azides
prefers consistently if weakly the (S) conformation (J(1’’,2’’)/J(3’’,4’’) ¼ 1.05 – 1.11)
whereas the (N) conformation predominates in the free amines (J(1’’,2’’)/J(3’’,4’’) ¼

1538
Helvetica Chimica Acta – Vol. 91 (2008)
Scheme 5
a) H5IO6, 4-Š mol. sieves, THF, 08. b) NaCNBH3, RNH2, AcOH, MeOH, 268; from 2 32% for 25; 36%
for 26. c) 0.1m aq. NaOH, 1m PMe3 in THF, THF, 508; 66% for 27; 68% for 28.
0.39 – 0.52)1). These conformational preferences differ from those of the analogous O-
Ac protected derivatives [12]. The idopyranosyl ring of all compounds adopts
4
predominantly the C1 conformation (J(2’’’,3’’’) ¼ 1.8 – 3.1, J(3’’’,4’’’) ¼ 2.7 – 3.1). The
small J(3’,4’) value of 3.3 and 2.7 Hz for 7 and 11, respectively, evidences the galacto-
configuration of ring I. A comparison of the chemical shift of C(4’) in 7 (59.14 ppm) and
15 (62.23 ppm) confirms the inversion of configuration2).
Biological Studies. – The experimental model used to investigate ribosomal drug
susceptibility is a single rRNA allelic derivative of the Gram-positive eubacterium
Mycobacterium smegmatis [25]. Genetic manipulations of its single rRNA operon using
site-directed mutagenesis and RecA-mediated gene conversion result in homogeneous
1
)
The range could not be determined precisely for all compounds, but were always > 1 for the azides,
and < 1 for the amines.
2
)
The C(4’)-atom of the gluco-configured derivatives resonates at lower field than that of the
corresponding galacto-configured isomer, as evidenced by the 13C chemical shifts of the hydroxy
(71. 7 (5) vs. 66.8 ppm (11)), trifloxy (68.6 (6) vs. 67.1 ppm (14)), azido (62.2 (15) vs. 59.1 ppm (7)),
and amino derivatives (57.2 (16) vs. 52.0 ppm (8)). The axial N3 substituent of 7 leads to an upfield
shift of ca. 3 ppm for C(3’) and C(5’).

Helvetica Chimica Acta – Vol. 91 (2008)
Table 1. Ribosomal Drug Susceptibilitya)
1539
Wild type
Mutant A1408G
Mutant G1491A
Mutant G1491C
Paromomycin (1)
1
128
32
4
2
> 512
ꢁ 512
64
> 512
> 512
256
32 – 64
> 512
512
256
128
> 512
> 512
512
> 512
> 512
> 512
> 512
> 512
> 512
9
13
17
23
27
28
256
> 512
ꢁ 512
a) Ribosomal drug susceptibility is given as minimal inhibitory concentration (MIC [mg/ml]).
populations of mutant ribosomes [26 – 29]. Ribosomal drug susceptibility was assessed
by determination of minimal inhibitory concentrations (MICs) [27 – 29].
All new derivatives proved less active against M. smegmatis wild type ribosomes
(Table 1). The derivatives 9 and 13 possessing a d-galacto-configured ring I displayed
lower antibacterial activity than the corresponding d-gluco-configured analogues 17
and paromomycin (1), respectively. This is in agreement with the postulated interaction
of a C(4’)ꢀOH (and also of a C(4’)ꢀNHþ3 ) group with the phosphate moiety of A1493.
The 4’-amino-4’-deoxy derivative 17 possessing a d-gluco-configured ring I proved
slightly less active than paromomycin. This result is in contradiction to the expectations
that were based on a comparison of the assumed interaction of a C(4’)ꢀOH or a
C(4’)ꢀNHþ3 with a phosphate group of A1493. This decrease of activity could be due to
the influence of counter ions and of hydration, and/or to an impaired cell wall
permeability. Similarly to previously reported results [13], 4’-deoxyparomomycin (23)
was found to be only slightly less active than the parent antibiotic.
To study the specificity of the interaction of the new paromomycin analogues with
prokaryotic ribosomes, we also investigated the drug susceptibility of ribosomes with
one of the mutations A1408G, G1491A, or G1491C. These 16SrRNA positions are
polymorphic, characteristic for eukaryotic ribosomes, and main determinants of
aminoglycoside selectivity [11][30]. As shown in Table 1, modification at C(4’) had
little, if any, effect on drug selectivity.
To exclude the effect of cell wall permeability, the IC50 values for the derivatives 9,
13, 17, and 23 were determined in cell-free translation assays on strains of M. smegmatis,
i.e., (UUU)12-mRNA-directed phenylalanine incorporation and the synthesis of
luciferase in coupled transcription – translation reactions [30] (see Tables 2 and 3).
Higher concentrations of the derivatives 9 and 13 (galacto-configured ring I) than those
of their analogues 17 and paromomycin (1) (gluco-configured ring I), respectively,
were required to inhibit by 50% the incorporation of [14C]phenylalanine or the
synthesis of luciferase. Similarly, compared to paromomycin (1), a higher concentration
of the 4’-amino-4’-deoxy derivative 17 and of the 4’-deoxy derivative 23 was required
for in-vitro inhibition of protein synthesis. Thus, the lower antibacterial activity of these
derivatives cannot be attributed to differences in cell-wall permeability, but is a direct
reflection of their interaction with the ribosome.
The results evidence that the orientation of the substituent at C(4’) is of importance
for drug binding and activity, while the nature of this substituent appears to have little

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Helvetica Chimica Acta – Vol. 91 (2008)
Table 2. Aminoglycoside-Induced Inhibition of
Table 3. Aminoglycoside-Induced Inhibition of
(UUU)12-Directed Phenylalanine Incorporation
Luciferase Synthesis
IC50 [mm]a)
IC50 [mm]a)
Paromomycin (1)
0.5
Paromomycin (1)
0.02
1.80
0.20
0.04
0.03
9
13
17
40.0 – 47.0
6.0 – 9.0
0.5 – 1.0
9
13
17
23
a) Aminoglycoside concentrations required to
inhibit [14C]phenylalanine incorporation to 50%
(IC50).
a) Aminoglycoside concentrations required to
inhibit synthesis of active luciferase to 50%
(IC50).
effect on the activity, as no significant difference was observed for paromomycin (1),
the d-gluco-configured 4’-amino-4’-deoxy derivative 17 (both possessing a H-bond
donor substituent at C(4’)), and 4’-deoxyparomomycin (23). An H-bond donating
substituent at C(4’) does not appear to be necessary for drug binding and activity.
We thank Dr. B. Bernet for checking the manuscript.
Experimental Part
General. See [12].
1,3,2’,2’’’,6’’’-Pentaazido-6,3’,2’’,5’’,3’’’,4’’’-hexa-O-benzyl-4’,6’-O-benzylidene-1,3,2’,2’’’,6’’’-pentade-
aminoparomomycin (4). Under Ar, a soln. of 3 [12] (1.5 g, 1.8 mmol) in THF (20 ml) was treated with
NaH (795 mg, 50 – 60% suspension in oil, ca. 18.2 mmol), BnBr (2.1 ml, 16.2 mmol), Bu4NI (204 mg,
0.54 mmol), stirred at 08 for 8 h and at 268 for 20 h, cooled to 08, and diluted portionwise with H2O. After
evaporation of THF, the aq. layer was extracted with AcOEt (5 Â 30 ml). The combined org. layers were
washed with brine, dried (MgSO4), filtered, and evaporated. FC (AcOEt/cyclohexane 3 :17) gave 4
(1.3 g, 55%). White solid. Rf (AcOEt/cyclohexane 1:3) 0.55. M.p. 62.2 – 65.78. [a]2D5 ¼ þ71.0 (c ¼ 0.14,
CHCl3). IR (ATR): 3091w, 3058w, 3032w, 2933w, 2867w, 2099s, 1607w, 1585w, 1496m, 1454m, 1368m,
1
1330m, 1261m, 1125m, 1088s, 1069s, 1027s, 999s. H-NMR (CDCl3, 400 MHz; assignments based on a
DQFCOSYand a HSQC spectrum): see Table 4; additionally, 7.51 – 7.12 (m, 35 arom. H); 5.50 (s, PhCH);
4.95 (d, J ¼ 10.7), 4.90 (d, J ¼ 11.3), 4.77 (d, J ¼ 11.3), 4.72 (d, J ¼ 10.7), 4.62 (d, J ¼ 12.0), 4.59 (d, J ¼ 11.2,
2 H), 4.53 (d, J ¼ 12.8), 4.46 (d, J ¼ 11.8), 4.44 (d, J ¼ 11.8), 4.41 (d, J ¼ 12.0), 4.26 (d, J ¼ 12.0)
(6 PhCH2). 13C-NMR (CDCl3, 100 MHz; assignments based on a HSQC spectrum): see Table 5;
additionally, 138.36, 138.10, 137.90, 137.60, 137.43, 137.04, 136.96 (7s, 7 arom. C); 128.69 – 126.11 (several
d, 35 arom. CH); 101.39 (d, PhCH); 75.03, 74.90, 73.26, 73.20, 72.42, 71.77 (6t, 6 PhCH2). HR-ESI-MS:
1256.5186 (21, [M ꢀ 5 N2 þ Na]þ, C72H75N5NaO1þ4 ; calc. 1256.5208), 689.2823 (100, [M ꢀ C39H41N6O7 ꢀ
2 H þ Na]þ, C33H32N9NaO7þ ; calc. 689.2322). Anal. calc. for C72H75N15O14 · H2O (1392.47): C 62.10, H
5.57, N 15.09; found: C 61.97, H 5.32, N 14.59.
1,3,2’,2’’’,6’’’-Pentaazido-6,3’,6’,2’’,5’’,3’’’,4’’’-hepta-O-benzyl-1,3,2’,2’’’,6’’’-pentadeaminoparomomycin
(5). Under N2, a soln. of 4 (2.25 g, 1.64 mmol) in THF (50 ml) was treated with 4-Š molecular sieves,
stirred for 1 h, cooled to 08, treated with NaCNBH3 (1.79 g, 28.48 mmol), and then dropwise with 0.7m
HCl in Et2O (45 ml, 31.5 mmol; methyl orange was added to indicate the pH), and stirred for 4 h. After
neutralization with NaHCO3 soln. and evaporation of THF, the aq. layer was extracted with AcOEt (3 Â
30 ml). The combined org. layers were washed with brine, dried (MgSO4), filtered, and evaporated. FC
(AcOEt/cyclohexane 9 :41) gave 5 (2.1 g, 93%). White solid. Rf (AcOEt/cyclohexane 1:4) 0.29. M.p.
1358. [a]2D5 ¼ þ71.4 (c ¼ 0.52, CHCl3). IR (CHCl3): 3574w, 3067w, 3007w, 2926w, 2870w, 2106s, 1603w,
1497w, 1454m, 1364w, 1330w, 1262m, 1207m, 1123m, 1074m, 1042m, 1028m, 914w. 1H-NMR (CDCl3,

Helvetica Chimica Acta – Vol. 91 (2008)
1541
Table 4. 1H-NMR Chemical Shifts [ppm] and Coupling Constants [Hz] of the Azido Derivatives 4 – 7, 10, 11, 14,
and 15 in CDCl3
4a)
5
6
7
10
6.36
3.64 – 3.57 3.52
11
6.18
3.42
4.01
14
15
6.18
3.02
3.93
3.54
HꢀC(1’)
HꢀC(2’)
HꢀC(3’)
HꢀC(4’)
HꢀC(5’)
6.10
6.16
3.00
6.17
2.85
6.14
6.23
3.42
4.07
5.51
3.08
4.10
3.44
4.08
3.98 – 3.87 4.04
3.53 – 3.39 4.77
4.15
4.08
4.47
–
4.14b)
3.78 – 3.72
4.11
3.92 – 3.88 4.29 – 4.27 4.84
4.28 – 4.22 3.99
HaꢀC(6’) 4.33 – 4.28 3.81
3.79 – 3.75 3.77 – 3.73 4.02 – 3.97 3.78 – 3.72
3.77 – 3.67 3.79 – 3.75 3.77 – 3.73 3.85 – 3.71 3.78 – 3.72
3.76 – 3.64 3.80 – 3.74
3.76 – 3.64 3.80 – 3.74
3.63 – 3.53 3.44
HbꢀC(6’) 3.65
HꢀC(1)
3.47 – 3.38 3.53 – 3.39 3.50 – 3.38 3.44 3.53 – 3.43 3.48 – 3.35
HaxꢀC(2) 1.41
HeqꢀC(2) 2.24
1.35
2.23
1.36
2.24
1.33
2.21
1.39
2.25
1.31
2.19
1.35
2.22
1.33
2.22
HꢀC(3)
HꢀC(4)
HꢀC(5)
HꢀC(6)
HꢀC(1’’)
HꢀC(2’’)
HꢀC(3’’)
HꢀC(4’’)
3.47 – 3.38 3.53 – 3.39 3.50 – 3.38 3.40
3.53 – 3.43 3.48 – 3.35
3.64 – 3.57 3.85 – 3.71 3.69
3.45 – 3.35 3.41
3.67
3.94
3.29
5.66
3.96
3.77 – 3.67 3.63
3.98 – 3.87 3.91
3.25
5.67
3.58
3.90
3.21
5.63
3.94
3.63
3.91
3.24
5.64
3.91
3.21
5.63
3.99
3.29
5.71
3.92
3.21
5.64
3.23
5.61
3.98 – 3.87 3.92 – 3.88 3.92
4.02 – 3.97 3.94
4.33 – 4.28 4.23 – 4.21
4.27 – 4.23
4.33 – 4.28 4.30 – 4.24 4.27 – 4.26 4.23
4.33 – 4.28 4.30 – 4.24 4.27 – 4.26 4.29 – 4.27 4.33 – 4.28 4.23 – 4.21
3.77 – 3.67 3.71 – 3.62 3.64 – 3.57 3.85 – 3.71 3.78 – 3.72
4.28 – 4.22 4.44 – 4.42
4.28 – 4.22 4.27 – 4.23
3.76 – 3.64 3.80 – 3.74
3.76 – 3.64 3.80 – 3.74
HaꢀC(5’’) 3.82
HbꢀC(5’’) 3.57
HꢀC(1’’’) 4.88
HꢀC(2’’’) 3.34b)
HꢀC(3’’’) 3.76
HꢀC(4’’’) 3.13b)
3.58
3.54
3.52
3.60
3.53
4.87
4.88
4.83
4.92
4.83
4.83
4.84
3.33
3.35b)
3.35b)
3.33b)
3.40b)
3.32b)
3.32b)
3.77 – 3.67 3.79 – 3.75 3.75
3.13c) 3.13b) 3.12b)
3.85 – 3.71 3.78 – 3.72
3.17b) 3.11b)
3.76 – 3.64 3.57
3.11b)
3.12b)
HꢀC(5’’’) 3.79 – 3.77 3.77 – 3.67 3.79 – 3.75 3.64 – 3.57 3.85 – 3.71 3.78 – 3.72
3.76 – 3.64 3.80 – 3.74
3.76 – 3.64 3.60
HaꢀC(6’’’) 3.65
HbꢀC(6’’’) 2.90
3.63
2.90
3.6
10.2
)
3.71 – 3.62 3.64 – 3.57 3.67
3.59
2.90
3.6
2.89
3.6
2.90
3.6
2.93
3.3
2.88
3.9
2.90
3.6
J(1’,2’)
J(2’,3’)
J(3’,4’)
J(4’,5’)
J(5’,6’a)
J(5’,6’b)
3.9
10.1
9.7
10.2
10.5
10.5
10.5
10.5
2.4
10.1
d
9.0
10.2
)
)
3.3
1.2
)
)
)
–
–
2.7
)
)
)
)
9.3
9.6
)
d
d
d
d
9.3
4.9
10.0
9.6
2.1
)
< 1.5
d
d
d
d
d
d
d
2.4
8.7
)
)
)
d
d
d
3.0
)
d
d
d
J((6’a,6’b) 10.0
10.5
12.6
4.8
)
)
J(1,2ax)
J(1,2eq)
J(1,6)
12.6
4.6
9.1
12.9
4.2
9.0
13.2
12.9
4.2
12.6
4.8
10.2
13.2
12.6
4.5
12.6
4.2
)
13.5
12.6
4.2
12.6
4.8
9.0
12.6
12.6
4.8
12.9
4.8
12.9
4.5
d
d
)
8.7
9.3
J(2ax,2eq) 13.2
13.2
12.6
4.8
13.5
12.9
4.8
12.9
12.9
4.2
J(2ax,3)
J(2eq,3)
J(3,4)
J(4,5)
J(5,6)
J(1’’,2’’)
J(2’’,3’’)
J(3’’,4’’)
J(4’’,5’’a)
J(4’’,5’’b)
12.6
4.6
8.6
8.6
8.7
5.4
9.4
)
9.0
9.3
9.3
9.0
9.0
9.3
d
9.0
9.3
9.3
8.7
8.7
9.4
9.4
5.7
)
)
)
8.7
9.3
9.3
9.0
8.7
9.3
5.7
)
)
)
5.7
5.1
2.1
)
6.0
5.7
5.1
)
)
6.0
5.7
6.0
)
)
)
d
d
d
d
d
)
)
)
)
d
d
d
d
d
d
d
)
)
)
d
d
d
d
d
d
d
1.8
3.1
)
d
d
2.4
3.0
3.6
2.7
3.3
)

1542
Helvetica Chimica Acta – Vol. 91 (2008)
Table 4 (cont.)
4a)
5
6
7
10
11
14
15
d
d
J(5’’a,5’’b)
J(1’’’,2’’’)
J(2’’’,3’’’)
J(3’’’,4’’’)
J(4’’’,5’’’)
J(5’’’,6’’’a)
J(5’’’,6’’’b)
J(6’’’a,6’’’b)
10.5
1.9
10.8
1.8
10.5
2.1
10.5
1.8
)
10.4
2.1
10.2
1.5
)
1.5
)
)
)
)
3.9
)
1.8
d
d
d
d
d
d
d
d
2.0
3.1
)
)
)
)
)
)
)
)
1.8
)
)
)
)
)
2.1
2.7
)
8.1
4.2
d
d
d
d
3.0
)
)
d
d
d
d
d
d
d
d
d
8.3
3.9
13.0
8.4
4.2
12.8
8.4
4.2
12.9
8.4
3.9
12.8
3.6
12.8
4.2
12.8
12.6
12.8
c
a) Assignments based on a DQFCOSYand a HSQC spectrum. b) w1/2 ¼ 6.0 Hz. ) w1/2 ¼ 6.9 Hz. d) Not assigned.
300 MHz): see Table 4; additionally, 7.40 – 7.19 (m, 35 arom. H); 4.97 (d, J ¼ 10.5), 4.91 (d, J ¼ 11.7), 4.77
(d, J ¼ 11.4), 4.69 (d, J ¼ 10.8), 4.66 (d, J ¼ 12.0), 4.63 (d, J ¼ 11.1), 4.59 (d, J ¼ 11.1), 4.57 (d, J ¼ 11.8),
4.54 (d, J ¼ 11.8), 4.47 (d, J ¼ 12.0), 4.44 (d, J ¼ 12.0), 4.42 (d, J ¼ 12.9), 4.32 (d, J ¼ 11.8), 4.26 (d, J ¼ 11.8)
(7 PhCH2); 2.48 (d, J ¼ 2.1, OH). 13C-NMR (CDCl3, 75 MHz): see Table 5; additionally, 138.46, 138.40,
138.23, 138.15, 137.78, 137.23, 137.17 (7s, 7 arom. C); 130.27 – 127.61 (several d, 35 arom. CH); 75.22 (2 C),
73.63 (2 C), 73.51, 73.08, 72.58 (5t, 7 PhCH2). HR-ESI-MS: 1414.5416 (26, [M þ K]þ, C72H77KN15O1þ4
;
calc. 1414.5411), 1398.5698 (100, [M þ Na]þ, C72H77N15NaO1þ4 ; calc. 1398.5672). Anal. calc. for
C72H77N15O14 (1376.49): C 62.83, H 5.64, N 15.26; found: C 62.65, H 5.72, N 15.05.
1,3,2’,2’’’,6’’’-Pentaazido-6,3’,6’,2’’,5’’,3’’’,4’’’-hepta-O-benzyl-1,3,2’,2’’’,6’’’-pentadeamino-4’-O-[(tri-
fluoromethyl)sulfonyl]paromomycin (6). Under N2, a soln. of pyridine (0.12 ml, 1.54 mmol) and Tf2O
(0.26 ml, 1.54 mmol) in CH2Cl2 (10 ml) was stirred for 30 min at ꢀ 108, treated portionwise with a soln. of
5 (950 mg, 0.69 mmol) over 30 min, and stirred at ꢀ 108 for 3 h. After neutralization with aq. NaHCO3
and extraction with CH2Cl2 (3 Â 30 ml), the combined org. layers were washed with brine, dried
(MgSO4), filtered, and evaporated. FC (AcOEt/cyclohexane 3 :22) gave 6 (648 mg, 62%). White solid. Rf
(AcOEt/cyclohexane 1:4) 0.44. M.p. 49 – 518. [a]2D5 ¼ þ89.0 (c ¼ 0.13, CHCl3). IR (CHCl3): 2927w,
2857w, 2105s, 1602m, 1491w, 1454m, 1410m, 1357w, 1329w, 1243w, 1141m, 1079m, 1037m, 1024m, 928m.
1H-NMR (CDCl3, 300 MHz): see Table 4; additionally, 7.37 – 7.17 (m, 35 arom. H); 4.93 (d, J ¼ 10.8), 4.85
(d, J ¼ 11.7), 4.72 (d, J ¼ 10.2), 4.71 (d, J ¼ 10.5), 4.63 (d, J ¼ 11.7), 4.61 (d, J ¼ 11.4), 4.55 (d, J ¼ 12.0),
4.54 (d, J ¼ 11.1), 4.41 (d, J ꢂ 9.9), 4.41 (d, J ¼ 11.7, 3 H), 4.32 (d, J ¼ 11.7), 4.25 (d, J ¼ 12.3), (7 PhCH2).
13CçNMR (CDCl3, 75 MHz): see Table 4; additionally, 137.81, 137.67, 137.61, 137.39, 136.91, 136.84,
136.79 (7s, 7 arom. C); 128.58 – 127.48 (several d, 35 arom. CH); 118.26 (q, 1J(C,F) ¼ 316.6, CF3); 75.13,
74.90, 73.46, 73.38, 73.16, 72.40, 71.73 (7t, 7 PhCH2). 19F-NMR (CDCl3, 282 MHz): ꢀ 74.60. HR-ESI-MS:
1530.5179 (22, [M þ Na]þ, C73H76F3N15NaO16Sþ; calc. 1530.5165), 803.5333 (100, [M ꢀ C39H41N6O7 þ
H]þ, C34H36F3N9O9Sþ; calc. 803.2309). Anal. calc. for C73H76F3N15O16S(1508.54): C 58.12, H 5.08, N
13.93; found: C 58.64, H 5.09, N 12.76.
4’-epi-1,3,2’,4’,2’’’,6’’’-Hexaazido-6,3’,6’,2’’,5’’,3’’’,4’’’-hepta-O-benzyl-1,3,2’,2’’’,6’’’-pentadeamino-4’-de-
oxyparomomycin (7). Under N2, a soln. of 6 (514 mg, 0.34 mmol) in MeCN (10 ml) was treated with
Bu4NN3 (1 g, 3.52 mmol), stirred for 10 h at r.t., and evaporated. A soln. of the residue in CH2Cl2 was
washed twice with brine, dried (MgSO4), filtered, and evaporated. FC (amino-phase silica gel; AcOEt/
cyclohexane 3 :22) gave 7 (376 mg, 79%). White solid. Rf (AcOEt/cyclohexane 1:4) 0.44. M.p. 48.48.
[a]2D5 ¼ þ71.0 (c ¼ 0.18, CHCl3). IR (CHCl3): 2928m, 2852w, 2105s, 1603w, 1493w, 1454m, 1364w, 1324w,
1
1263m, 1090m, 1029m, 915w. H-NMR (CDCl3, 300 MHz): see Table 4; additionally, 7.44 – 7.13 (m, 35
arom. H); 4.94 (d, J ¼ 10.5), 4.75 (d, J ¼ 11.4), 4.70 (d, J ¼ 11.4), 4.65 (d, J ꢂ 9.6, 2 H), 4.62 (d, J ¼ 12.0,
2 H), 4.57 (d, J ¼ 11.4, 2 H), 4.55 (d, J ¼ 11.7), 4.42 (d, J ¼ 11.7), 4.41 (d, J ¼ 11.4), 4.31 (d, J ¼ 12.0), 4.25
(d, J ¼ 12.0) (7 PhCH2). 13C-NMR (CDCl3, 75 MHz): see Table 5; additionally, 138.07, 137.80 (2 C),
137.45, 137.03, 136.93, 136.86 (6s, 7 arom. C); 128.58 – 127.37 (several d, 35 arom. CH); 74.92, 73.28 (3 C),
72.28, 72.271, 71.62 (5t, 7 PhCH2). HR-MALDI-MS: 1439.5373 (10, [ M þ K]þ, C72H76KN18O1þ3 ; calc.

Helvetica Chimica Acta – Vol. 91 (2008)
Table 5. 13C-NMR Chemical Shifts [ppm] of the Azido Derivatives 4 – 7, 10, 11, 14, and 15 in CDCl3
4a)
10 11 14 15
96.50 95.62 95.97 95.68 95.99
1543
5
6
7
C(1’)
C(2’)
C(3’)
C(4’)
C(5’)
C(6’)
C(1)b)
C(2)
96.21
62.76
79.91
70.40
71.66
70.47
60.58
32.77
60.38
75.00
82.06
84.38
106.26
82.62
75.74
82.25
71.93
98.87
57.49
73.40
72.31
74.50
51.25
95.51
63.02
81.53
68.60
71.44
67.84
60.33
32.39
59.88
75.40
81.99
84.08
106.34
82.23
76.75
82.09
70.08
98.55
57.29
74.39
72.84
75.28
51.16
95.72
59.59
75.82
59.14
67.41
68.72
60.17
32.29
59.88
74.68
81.64
84.03
105.92
82.40
75.52
82.08
69.96
98.57
57.19
72.80
71.33
74.13
50.89
62.87
75.98
82.53
62.97
69.00
60.34
32.39
59.87
75.42
81.85
84.14
106.25
82.42
75.51
82.11
70.37
98.60
57.31
72.96
71.60
74.39
51.14
63.80
79.18
202.27
72.93
68.13
60.37
32.49
60.01
75.52
81.94
84.23
106.08
82.60
75.62
82.22
69.98
98.76
57.37
74.03
71.47
74.39
51.17
60.38
75.92
66.83
68.82
69.76
60.14
32.60
58.78
74.67
81.73
84.16
105.94
82.47
75.67
82.14
70.15
98.67
57.38
72.94
71.48
74.24
51.05
60.10
81.63
67.07
72.77
67.24
59.64
32.08
58.62
74.94
81.99
84.00
105.98
82.47
75.49
82.18
69.91
98.57
57.16
72.77
71.33
74.19
50.94
62.93
78.47
62.23
70.13
69.00
60.35
32.57
60.08
74.79
81.85
84.09
106.02
82.30
75.49
81.99
70.02
98.61
57.29
72.86
71.41
74.23
51.04
C(3)b)
C(4)
C(5)
C(6)
C(1’’)
C(2’’)
C(3’’)
C(4’’)
C(5’’)
C(1’’’)
C(2’’’)
C(3’’’)
C(4’’’)
C(5’’’)
C(6’’’)
a) Assignments based on a DQFCOSY and a HSQC spectrum. b) Assigments for 5 – 7, 10, 11, 14, and 15
may be interchanged.
1439.5476), 1423.5701 (100, [M þ Na]þ, C72H76N18NaO1þ3 ; calc. 1423.5737), 1373.5812 (22, [M ꢀ N2 þ
H]þ, C72H77N16Oþ13 ; calc. 1373.5856). Anal. calc. for C72H76N18O13 (1401.49): C 61.70, H 5.47, N 17.99;
found: C 61.54, H 5.48, N 17.76.
4’-epi-4’-Amino-6,3’,6’,2’’,5’’,3’’’,4’’’-hepta-O-benzyl-4’-deoxyparomomycin (8). A soln. of 7 (198 mg,
0.14 mmol) in THF (20 ml) was treated with 0.1m aq. NaOH. (4.2 ml) and 1m PMe3 in THF (2.24 ml,
2.24 mmol), stirred for 2 h at 508, and evaporated. FC (amino phase silica gel; MeOH/cyclohexane 3 :97)
gave 8 (125 mg, 71%). White solid. Rf (MeOH/CHCl3 1:19) 0.34. M.p. 45.2 – 47.28. [a]2D5 ¼ þ37.8 (c ¼ 0.37,
CHCl3). IR (ATR): 3373w, 3029w, 2912m, 2865m, 1587m, 1496m, 1453s, 1363m, 1328m, 1207m, 1072s,
1025s, 908m. 1H-NMR (CDCl3, 300 MHz): see Table 6; additionally, 7.36 – 7.18 (m, 35 arom. H); 4.95 (d,
J ¼ 11.4), 4.64 (d, J ¼ 11.7), 4.63 (d, J ¼ 11.7), 4.58 (d, J ¼ 12.3), 4.51 (d, J ꢂ 9.0, 5 H), 4.51 (d, J ¼ 11.7),
4.49 (d, J ¼ 12.6), 4.44 (d, J ¼ 12.0), 4.37 (d, J ¼ 12.0), 4.24 (d, J ¼ 11.7) (7 PhCH2); 1.50 (br. s, w1/2 ¼ 72.0,
6 NH2). 13C-NMR (CDCl3, 75 MHz): see Table 7; additionally, 138.23, 138.19, 138.00, 137.94, 137.67
(2 C), 137.14 (6s, 7 arom. C); 128.38 – 127.39 (several d, 35 arom. CH); 75.01, 73.44, 73.02 (3 C), 71.90,
71.33 (5t, 7 PhCH2). HR-ESI-MS: 1267.6584 (2, [M þ Na]þ, C72H88N6NaO1þ3 ; calc. 1267.6301); 1245.6464
(14, [M þ H]þ, C72H89N6Oþ13 ; calc. 1245.6488), 905.4517 (100, [M ꢀ C20H25N2O3 þ 2 H]þ, C52H65N4Oþ10
;
calc. 905.4701). Anal. calc. for C72H88N6O13 · 4 H2O (1317.56): C 65.63, H 7.34, N 6.38; found: C 65.96, H
7.26, N 6.72.
4’-epi-4’-Amino-4’-deoxyparomomycin Hexaacetate (9). A soln. of 8 (87 mg, 0.07 mmol) in AcOH/
H2O/D2O 2 :2 :1 (3 ml) was treated with Pd(OH)2 (65 mg) and stirred for 12 h at r.t. under 6 bar of H2.
Filtration and evaporation gave 9 (53 mg, 78%). White solid. Rf (CHCl3/MeOH/25% aq. NH3 1:4 :3)
1
0.40. M.p. 1258 (dec.). [a]2D5 ¼ þ53.1 (c ¼ 0.27, H2O). H-NMR (D2O, 300 MHz): see Table 6; addition-
ally, 1.96 (s, 6 AcO). 13C-NMR (CDCl3, 75 MHz): see Table 7; additionally, 182.19 (s, 6 C¼O); 25.77 (q,

1544
Helvetica Chimica Acta – Vol. 91 (2008)
Table 6. 1H-NMR Chemical Shifts [ppm] and Coupling Constants [Hz] of the Protected Amines 8, 12, and
16 in CDCl3, and of the Deprotected Ammonium Salts 9, 13, and 17 in D2O
8
12
16
5.42
9
13a)
17
5.67
HꢀC(1’)
HꢀC(2’)
HꢀC(3’)
HꢀC(4’)
HꢀC(5’)
HaꢀC(6’)
HbꢀC(6’)
HꢀC(1)
HaxꢀC(2)
HeqꢀC(2)
HꢀC(3)
HꢀC(4)
HꢀC(5)
HꢀC(6)
HꢀC(1’’)
HꢀC(2’’)
HꢀC(3’’)
HꢀC(4’’)
HaꢀC(5’’)
HbꢀC(5’’)
HꢀC(1’’’)
HꢀC(2’’’)
HꢀC(3’’’)
HꢀC(4’’’)
HꢀC(5’’’)
HaꢀC(6’’’)
HbꢀC(6’’’)
J(1’,2’)
J(2’,3’)
J(3’,4’)
J(4’,5’)
J(5’,6’a)
J(5’,6’b)
J((6’a,6’b)
J(1,2ax)
J(1,2eq)
J(1,6)
J(2ax,2eq)
J(2ax,3)
J(2eq,3)
J(3,4)
J(4,5)
J(5,6)
J(1’’,2’’)
J(2’’,3’’)
J(3’’,4’’)
5.45
5.41
5.78
5.73
3.04
3.17
3.95 – 3.93
3.99
4.26 – 4.18
3.73
3.73
2.78 – 2.66
1.11
1.84
2.78 – 2.66
3.37
3.51
2.90
3.69 – 3.58
2.84
4.28 – 4.21
3.42 – 3.35
3.42 – 3.35
2.79 – 2.69
1.11
1.85
2.79 – 2.69
3.04
3.49
3.36
3.46
4.36
3.56
4.08
4.02
4.09 – 4.08
3.77
3.75
3.28
1.71
2.37
3.35 – 3.32
3.86
3.84
3.64
5.37
4.37
4.53
4.19
3.91
3.78
5.29
3.58
4.22
3.81
4.30
3.42
3.36
4.1
3.34–3.17
3.56 – 3.53
2.93
3.83 – 3.63
3.83 – 3.63
3.83 – 3.63
3.34 – 3.17
1.60
3.95 – 3.92
3.20b)
4.30 – 4.22
3.72 – 3.60
3.72 – 3.60
2.78 – 2.68
1.08
1.82
2.78 – 2.68
3.37
3.55
3.04
5.37
3.95 – 3.92
4.30 – 4.22
4.30 – 4.22
3.72 – 3.60
3.51
3.55 – 3.47
4.29
3.78 – 3.65
3.78 – 3.65
3.35 – 3.22
1.78
2.38
3.35 – 3.22
3.84
4.00
3.63
5.29
4.22
2.23
3.34 – 3.17
3.83 – 3.63
3.83 – 3.63
3.56 – 3.53
5.27
4.27
4.40
4.19
3.05
5.39
5.33
3.95 – 3.93
4.26 – 4.18
4.26 – 4.18
3.63 – 3.61
3.56
3.92b)
3.97
4.44
4.28 – 4.21
3.70
3.61
4.64
2.92
3.42 – 3.35
3.19b)
3.91 – 3.84
2.76
2.51
3.3
4.12 – 4.08
3.78 – 3.65
3.78 – 3.65
5.18
3.83 – 3.63
3.83 – 3.63
5.17c)
4.56
4.61
2.94b)
3.72 – 3.60
3.20b)
3.44
2.94d)
3.63 – 3.61
3.19d)
3.63 – 3.61
2.77
3.47e)
4.12 – 4.08
3.81b)
4.22
3.46f)
4.11
3.83 – 3.63
4.10 – 4.05
2.78
2.52
3.9
10.2
)
)
)
)
3.55 – 3.47
3.25
4.2
11.4
4.5
< 0.8
2.1
3.31
3.25
3.7
)
10.2
10.2
)
)
)
12.1
4.2
)
12.9
12.1
4.2
)
)
)
)
2.52
3.9
10.5
2.4
< 1.5
5.6
g
10.5
11.1
g
9.6
9.6
)
)
)
12.9
4.2
9.6
12.9
13.2
4.2
2.8
< 1.5
4.3
g
g
g
g
g
g
g
g
5.6
4.8
3.2
g
g
g
g
)
)
)
12.0
12.5
4.1
10.5
12.6
12.6
4.3
9.3
9.3
9.1
2.5
12.6
4.2
9.0
13.2
12.6
4.2
9.3
9.0
9.0
3.6
12.9
4.2
9.3
12.9
12.9
4.2
12.9
4.2
9.6
12.9
12.9
4.2
9.3
9.3
9.6
2.1
g
g
9.3
9.3
9.3
3.9
9.3
9.6
9.3
3.6
g
g
g
g
g
g
)
)
)
3.3
)
)
)
)
5.4
4.8
6.8
3.0
4.3
5.0
5.0
)
g
g
3.9
2.4
5.4
6.4
g
g
g
g
J(4’’,5’’a)
J(4’’,5’’b)
)
)
g
3.3
5.0

Helvetica Chimica Acta – Vol. 91 (2008)
1545
Table 6 (cont.)
8
12
16
9
13a)
17
g
g
J(5’’a,5’’b)
J(1’’’,2’’’)
J(2’’’,3’’’)
J(3’’’,4’’’)
J(4’’’,5’’’)
J(5’’’,6’’’a)
J(5’’’,6’’’b)
J(6’’’a,6’’’b)
10.1
2.1
10.5
10.5
2.1
)
)
)
)
)
)
4.5
12.4
1.7
3.1
3.1
1.5
6.7
4.0
13.7
)
)
)
2.9
)
6.7
4.5
g
g
g
g
g
g
g
)
)
)
)
g
g
g
g
)
)
)
)
)
)
g
g
g
g
g
g
g
7.5
5.1
13.2
7.5
4.9
13.4
7.2
5.1
13.7
14.1
13.5
a) Assignment based on a DQFCOSY and a HSQC spectrum. b) w1/2 ¼ 6 Hz. c) w1/2 ¼ 5.5 Hz. d) w1/2
¼
f
10 Hz. e) w1/2 ¼ 4.2 Hz. ) w1/2 ¼ 6.5 Hz. g) Not assigned.
6 MeC¼O). HR-ESI-MS: 637.3024 (36, [M þ Na]þ, C23H46N6NaO1þ3 ; calc. 637.3021), 615.3204 (17, [M þ
H]þ, C23H47N6O1þ3 ; calc. 615.3201), 455.2325 (100, [M ꢀ C6H13N2O3 þ 2 H]þ, C17H35N4O1þ0 ; calc.
455.2353).
1,3,2’,2’’’,6’’’-Pentaazido-6,3’,6’,2’’,5’’,3’’’,4’’’-hepta-O-benzyl-1,3,2’,2’’’,6’’’-pentadeamino-4’-deoxy-4’-
oxoparomomycin (10). Under N2, a soln. of 5 (713 mg, 0.52 mmol) in CH2Cl2 (10 ml) was treated with a
15 wt-% soln. of Dess – Martin periodinane in CH2Cl2 (2.21 ml, 0.70 mmol), and stirred for 12 h at 268.
After neutralization with aq. NaHCO3 and extraction with CH2Cl2 (3 Â 30 ml), the combined org. layers
were washed with brine, dried (MgSO4), filtered, and evaporated. FC (AcOEt/cyclohexane 9 :41) gave
10 (576 mg, 81%). White sticky solid. Rf (AcOEt/cyclohexane 1:4) 0.29. M.p. ca. 558. [a]2D5 ¼ þ93.1 (c ¼
0.60, CHCl3). IR (ATR): 3030w, 2924w, 2864w, 2098s, 1736m, 1496m, 1453m, 1366m, 1328w, 1260m,
1208w, 1026s, 914w, 819w. 1H-NMR (CDCl3, 300 MHz): see Table 4; additionally, 7.48 – 7.19 (m, 35 arom.
H); 5.02 (d, J ¼ 10.5), 4.97 (d, J ¼ 10.8), 4.73 (d, J ¼ 10.5), 4.67 (d, J ¼ 10.8), 4.66 (s, 2 H), 4.64 (d, J ¼ 12.9,
2 H), 4.53 (d, J ¼ 11.7), 4.50 (d, J ¼ 12.0), 4.46 (d, J ¼ 12.3), 4.44 (d, J ¼ 12.0), 4.36 (d, J ¼ 12.3), 4.30 (d,
J ¼ 12.3) (7 PhCH2). 13C-NMR (CDCl3, 75 MHz): see Table 5; additionally, 138.08, 138.00, 137.85, 137.45,
136.97 (2 C), 136.90 (6s, 7 arom. C); 129.03 – 127.51 (several d, 35 arom. CH); 75.19, 73.95, 73.46 (2 C),
73.37, 72.46, 71.78 (6t, 7 PhCH2). HR-MALDI-MS: 1412.5260 (45, [ M þ K]þ, C72H75KN15O1þ4 ; calc.
1412.5255), 1396.5536 (100, [M þ Na]þ, C72H75N15NaOþ14 ; calc. 1396.5516). Anal. calc. for C72H75N15O14
(1374.46): C 62.92, H 5.50, N 15.29; found: C 62.66, H 5.59, N 15.09.
4’-epi-1,3,2’,2’’’,6’’’-Pentaazido-6,3’,6’,2’’,5’’,3’’’,4’’’-hepta-O-benzyl-1,3,2’,2’’’,6’’’-pentadeaminoparo-
momycin (11). Under N2, a soln. of 10 (376 mg, 0.27 mmol) in THF/H2O 1:1 (4 ml) was treated with
NaBH4 (33 mg, 0.87 mmol) and stirred for 3 h at 268. After neutralization with 10% aq. AcOH (0.5 ml)
and evaporation, a suspension of the residue in H2O was extracted with CH2Cl2 (3 Â 15 ml). The
combined org. layers were washed with brine, dried (MgSO4), filtered, and evaporated. FC (AcOEt/
cyclohexane 9 :41) gave 10 (290 mg, 77%). White solid. Rf (AcOEt/cyclohexane 1:3) 0.26. M.p. 49.6 –
51.68. [a]2D5 ¼ þ75.9 (c ¼ 0.30, CHCl3). IR (ATR): 3390w, 3081w, 3059w, 2928w, 2105s, 1602w, 1497w,
1454w, 1360w, 1324w, 1262m, 1118m, 1087m, 1039m, 956w, 906w. 1H-NMR (CDCl3, 300 MHz): see
Table 4; additionally, 7.38 – 7.13 (m, 35 arom. H); 4.94 (d, J ¼ 10.5), 4.70 (s, 2 H), 4.64 (d, J ¼ 10.8, 2 H),
4.61 (d, J ꢂ 9.3, 2 H), 4.56, (d, J ¼ 11.4, 2 H), 4.44 (d, J ¼ 10.2), 4.40 (d, J ¼ 10.8), 4.30 (d, J ¼ 12.0), 4.25
(d, J ¼ 11.7, 2 H) (7 PhCH2); 2.38 (br. s, OH). 13C-NMR (CDCl3, 75 MHz): see Table 5; additionally,
138.17, 138.12, 137.87, 137.49, 137.29, 136.94, 136.89 (7s, 7 arom. C); 128.62 – 127.44 (several d, 35 arom.
CH); 75.06, 73.36 (3 C), 72.41, 71.92, 71.76 (5t, 7 PhCH2). HR-MALDI-MS: 1414.5402 (15, [ M þ K]þ,
C72H77KN15Oþ14 ; calc. 1414.5411), 1398.5691 (100, [M þ Na]þ, C72H77N15NaO1þ4 ; calc. 1398.5672),
1348.5719 (23, [M ꢀ N2 þ H]þ, C72H78N13Oþ14 ; calc. 1348.5791). Anal. calc. for C72H77N15O14 · 2 H2O
(1412.50): C 61.22, H 5.78, N 14.87; found: C 60.95, H 5.24, N 14.80.
4’-epi-6,3’,6’,2’’,5’’,3’’’,4’’’-Hepta-O-benzylparomomycin (12). A soln. of 11 (71 mg, 0.05 mmol) in
THF (10 ml) was treated with 0.1m aq. NaOH (1.8 ml) and 1m PMe3 in THF (0.45 ml, 0.45 mmol), and
stirred for 2 h at 508. Evaporation and FC (amino-phase silica gel; MeOH/CHCl3 3 :97) gave 12 (42 mg,

1546
Helvetica Chimica Acta – Vol. 91 (2008)
Table 7. 13C-NMR Chemical Shifts [ppm] of the Protected Amino Derivatives 8, 12, and 16 in CDCl3, and
of the Deprotected Amino Derivatives 9, 13, and 17 in D2O
8
12
99.81
16
99.72
9
13a)
17
98.11
C(1’)
C(2’)
C(3’)
C(4’)
C(5’)
C(6’)
C(1)
C(2)
C(3)
C(4)
C(5)
99.87
51.60
80.71
51.96
70.11
70.11
51.40b)
38.62
51.04b)
76.32
86.04
87.27
107.00
81.74
80.28
81.16
70.71
100.81
49.10
75.64
71.90
74.57
42.46
100.86
55.12
67.67
57.17
75.13
65.51
54.66b)
33.06
53.81b)
82.83
89.03
77.17
114.83
78.37
80.06
86.42
65.09
101.09
55.84b)
72.61
72.26
73.16
45.44
99.21
52.01
80.63
66.38
69.97
70.26
51.73b)
38.63
51.41b)
76.36
86.54
87.17
107.19
81.57
79.67
81.18
70.26
101.18
51.30
75.63
73.00
74.61
42.29
53.26
80.43
56.88
73.39
70.04
51.99b)
38.49
51.72b)
76.29
86.55
86.96
107.45
83.54
81.19
81.57
70.35
101.65
51.41
75.71
72.95
74.56
42.42
53.59
68.91
70.57
75.30
63.79
52.72
32.51
51.81
82.12
87.31
75.62
112.65
76.14
77.85
83.92
62.74
98.20
53.62
70.43
70.03
72.99
43.15
57.11
69.99
55.17
74.90
62.85
53.57b)
32.66
52.76b)
83.93
87.30
75.53
112.62
76.08
77.85
81.78
63.12
99.12
51.69
70.53
70.39
72.94
43.10
C(6)
C(1’’)
C(2’’)
C(3’’)
C(4’’)
C(5’’)
C(1’’’)
C(2’’’)
C(3’’’)
C(4’’’)
C(5’’’)
C(6’’’)
a) Assignments based on a DQFCOSY and a HSQC spectrum. b) Assignments may be interchanged.
65%). White solid. Rf (MeOH/CHCl3 1:19) 0.21. M.p. 558. [a]2D5 ¼ þ33.4 (c ¼ 0.18, CHCl3). IR (CHCl3):
3375w (br.), 3309w, 3081w, 3054w, 2926m, 2874m, 1601m, 1493m, 1454m, 1366m, 1093m, 1070m, 1025m,
1
917w. H-NMR (CDCl3, 300 MHz): see Table 6; additionally, 7.37 – 7.19 (m, 35 arom. H); 4.94 (d, J ¼
11.4), 4.69 (d, J ¼ 11.7), 4.63 (d, J ¼ 12.0), 4.59 (d, J ¼ 12.9), 4.57 – 4.52 (m, 7 H), 4.50 (d, J ¼ 12.3),
4.44 (d, J ¼ 12.3), 4.39 (d, J ¼ 12.3) (7 PhCH2); 1.60 (br. s, w1/2 ꢂ 53.0, 7 NH2, OH). 13C-NMR (CDCl3,
75 MHz): see Table 7; additionally, 138.24, 137.98 (3 C), 137.71 (2 C), 137.18 (4s, 7 arom. C); 128.45 –
127.61 (several d, 35 arom. CH); 75.08, 73.62, 73.17, 73.09, 71.96 (2 C), 71.86 (6t, 7 PhCH2). HR-
MALDI-MS: 1284.5860 (54, [M þ K]þ, C72H87KN5O1þ4 ; calc. 1284.5887), 1268.6147 (92, [M þ Na]þ,
C72H87N5NaOþ14 ; calc. 1268.6147), 1246.6307 (98, [M þ H]þ, C72H88N5O1þ4 ; calc. 1246.6328), 905.4727 (79,
[M ꢀ C20H25N2O3 þ H]þ, C52H63N3O1þ1 ; calc. 905.4463), 565.2919 (100, [M ꢀ C20H25N2O3 ꢀ C20H24NO4 þ
3 H]þ, C32H41N2Oþ7 ; calc. 565.2914). Anal. calc. for C72H87N5O14 · 3 H2O (1300.53): C 66.49, H 7.21, N
5.38; found: C 66.69, H 7.15, N 5.48.
4’-epi-Paromomycin Pentaacetate Trihydrate (13). A soln. of 12 (30 mg, 0.02 mmol) in AcOH/H2O/
D2O 2 :2 :1 (3 ml) was treated with Pd(OH)2 (20 mg) and stirred for 36 h at r.t. under 6 bar of H2.
Filtration and evaporation gave 13 (18 mg, 82%). White solid. Rf (CHCl3/MeOH/25% aq. NH3 1:4 :3)
0.44. M.p. 1258 (dec.). [a]2D5 ¼ þ55.5 (c ¼ 0.19, H2O). 1H-NMR (D2O, 600 MHz; assignments based on a
DQFCOSY and a HSQC spectrum): see Table 6; additionally, 1.90 (s, 5 AcO). 13C-NMR (CDCl3,
150 MHz; assignments based on a HSQC spectrum): see Table 7; additionally, 183.70 (s, 5 C¼O); 25.72
(q, 5 MeC¼O). HR-ESI-MS: 638.2846 (100, [M þ Na]þ, C23H45N5NaO1þ4 ; calc. 638.2861), 616.3025 (78,
[M þ H]þ, C23H46N5O1þ4 ; calc. 616.3041), 455.2325 (100, [M ꢀ C6H13N2O3 þ 2 H]þ, C17H35N4O1þ0 ; calc.
455.2353). Anal. calc. for C23H45N5O14 · 5 AcOH · 3 H2O (969.93): C 40.86, H 7.38, N 7.22; found: C 41.14,
H 7.15, N 7.29.

Helvetica Chimica Acta – Vol. 91 (2008)
1547
4’-epi-1,3,2’,2’’’,6’’’-Pentaazido-6,3’,6’,2’’,5’’,3’’’,4’’’-hepta-O-benzyl-1,3,2’,2’’’,6’’’-pentadeamino-4’-O-
[(trifluoromethyl)sulfonyl]paromomycin (14). Under N2, a soln. of pyridine (0.03 ml, 0.36 mmol) and
Tf2O (0.06 ml, 0.36 mmol) in CH2Cl2 (5 ml) was stirred for 30 min at ꢀ 108, treated portionwise with a
soln. of 11 (206 mg, 0.15 mmol) in CH2Cl2 (5 ml) during 30 min, and stirred for 5 h at ꢀ 108 and for 4 h at
208. After neutralization with aq. NaHCO3 and extraction with CH2Cl2 (3 Â 30 ml), the combined org.
layers were washed with brine, dried (MgSO4), filtered, and evaporated. FC (AcOEt/cyclohexane 3 :22)
gave 14 (170 mg, 75%). White solid. Rf (AcOEt/cyclohexane 1:4) 0.44. M.p. 48.0 – 48.98. [a]2D5 ¼ þ79.1
(c ¼ 0.17, CHCl3). IR (CHCl3): 3054w, 2923w, 2870w, 2104s, 1601w, 1493w, 1454w, 1408m, 1362w, 1333w,
1259w, 1239w, 1141m, 1077w, 1031w, 916m. 1H-NMR (CDCl3, 300 MHz): see Table 4; additionally, 7.42 –
7.12 (m, 35 arom. H); 4.95 (d, J ¼ 10.5), 4.87 (d, J ¼ 11.4), 4.66 (d, J ¼ 10.5), 4.661 (d, J ¼ 11.4), 4.58 (d,
J ¼ 12.0), 4.54 (d, J ¼ 12.0), 4.52 (d, J ¼ 12.0, 2 H), 4.50 (d, J ¼ 11.7), 4.41 (d, J ¼ 11.7), 4.39 (d, J ¼ 11.7),
4.36 (d, J ꢂ 12.5), 4.28 (d, J ꢂ 12.3), 4.24 (d, J ¼ 12.3) (7 PhCH2). 13C-NMR (CDCl3, 75 MHz): see
Table 5; additionally, 137.93, 137.75, 137.46, 137.40, 136.91, 136.84, 136.28 (7s, 7 arom. C); 128.58 – 127.30
1
(several d, 35 arom. CH); 118.26 (q, J(C,F) ¼ 316.6, CF3); 74.94, 73.24, 73.23 (2 C), 72.29, 72.28, 71.60
(6t, 7 PhCH2). 19F-NMR (CDCl3, 282 MHz): ꢀ 73.85. HR-MALDI-MS: 1530.5119 (100, [ M þ Na]þ,
C73H76F3N15NaO16Sþ; calc. 1530.5165), 1380.5642 (56, [M ꢀ TfOH þ Na]þ, C72H75N15NaO1þ3 ; calc.
1380.5566). Anal. calc. for C73H76F3N15O16S(1508.54): C 58.12, H 5.08, N 13.93, S2.13; found: C
58.22, H 5.29, N 13.76, S2.27.
1,3,2’,4’,2’’’,6’’’-Hexaazido-6,3’,6’,2’’,5’’,3’’’,4’’’-hepta-O-benzyl-1,3,2’,2’’’,6’’’-pentadeamino-4’-deoxy-
paromomycin (15). Under N2, a soln. of 14 (106 mg, 0.076 mmol) in MeCN (3 ml) was treated with
Bu4NN3 (238 mg, 0.84 mmol) and stirred for 10 h at r.t. After evaporation, a soln. of the residue in CH2Cl2
(10 ml) was washed twice with brine, dried (MgSO4), filtered, and evaporated. FC (amino-phase silica
gel; AcOEt/cyclohexane 3 :22) gave 15 (62 mg, 63%). Colourless oil. Rf (amino-phase silica gel; AcOEt/
cyclohexane 1:3) 0.45. [a]2D5 ¼ þ98.1 (c ¼ 0.17, CHCl3). IR (CHCl3): 2868w, 2105s, 1603w, 1495w, 1454w,
1364w, 1322w, 1264m, 1081m, 1027m, 919w. 1H-NMR (CDCl3, 300 MHz): see Table 4; additionally, 7.45 –
7.14 (m, 35 arom. H); 4.96 (d, J ¼ 10.5), 4.85 (d, J ¼ 10.8), 4.81 (d, J ꢂ 11.0), 4.67 (d, J ¼ 12.3, 2 H), 4.62 (d,
J ¼ 12.0), 4.58 (d, J ¼ 12.0), 4.56 (d, J ¼ 11.7, 2 H), 4.49 (d, J ¼ 12.6), 4.42 (d, J ¼ 12.0), 4.41 (d, J ¼ 12.0),
4.31 (d, J ¼ 12.0), 4.25 (d, J ¼ 12.0) (7 PhCH2). 13C-NMR (CDCl3, 75 MHz): see Table 5; additionally,
138.02, 137.82, 137.74, 137.41, 137.21, 136.87, 136.80 (7s, 7 arom. C), 128.56 – 127.31 (several d, 35 arom.
CH); 75.35, 75.07, 73.40, 73.26, 73.23, 72.36, 71.71 (7t, 7 PhCH2). HR-ESI-MS: 1439.5408 (13, [M þ K]þ,
C72H76KN18Oþ13 ; calc. 1439.5476), 1423.5705 (100, [M þ Na]þ, C72H76N18NaO1þ3 ; calc. 1423.5737),
1373.5836 (18, [M ꢀ N2 þ H]þ, C72H77N16Oþ13 ; calc. 1373.5856). Anal. calc. for C72H76N18O13 · H2O
(1419.50): C 60.92, H 5.54, N 17.76; found: C 60.70, H 5.31, N 17.33.
4’-Amino-6,3’,6’,2’’,5’’,3’’’,4’’’-hepta-O-benzyl-4’-deoxyparomomycin (16). A soln. of 15 (38 mg,
0.03 mmol) in THF (6 ml) was treated with 0.1m aq. NaOH (1.08 ml) and 1m PMe3 in THF (0.32 ml,
0.32 mmol), stirred for 2 h at 508, and evaporated. FC (amino-phase silica gel; MeOH/cyclohexane 3 :97)
gave 16 (27 mg, 80%). White solid. Rf (MeOH/CHCl3 1:19) 0.23. M.p. 45.3 – 47.28. [a]2D5 ¼ þ21.5 (c ¼
0.22, CHCl3). IR (CHCl3): 3389w (br.), 3305w (br.), 3059w, 2924m, 2865m, 1601m, 1496w, 1454m,
1361w, 1311w, 1094m, 1068m, 1026m, 908m. 1H-NMR (CDCl3, 300 MHz): see Table 6; additionally, 7.35 –
7.20 (m, 35 arom. H); 4.93 (d, J ¼ 11.1), 4.89 (d, J ¼ 11.7), 4.68 (d, J ¼ 11.4), 4.58 (d, J ꢂ 13.0), 4.57 (d, J ¼
12.3, 2 H), 4.53 (d, J ꢂ 13.0), 4.532 (d, J ¼ 11.1, 2 H), 4.51 (d, J ¼ 11.7), 4.44 (d, J ¼ 12.0, 2 H), 4.42 (d, J ¼
12.0), 4.23 (d, J ¼ 11.4) (7 PhCH2); 1.53 (br. s, w1/2 ¼ 72.0, 6 NH2). 13C-NMR (CDCl3, 75 MHz): see
Table 7; additionally, 138.65, 138.18, 137.98, 137.94, 137.67 (2 C), 137.13 (6s, 7 arom. C), 128.44 – 127.46
(several d, 35 arom. CH); 75.12, 74.43, 73.39, 73.21, 73.05, 71.92 (2 C) (6t, 7 PhCH2). Anal. calc. for
C72H88N6O13 · 3 H2O (1299.55): C 66.54, H 7.29, N 6.47; found: C 66.23, H 7.01, N 6.48.
4’-Amino-4’-deoxyparomomycin Hexaacetate (17). A soln. of 16 (16 mg, 0.01 mmol) in AcOH/H2O/
D2O 2 :2 :1 (3 ml) was treated with Pd(OH)2 (13 mg) and stirred for 12 h at r.t. under 6 bar of H2.
Filtration and evaporation gave 17 (11 mg, 88%). White solid. Rf (CHCl3/MeOH/25% aq. NH3 1:4 :3)
0.40. M.p. 1258 (dec.). [a]2D5 ¼ þ44.6 (c ¼ 0.21, H2O). 1H-NMR (D2O, 400 MHz): see Table 6;
additionally, 1.79 (s, 6 AcO). 13C-NMR (CDCl3, 100 MHz): see Table 7; additionally, 183.89 (s,
6 OC¼O); 25.77 (q, 6 MeC¼O). HR-ESI-MS: 637.3035 (86, [M þ Na]þ, C23H46N6NaO1þ3 ; calc.
637.3021), 477.2178 (100, [M ꢀ C6H13N2O3 þ Na þ H]þ, C17H34N4NaO1þ0 ; calc. 477.2178).

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Helvetica Chimica Acta – Vol. 91 (2008)
1,3,2’,2’’’,6’’’-Pentaazido-6,3’,2’’,5’’,3’’’,4’’’-hexa-O-benzyl-1,3,2’,2’’’,6’’’-pentadeaminoparomomycin
(18). A soln. of 4 (3.37 g, 2.45 mmol) in CH2Cl2/MeOH 4 :1 (25 ml) was treated at r.t. with TsOH · H2O
(0.70 g, 3.68 mmol), and stirred for 12 h. After evaporation, FC (AcOEt/cyclohexane 2 :3) gave 18
(1.33 g, 42%) and 4 (1.4 g, 42%). Rf (AcOEt/cyclohexane 1:1) 0.59. White solid. IR (ATR): 3446w,
1
3030w, 2871w, 2099s, 1605w, 1496w, 1454m, 1361m, 1215m, 1026s, 913m. H-NMR (CDCl3, 400 MHz;
assignments based on a DQFCOSY and a HSQC spectrum): 7.51 – 7.14 (m, 30 arom. H); 6.17 (d, J ¼ 3.6,
HꢀC(1’)); 5.69 (d, J ¼ 5.8, HꢀC(1’’)); 4.99 (d, J ¼ 10.5, PhCH); 4.93 (d, J ꢂ 11.4, PhCH); 4.91 (br. s,
HꢀC(1’’’)); 4.72 (d, J ¼ 10.7, PhCH); 4.67 (d, J ¼ 11.4, PhCH); 4.63 (d, J ¼ 12.0, PhCH); 4.59 (d, J ¼ 11.8,
PhCH); 4.482 (s, PhCH2); 4.475 (d, J ¼ 11.8, PhCH); 4.41 (d, J ¼ 12.0, PhCH); 4.33 (br. s, HꢀC(4’’)); 4.32
(br. s, HꢀC(3’’)); 4.32 (d, J ¼ 12.0, PhCH); 4.26 (d, J ¼ 12.1, PhCH); 3.99 (t, J ¼ 5.1, HꢀC(2’’)); 3.97 (d,
J ¼ 8.9, HꢀC(5)); 3.91 – 3.82 (m, HꢀC(3’), HꢀC(5’), HaꢀC(5’’)); 3.82 – 3.70 (m, 2 HꢀC(6’), HꢀC(3’’’),
HꢀC(5’’’)); 3.70 – 3.57 (m, HꢀC(4), HbꢀC(5’’), HaꢀC(6’’’)); 3.48 – 3.36 (m, HꢀC(1), HꢀC(3),
HꢀC(4’)); 3.36 (br. t, J ꢂ 2.0, HꢀC(2’’’)); 3.30 (t, J ¼ 9.3, HꢀC(6)); 3.13 (br. t, J ꢂ 1.9, HꢀC(4’’’));
2.89 (dd, J ¼ 12.9, 3.75, HbꢀC(6’’’)); 2.86 (dd, J ¼ 10.2, 3.65, HbꢀC(2’)); 2.23 (dt, J ¼ 13.2, 4.5, HeqꢀC(2));
2.16 (d, J ¼ 3.3, HOꢀC(4’)); 1.67 (dd, J ¼ 8.1, 4.9, HOꢀC(6’)); 1.40 (q, J ¼ 12.7, HaxꢀC(2)). 13C-NMR
(CDCl3, 100 MHz; assignments based on a HSQC spectrum): 138.31, 138.11, 137.93, 137.56, 137.02, 136.95
(6s, 6 arom. C); 128.70 – 127.15 (several d, 30 arom. CH); 106.17 (d, C(1’’)); 98.69 (d, C(1’’’)); 95.97 (d,
C(1’)); 84.29 (d, C(6)); 82.54 (d, C(2’’)); 82.19 (d, C(4’’)); 82.01 (d, C(5)); 79.71 (d, C(3’)); 75.54 (d,
C(3’’)); 75.09, 74.94 (2t, 2 PhCH2); 74.94 (d, C(4)); 74.45 (d, C(5’’’)); 73.22 (t, 2 PhCH2); 72.93 (d, C(3’’’));
72.42, 71.76 (2t, 2 PhCH2); 71.61 (d, C(4’’’)); 71.56 (d, C(5’)); 70.46 (d, C(4’)); 70.32 (t, C(5’’)); 62.66 (d,
C(2’)); 62.16 (t, C(6’)); 60.41, 60.29 (2d, C(1), C(3)); 57.28 (d, C(2’’’)); 51.19 (t, C(6’’’)); 32.56 (t, C(2)).
HR-MALDI-MS: 1308.5197 (100, [M þ Na]þ, C67H71N15NaO1þ4 ; calc. 1308.5203).
6,3’,2’’,5’’,3’’’,4’’’-Hexa-O-benzyl-1,3,2’,2’’’,6’’’-pentakis-N-[(benzyloxy)carbonyl]paromomycin (19).
A soln. of 18 (1.15 g, 0.89 mmol) in THF (24 ml) was treated with 0.1m aq. NaOH (6 ml) and 1m PMe3
in THF (6.7 ml, 6.7 mmol), stirred for 2.5 h at 508, and evaporated. An ice-cold soln. of the crude product
and Na2CO3 (1.3 g, 12.3 mmol) in MeOH (20 ml) was treated dropwise with CbzCl (0.76 ml, 5.4 mmol),
and stirred at r.t. for 6 h. After filtration and evaporation, a soln. of the residue in AcOEt (20 ml) was
washed with H2O and brine, dried (Na2SO4), filtered, and evaporated. FC (AcOEt/cyclohexane 2 :3)
gave 19 (1.3 g, 80%). Rf (AcOEt/cyclohexane 2 :1) 0.58. IR (ATR): 3419w, 3330w, 3063w, 3032w, 2933w,
1704s, 1514s, 1497m, 1454m, 1331m, 1299m, 1216s, 1043s, 1025s, 912m. HR-MALDI-MS: 1849.7579 (100,
[M þ Na]þ, C105H111N5NaO2þ4 ; calc. 1849.7550).
6,3’,2’’,5’’,3’’’,4’’’-Hexa-O-benzyl-1,3,2’,2’’’,6’’’-pentakis-N-[(benzyloxy)carbonyl]-6’-O-(2,2-dimethyl-
propanoyl)paromomycin (20). A soln. of 19 (600 mg, 0.47 mmol) in CH2Cl2/pyridine 5 :3 (16 ml) was
treated with pivaloyl chloride (0.3 ml, 2.45 mmol) and stirred for 12 h at r.t. Evaporation and FC (AcOEt/
cyclohexane 2 :3) gave 20 (523 mg, 83%). Rf (AcOEt/cyclohexane 1:1) 0.63. IR (ATR): 3415w, 3036w,
2938w, 2844w, 1715s, 1513m, 1454m, 1212m, 1052s, 1032s, 1016s, 910w. HR-MALDI-MS: 1933.8059 (100,
[M þ Na]þ, C110H119N5NaO2þ5 ; calc. 1933.8125).
6,3’,2’’,5’’,3’’’,4’’’-Hexa-O-benzyl-1,3,2’,2’’’,6’’’-pentakis-N-[(benzyloxy)carbonyl]-6’-O-(2,2-dimethyl-
propanoyl)-4’-O-[(methylsulfanyl)carbothioyl]paromomycin (21). A suspension of NaH (16 mg,
0.41 mmol) in THF (10 ml) at 08 was treated with a soln. of 20 (523 mg, 0.27 mmol) in THF (10 ml),
stirred for 10 min, treated with CS2 (0.3 ml, 5 mmol) and 1H-imidazole (10 mg, 0.15 mmol), stirred at r.t.
for 2 h, treated with MeI (50 ml, 0.81 mmol), and stirred for 20 min. The mixture was treated with sat. aq.
NH4Cl (10 ml), and extracted with AcOEt (3 Â 15 ml). The combined org. layers were washed with
brine, dried (Na2SO4), filtered, and evaporated. FC (AcOEt/cyclohexane 1:3) gave 21 (460 mg, 84%). Rf
(AcOEt/cyclohexane 2 :3) 0.50. IR (ATR): 3424w, 3334w, 3063w, 3031w, 2932w, 1716s, 1586w, 1513m,
1454m, 1331m, 1212s, 1045s, 1026s, 964m, 912w. HR-MALDI-MS: 2023.7670 (100, [ M þ Na]þ,
C112H121N5NaO25Sþ2 ; calc. 2023.7723).
6,3’,2’’,5’’,3’’’,4’’’-Hexa-O-benzyl-1,3,2’,2’’’,6’’’-pentakis-N-[(benzyloxy)carbonyl]-4’-deoxy-6’-O-(2,2-
dimethylpropanoyl)paromomycin (22). A soln. of 21 (355 mg, 0.18 mmol), Et3N (0.2 ml, 1.44 mmol),
and H3PO2 (0.2 ml, 3.65 mmol) in 1,4-dioxane (10 ml) was heated to reflux, treated with a soln. of AIBN
(300 mg, 1.83 mmol) in 1,4-dioxane (5 ml) during 15 min, and kept at reflux for 24 h. The mixture was
cooled to r.t., diluted with AcOEt (20 ml), washed with H2O (2 Â 12 ml) and brine, dried (Na2SO4),
filtered, and evaporated. FC (AcOEt/cyclohexane 1:3) afforded 22 (303 mg, 84%). Rf (AcOEt/

Helvetica Chimica Acta – Vol. 91 (2008)
1549
cyclohexane 2 :3) 0.50. IR (ATR): 3424w, 3021w, 2972w, 1715s, 1605w, 1515m, 1454m, 1233m, 1092m,
1047s, 1032s, 1020s, 912w. HR-MALDI-MS: 1916.8143 (100, [ M þ Na]þ, C110H119N5NaO2þ4 ; calc.
1917.8176).
4’-Deoxyparomomycin (23). A cold (ꢀ 788) soln. of condensed NH3 (ca. 20 ml) in THF (5 ml) was
treated portionwise with Na (70 mg, 3.04 mmol). The resulting dark blue soln. was treated dropwise with
a soln. of 22 (230 mg, 0.12 mmol) in THF (5 ml) and then allowed to reach r.t. over 5 h. After dilution
with MeOH (10 ml), the mixture was neutralized with Amberlite IR-120 (Hþ form), filtered, and
evaporated. FC (amino-phase silica gel: CHCl3/MeOH/25% NH3 15 :45 :1), evaporation, filtration, and
lyophilization gave 23 (51 mg, 68%). White solid. Rf (CHCl3/MeOH/25% aq. NH3 1:3 :2) 0.39. IR
(ATR): 3103m, 2909m, 1543s, 1454w, 1379w, 1344w, 1138w, 1015s, 932w. 1H-NMR (D2O, 400 MHz;
assignments based on a DQFCOSY and a HSQC spectrum): 5.25 (d, J ¼ 3.7, HꢀC(1’)); 5.23 (d, J ¼ 2.9,
HꢀC(1’’)); 4.83 (d, J ¼ 1.8, HꢀC(1’’’)); 4.29 (dd, J ¼ 6.1, 5.2, HꢀC(3’’)); 4.13 (dd, J ¼ 5.1, 2.9, HꢀC(2’’));
4.01 – 3.95 (m, HꢀC(5’), HꢀC(4’’)); 3.88 (t, J ¼ 3.3, HꢀC(3’’’)); 3.82 (dt, J ꢂ 8.4, 4.0, HꢀC(5’’’)); 3.76 –
3.67 (m, HꢀC(3’), HaꢀC(6’)); 3.59 (dd, J ¼ 12.4, 5.3, HbꢀC(6’)); 3.53 (dd, J ¼ 12.2, 3.5, HaꢀC(5’’)); 3.50
(t, J ¼ 9.3, HꢀC(5)); 3.50 (t, J ꢂ 3.0, HꢀC(4’’’)); 3.44 (dd, J ¼ 12.2, 6.4, HbꢀC(5’’)); 3.29 (t, J ¼ 9.3,
HꢀC(4)); 3.13 (t, J ¼ 9.6, HꢀC(6)); 2.89 (dd, J ¼ 13.5, 8.5, HaꢀC(6’’’)); 2.89 (dd, J ¼ 3.8, 1.8, HꢀC(2’’’));
2.76 (dd, J ¼ 13.6, 4.3, HbꢀC(6’’’)); 2.72 (ddd, J ¼ 12.4, 9.7, 4.2, HꢀC(3)); 2.58 (ddd, J ¼ 12.0, 9.8, 3.8,
HꢀC(1)); 2.50 (dd, J ¼ 10.1, 3.6, HꢀC(2’)); 1.86 – 1.76 (m, HeqꢀC(2), HeqꢀC(4’)); 1.28 (q, J ¼ 12.0,
HaxꢀC(4’)); 1.06 (q, J ¼ 12.4, HaxꢀC(2)). 13C-NMR (D2O, 100 MHz; assignments based on a HSQC
spectrum): 110.99 (d, C(1’’)); 102.80 (d, C(1’)); 102.08 (d, C(1’’’)); 86.87 (d, C(5)); 85.84 (d, C(4)); 84.21
(d, C(4’’)); 80.11 (d, C(6)); 78.64 (d, C(3’’)); 77.82 (d, C(5’’’)); 76.08 (d, C(2’’)); 73.27 (d, C(3’’’)); 72.63 (d,
C(5’)); 71.17 (d, C(4’’’)); 70.42 (d, C(3’)); 66.48 (t, C(5’’)); 63.88 (t, C(6’)); 59.64 (d, C(2’)); 55.29 (d,
C(2’’’)); 52.92 (d, C(1), C(3)); 43.70 (t, C(6’’’)); 38.14 (t, C(2)); 37.10 (t, C(4’)). HR-MALDI-MS: 600.3087
(100, [M þ H]þ, C23H46N5O1þ3 ; calc. 600.3092).
d-Glyceraldehyde-(2 ! 1)-(1R)-2-azido-2-deoxy-d-glycerodialdehyde-(1 ! 4)-1,3,2’’’,6’’’-tetraazido-
1,3,2’’’,6’’’-tetradeamino-4-O-de(2-amino-2-deoxy-a-d-glucopyranosyl)paromomycin (24). Under N2, a
soln. of 2 (163 mg, 0.22 mmol) in THF (20 ml) was treated with 4-Š molecular sieves, stirred for 1 h,
cooled to 08, treated with H5IO6 (72 mg, 0.32 mmol), stirred for 3 h, neutralized with sat. aq. NaHCO3,
and extracted with AcOEt (3 Â 20 ml). The combined org. layers were washed with brine, dried
(MgSO4), filtered, and evaporated to afford crude 24 (90 mg, ca. 55%) that was used as such for further
reactions. Yellow oil. Rf (CHCl3/AcOEt/MeOH 4 :8 :1) 0.42.
1,3,2’’’,6’’’-Tetraazido-1,3,2’’’,6’’’-tetradeamino-4-O-de(2-amino-2-deoxy-a-d-glucopyranosyl)-4-O-
(3,4,5-trideoxy-4-N-phenyl-4-aza-a-d-erythro-heptoseptanosyl)paromomycin (25). Under N2, a soln. of
crude 24 (300 mg, 0.4 mmol) in MeOH (10 ml) was treated with NaCNBH3 (102 mg, 1.61 mmol) and
portionwise, over 1 h, with a soln. of PhNH2 (37 ml, 0.40 mmol) and AcOH (97 ml, 1.7 mmol) in MeOH
(3 ml), and stirred at r.t. for 12 h. Et3N (1 ml) was added, and the mixture was taken to dryness. A
suspension of the residue in sat. aq. NaHCO3 was extracted with AcOEt (3 Â 20 ml). The combined org.
layers were washed with brine, dried (MgSO4), filtered, and evaporated. FC (CHCl3/AcOEt 5 :9) gave 25
(90 mg, 32% from 2). Yellow oil. Rf (CHCl3/AcOEt/MeOH 4 :8 :1) 0.52. 1H-NMR (CD3OD, 500 MHz;
assignments based on a DQFCOSYand a HSQC spectrum): see Table 8; additionally, 7.17 (dt, J ¼ 8.8, 1.6,
2 arom. H); 6.88 (dd, J ¼ 8.9, 0.9, 2 arom. H); 6.80 (dt, J ¼ 7.3, 0.9, 1 arom. H). 13C-NMR (CD3OD,
125 MHz; assignments based on a HSQC spectrum): see Table 9; additionally, 151.48 (s); 130.13 (d, 2 C);
118.65 (d); 114.99 (d, 2 C). HR-MALDI-MS: 827.2883 (32, [M þ Na]þ, C29H40N16NaO1þ2 ; calc. 827.2909),
805.3070 (100, [M þ H]þ), C29H41N16Oþ12 ; calc. 805.3090), 461.2003 (46, [M ꢀ C11H17N6O7 þ 2 H]þ,
C18H25N10NaOþ5 ; calc. 461.2009).
1,3,2’’’,6’’’-Tetraazido-1,3,2’’’,6’’’-tetradeamino-4-O-de(2-amino-2-deoxy-a-d-glucopyranosyl)-4-O-(4-
N-benzyl-3,4,5-trideoxy-4-aza-a-d-erythro-heptoseptanosyl)paromomycin (26). Under N2, a soln. of
crude 24 (300 mg, 0.4 mmol) in MeOH (10 ml) was treated with NaCNBH3 (102 mg, 1.61 mmol) and
portionwise, during 1 h, with a soln. of BnNH2 (44 ml, 0.40 mmol) and AcOH (97 ml, 1.7 mmol) in MeOH
(3 ml), stirred at r.t. for 12 h, basified with Et3N (1 ml), and evaporated. A suspension of the residue in
sat. aq. NaHCO3 was extracted with AcOEt (3 Â 20 ml). The combined org. layers were washed with
brine, dried (MgSO4), filtered, and evaporated. FC (CHCl3/AcOEt 5 :9) gave 26 (100 mg, 36% from 2).
Yellow oil. Rf (CHCl3/AcOEt/MeOH 20 :40 :7) 0.63. 1H-NMR (CD3OD, 300 MHz): see Table 8;

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Helvetica Chimica Acta – Vol. 91 (2008)
Table 8. 1H-NMR Chemical Shifts [ppm] and Coupling Constants [Hz] of the Azides 25 and 26 in
CD3OD, and the Amines 27 and 28 in D2O
25a)
26
27a)
28a)
25
26
27
1.4
5.0
3.2
28
1.9
)
2.3
HꢀC(1’)
HꢀC(2’)
HaꢀC(3’)
HbꢀC(3’)
HaꢀC(5’)
HbꢀC(5’)
HꢀC(6’)
HaꢀC(7’)
HbꢀC(7’)
5.86
5.62
5.45
5.22
J(1’,2’)
2.3
3.6
)
)
b
b
b
b
b
b
4.04 – 3.99 3.54 – 3.29 3.38 – 3.39 3.22 – 3.21 J(2’,3’a)
4.04 – 3.99 3.00
3.44 – 3.40 2.57
)
)
)
3.94
3.33
3.01 – 2.95 J(2’,3’b)
2.53 J(3’a,3’b)
13.2 14.9 13.4
3.81
3.18
4.41
3.83
3.65
3.06
2.33
3.83 – 3.79 3.01 – 2.95 J(5’a,5’b)
3.21 2.33 J(5’a,6’)
4.04 – 4.01 J(5’b,6’)
3.73 J(6’,7’a)
14.5 12.8 15.3 14.2
10.4
5.3
5.3
3.1
9.9 10.6 10.6
b
4.36 – 4.32 4.38
3.78 3.81
3.73 – 3.62 3.74
6.0
2.7
)
7.6
7.0
3.1
)
3.5
)
b
b
3.63 – 3.59 J(6’,7’b)
J(7’a,7’b)
11.8 11.9 12.1 13.1
12.5 12.3 12.0 12.0
HꢀC(1)
HaxꢀC(2)
HeqꢀC(2)
HꢀC(3)
HꢀC(4)
HꢀC(5)
HꢀC(6)
3.44 – 3.40 3.54 – 3.29 2.73
2.68
1.15
1.96
2.78
3.43
3.50
3.19
J(1,2ax)
J(1,2eq)
J(1,6)
J(2ax,2eq)
J(2ax,3)
J(2eq,3)
J(3,4)
1.29
2.13
3.50
3.78
3.57
1.23
2.10
1.21
2.00
4.4
9.8
4.2
)
4.2
9.9
4.1
9.8
b
3.54 – 3.29 2.83
3.54 – 3.29 3.53
3.54 – 3.29 3.56
13.0 12.6 12.9 13.0
12.5 12.3 12.3 12.2
4.1
9.4
9.4
9.2
2.7
4.8
5.9
2.6
5.9
4.2
)
)
4.1
9.2
9.2
9.1
2.4
5.0
6.4
3.4
)
4.2
9.6
9.4
9.3
2.4
4.9
6.4
3.6
6.9
b
b
b
3.44 – 3.40 3.54 – 3.29 3.25
J(4,5)
J(5,6)
)
HꢀC(1’’)
HꢀC(2’’)
HꢀC(3’’)
HꢀC(4’’)
5.24
4.25
4.33
4.13
5.19
4.21
4.27
5.28
4.28
4.43
5.21
4.24
4.35
4.07
3.58
3.50
4.91
J(1’’,2’’)
J(2’’,3’’)
J(3’’,4’’)
J(4’’,5’’a)
J(4’’,5’’b)
J(5’’a,5’’b)
J(1’’’,2’’’)
3.0
4.9
6.0
)
b
4.15 – 4.04 4.15
3.73 – 3.62 3.87
3.58
5.12
3.54 – 3.29 3.04
3.93 4.03
b
HaꢀC(5’’) 3.81
HbꢀC(5’’) 3.66
6.0
3.73
4.97
11.5 11.9 12.1 11.7
HꢀC(1’’’)
HꢀC(2’’’)
HꢀC(3’’’)
HꢀC(4’’’)
HꢀC(5’’’)
5.14
3.68
6.34
1.9
1.5
3.4
)
5.7
4.4
1.8
3.3
3.3
1.8
8.7
4.2
)
1.9
3.3
3.3
1.7
8.0
4.3
1.9
3.3
3.3
1.6
8.4
4.3
3.01 – 2.95 J(2’’’,3’’’)
4.00
3.62
3.89
J(3’’’,4’’’)
J(4’’’,5’’’)
J(5’’’,6’’’a)
b
3.44 – 3.40 3.54 – 3.29 3.66
4.04 – 3.99 4.00
3.98
3.54 – 3.29 3.06
3.54 – 3.29 2.95
HaꢀC(6’’’) 3.64
3.01 – 2.95 J(5’’’,6’’’b)
2.85
b
HbꢀC(6’’’) 3.37
J(6’’’a,6’’’b) 13.0
13.5 13.5
a) Assignments based on a DQFCOSY and a HSQC spectrum. b) Not assigned.
additionally, 7.39 – 7.20 (m, 5 arom. H); 3.68 (s, PhCH2). 13C-NMR (CD3OD, 75 MHz): see Table 9;
additionally, 138.86 (s); 129.87 (d, 2 C); 128.88 (d, 2 C); 127.83 (d); 63.72 (t, PhCH2). HR-MALDI-MS:
841.3097 (12, [M þ Na]þ), C30H42N16NaOþ12 ; calc. 841.3066), 819.3225 (100, [M þ H]þ), C30H43N16Oþ12
;
calc. 819.3246).
4-O-De(2-amino-2-deoxy-a-d-glucopyranosyl)-4-O-(3,4,5-trideoxy-4-N-phenyl-4-aza-a-d-erythro-
heptoseptanosyl)paromomycin (27). A soln. of 25 (90 mg, 0.11 mmol) in THF (10 ml) was treated with
0.1m aq. NaOH (3.1 ml) and 1m PMe3 in THF (0.84 ml, 0.84 mmol), heated to 508 for 2 h, and evaporated.
FC (MeOH/25% aq. NH3 4 :3) gave 27 (50 mg, 66%). White solid. Rf (CHCl3/MeOH/25% aq. NH3
1:3 :4) 0.55. [a]2D5 ¼ þ18.6 (c ¼ 0.23, H2O). 1H-NMR (D2O, 500 MHz; assignments based on a
DQFCOSY and a HSQC spectrum): see Table 8; additionally, 7.32 (dt, J ¼ 7.3, 1.5, 2 arom. H); 6.95
(dd, J ¼ 8.1, 2 arom. H); 6.80 (t, J ¼ 7.3, 1 arom. H). 13C-NMR (D2O, 125 MHz; assignments based on a
HSQC spectrum): see Table 9; additionally, 151.55 (s); 132.44 (d, 2 C); 119.94 (d); 115.10 (d, 2 C). HR-
MALDI-MS: 697.3384 (50, [M þ Na]þ), C29H50N6NaO1þ2 ; calc. 697.3384), 675.3551 (100, [M þ H]þ),

Helvetica Chimica Acta – Vol. 91 (2008)
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Table 9. 13C-NMR Chemical Shifts [ppm] and Coupling Constants [Hz] of the Azides 19 and 20 in
CD3OD, and the Amines 21 and 22 in D2O
25a)
26
99.56
27a)
28a)
25a)
26
27a)
28a)
C(1’)
C(2’)
C(3’)
C(5’)
C(6’)
C(7’)
C(1)
C(2)
C(3)
C(4)
C(5)
C(6)
100.48
64.44
51.59
54.26
73.75
63.78
61.46
33.46
62.03
78.94
86.51
77.38
101.29
57.34
53.96
53.86
74.84
63.78
52.99
37.48
52.06
83.70
86.44
79.52
101.08
55.33
59.85
59.29
75.79
64.24
53.03
37.90
52.06
86.68
84.24
79.76
C(1’’)
C(2’’)
C(3’’)
C(4’’)
C(5’’)
111.42
75.17
77.52
83.63
63.92
111.11
74.26
77.16
83.13
64.02
111.65
75.79
78.90
84.08
64.41
111.67
75.64
78.94
84.02
64.41
61.26
55.36
59.77
74.76
63.34
61.38
33.21
61.68
78.70
86.10
77.16
C(1’’’)
C(2’’’)
C(3’’’)
C(4’’’)
C(5’’’)
C(6’’’)
100.00
61.77
71.18
69.62
75.71
52.57
100.89
61.24
70.74
69.25
75.33
52.24
101.94
55.27
73.19
71.15
77.35
43.65
102.03
55.25
73.26
71.14
78.10
43.73
a) Assignments based on a DQFCOSY and a HSQC spectrum.
C29H51N6Oþ12 ; calc. 675.3565). Dissolution of 27 in 50% aq. AcOH, followed by evaporation and
lyophilization, gave 27 · 5 AcOH · H2O. Anal. calc. for C29H50N6O12 · 5 AcOH · H2O (993.02): C 47.17, H
7.31, N 8.46; found: C 46.80, H 7.27, N 8.46.
4-O-De(2-amino-2-deoxy-a-d-glucopyranosyl)-4-O-(4-N-benzyl-3,4,5-trideoxy-4-aza-a-d-erythro-
heptoseptanosyl)paromomycin (28). A soln. of 26 (90 mg, 0.11 mmol) in THF (10 ml) was treated with
0.1m aq. NaOH (3.1 ml) and 1m PMe3 in THF (0.84 ml, 0.84 mmol), heated to 508 for 2 h, and evaporated.
FC (MeOH/25% aq. NH3 4 :3) gave 28 (52 mg, 68%). White solid. Rf (CHCl3/MeOH/25% aq. NH3
1:3 :4) 0.55. [a]2D5 ¼ þ24.7 (c ¼ 0.22, H2O). IR (KBr): 3360s (br.), 3287s (br.), 2922s, 1595s, 1491m,
1454m, 1381m, 1356m, 1237w, 1149s, 1028s, 930s, 856w. 1H-NMR (D2O, 500 MHz; assignments based on a
DQFCOSYand a HSQC spectrum): see Table 8; additionally, 7.43 – 7.34 (m, 5 arom. H); 3.74 (s, PhCH2).
13C-NMR (D2O, 125 MHz; assignments based on a HSQC spectrum): see Table 9; additionally, 139.60
(s); 132.61 (d, 2 C); 131.21 (d, 2 C); 130.45 (d); 65.15 (t, PhCH2). HR-MALDI-MS: 711.3555 (23, [ M þ
Na]þ), C30H52N6NaO1þ2 ; calc. 711.3541), 689.3705 (100, [M þ H]þ), C30H53N6O1þ2 ; calc. 689.3721).
Dissolution of 28 in 50% aq. AcOH, followed by evaporation and lyophilization, gave 28 · 5 AcOH · H2O.
Anal. calc. for C30H52N6O12 · 5 AcOH · H2O (1007.04): C 47.71, H 7.41, N 8.35; found: C 47.33, H 7.13, N
8.36.
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Received April 3, 2008