Mannich reaction: An approach for the synthesis of water soluble mulundocandin analogues

Article abstract of DOI:10.1016/j.bmc.2004.01.015

Semisynthetic modifications at Hydroxy tyrosine (Htyr) unit of mulundocandin (1) were carried out to improve its aqueous solubility. A single step introduction of substituted aminomethyl groups at the ortho position(s) of phenolic hydroxyl of HTyr unit of

Full text of DOI:10.1016/j.bmc.2004.01.015

Bioorganic & Medicinal Chemistry 12(2004) 1751–1768
Mannich reaction: an approach for the synthesis of water soluble
?
mulundocandin analogues§,
Bansi Lal,* Vitthal Genbhau Gund, Nandu Baban Bhise
and Ashok Kumar Gangopadhyay
Quest Institute of LifeSciences, Nicholas Piramal India Limited, Mulund (w), Mumbai 4000 80, India
Received 6 November 2003; revised 2 January 2004; accepted 5 January 2004
Abstract—Semisynthetic modifications at Hydroxy tyrosine (Htyr) unit of mulundocandin (1) were carried out to improve its
aqueous solubility. A single step introduction of substituted aminomethyl groups at the ortho position(s) of phenolic hydroxyl of
HTyr unit of mulundocandin has been achieved in 7–85% yield. The in vitro screening of Mannich products against Candida albi-
cans and Aspergillus fumigatus, retained the in vivo activity of parent by oral and intraperitoneal route. Compound 20, showed
significant improvement in activity over mulundocandin (1) and activity compares well with that of fluconazole.
# 2004 Elsevier Ltd. All rights reserved.
1. Introduction
the cyclic lipopeptides belonging to echinocandin
family.6 They have a unique mechanism of action in
which they irreversibly inhibit the synthesis of (1,3)-b-d-
glucan, an essential polysaccharide in the wall of many
important fungal pathogens. Glucans are vital for
maintaining the structural integrity of the fungal cell
wall; therefore, the (1,3)-b-d-glucan synthase inhibitors
are cidal for many different fungi.7 Since this poly-
saccharide is not present in mammalian cells the glucan
synthase became an attractive target for the develop-
ment of new antifungal agents. Uniquely, unlike other
antifungal agents, the echinocandins are active against
Pneumocystis carinii. This novel mode of action and
potent antifungal activity has led to the development of
several interesting drug candidates during the past sev-
eral years. Caspofungin8 (Cancidas1) and Micafungin
(FK 463 )9 new echinocandin antifungals for the treat-
ment of invasive Aspergillus infections in patients unre-
sponsive to or unable to receive amphotericin B10 are
introduced in the market. Other echinocandin (LY
303366) and triazole antifungals are under development
in attempts to provide improved therapy for fungal
infections.11
The incidence of systemic fungal infections essentially
invasive candidiasis, but also invasive aspergillosis, has
increased dramatically over the past few decades,
resulting in an ever-increasing therapy challenge for
clinicians.1,2 Despite the currently available antifungal
drugs, amphotericin B (AmB), azole compounds (e.g.,
fluconazole, itraconazole), these infections are asso-
ciated with significant morbidity and mortality. The
need for a new antifungal agent is due to the alarming
rise in the number of AIDS cases and the subsequent
suppression of the immune system in these patients.
Other reasons that have spurred the development of
new systemic antifungal agents include the increase in
the frequent use of antineoplastic agents and long term
use of antibiotics. The polyenes and azoles, target the
fungal cell membrane, a structure shared by both mam-
malian and fungal cells, and thus these drugs have
inherent toxicity.3ꢀ5
A newer class of cell wall active agents that have been
developed to the point of seeing clinical candidates is
Mulundocandin (MCN or 1), an echinocandins class of
antifungal lipopeptide, was isolated from the culture
broth of a strain of Aspergillus sydowi.10 It exhibited
excellent in vitro activity against Candida species, espe-
cially against C. albicans and C. glabrata isolates.12
Naturally occurring echinocandins have very poor
Keywords: Mulundocandin; Mannich reaction; Antifungal.
§Supplementary data associated with this article can be found, in the
online version at, doi:10.1016/j.bmc.2004.01.015
?
This work was completed at Research Centre, Hoechst Marion
Roussel Limited, Mulund (w), Mumbai 400 080, India.
* Corresponding author. Tel.: +91-022-564-1286; fax: +91-022-564-
1953; e-mail: bansilal@nicholaspiramal.co.in
0968-0896/$ - see front matter # 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2004.01.015

1752
B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
Scheme 1. Mannich reaction on Mulundocandin (1).
Table 1. Structures of Mannich products 2–5a
groups into its HTyr unit in 7–85% yield, with an aim
to increase water solubility through salt formation.
Compd no.
R2
R3
2
–H
–H
2. Chemistry
3
4
5
–H
–H
We have observed that benzyl ether or methyl ether
moieties at Orn-5 position (for nomenclature and num-
bering see Fig. 1) impart increased stability in MCN
without compromising its antifungal activity.20 There-
fore, it was of interest to further modify these analogues
using aminoalkylation reaction, to make water-soluble
derivatives. Such an idea can serve both the objectives,
of making MCN stable and water-soluble. For the pur-
pose of model experiment, it was decided to first intro-
duce aminoalkyl group in MCN itself without Orn-5-
hydroxyl protection.
a Small alphabets a–f is used to identify the protons position.
2.1. Mannich reaction on mulundocandin
Mulundocandin (1) when subjected to Mannich reac-
tion with secondary amine (e.g., pyrrolidine, piperidine)
and formaldehyde solution in the presence or absence of
an acid catalyst in dry dioxane, no Mannich products
were observed; instead a mixture of degraded products
were formed due to the instability of mulundocandin in
acidic conditions. When the reaction was carried out
with paraformaldehyde (instead of formaldehyde solu-
tion) and in the absence of the acid catalyst, Mannich
products were formed but corresponding to the base
degraded MCN. First there appears to occur a ring
opening at Orn-5 position under the basic reaction con-
ditions, and the degraded product 2 on Mannich reac-
tion furnishes the products 3–5 shown in Scheme 1 and
Table 1.
water solubility. To overcome this limitation, a variety
of approaches have been reported in literature. These
include, introduction of secondary amines at Orn-5
position, insertion of chemically or enzymatically labile
groups at phenolic hydroxyl group or introduction of
substituted amino groups at ortho position to phenolic
hydroxyl of HTyr unit.14
We have earlier reported the synthesis of mulundo-
candins carrying C-OR, C–C, C–N and C–H (deoxy)
linkages at Orn-5 position as its chemically stable ana-
logues.15 In this paper, we report the modifications in
aromatic ring of HTyr unit of mulundocandin (MCN)
in an attempt to increase its water solubility. Directing
our efforts towards synthetic methodologies for water-
soluble derivatives, the introduction of aminoalkyl
group into HTyr unit of MCN was attempted by use of
a Mannich reaction. Aminoalkyl group at ortho-posi-
tion of phenolic hydroxyl in echinocandins have typi-
cally been introduced over four steps in less than 40%
overall yield.16 Mannich base formation approach is
used in drug discovery,17ꢀ19 however not for echino-
candins. Here we report Mannich reaction on MCN as
a one step introduction of mono- and di-aminomethyl
After having developed a protocol for synthesis of
Mannich bases, we then chose Orn-5-stabilized mulun-
docandins as a starting material to avoid the degradation
Orn-5-benzyloxy MCN (6) was prepared from mulun-
docandin in 67% yield. Thus, treatment of 6 (1 equiv)
with piperidine (10 equiv) and paraformaldehyde (20
equiv) in anhydrous dioxane at 100–120 ꢁC for 2h, gave
the desired Mannich product 8 in 28% yield. Having

B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
1753
established a successful synthetic protocol, a variety of
Mannich bases were prepared using various alicyclic
and aliphatic secondary amines as the reactants as
shown in Scheme 2 and Table 2. The stoichiometric
ratios of the reactants used (starting compound: sec-
ondary amine: paraformaldehyde), the solvent, the time
required for completion of reaction, the yield, the melt-
ing points, molecular formula and molecular weights of
Mannich reaction products are summarized in Table 3
(see the supplement material)
such as dibenzylamine and N-(t-butyl)benzylamine, it
took 18–24 h, for the reaction to complete.
Mannich reactions, with different secondary amines,
gave a mixture of mono- and di-substituted Mannich
products. Even with one equivalent of secondary amine,
a mixture of mono- and di-substituted Mannich pro-
ducts were formed along with some unreacted starting
compound. The formation of mono- and di-Mannich
products indicates that the reaction may not be pro-
ceeding in a stepwise manner. The reaction rate of
starting material and that of the mono-Mannich pro-
duct does not differ much; hence, simultaneous forma-
tion of mono and di-Mannich products can be expected.
Therefore, in order to drive the reaction to completion,
excess reagents were used. Purification of the reaction
products of these large molecules is the major part of
the experimental. Relatively non-polar Mannich com-
pounds were purified by flash chromatography over
silica gel, polar Mannich compounds were purified by
HPLC over an RP-18 column eluting with CH3CN/H2O
Depending upon the type of secondary amine used, the
time required for the completion of Mannich reaction
ranged from 2–24 h. The Mannich products 8–35 were
obtained by treating the starting compound, a second-
ary amine and paraformaldehyde in the ratio 1:10:20, in
anhydrous dioxane at 80–120 ꢁC for 2–24 h. The reaction
yields ranged from 7–85% depending upon the second-
ary amine involved and the reaction time (Table 3).
With secondary amines like 4-piperidinopiperidine and
pyrrolidine, it took 2.5–4 h, while with hindered amines
Scheme 2. Mannich reaction on Orn-5-benzyloxy Mulundocandin (6).

1754
B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
Table 2. Structures of Mannich products 8–35a
Table 2( continued)
Compd
R2
R3
Compd
R2
R3
8
–H
20
–H
9
–H
21
22
23
–H
–H
CH2dNðCH2ePhÞ2
10
11
–H
24
25
26
27
28
29
30
31
–H
–H
12
13
14
15
16
17
18
19
–H
–H
–H
–H
–H
–H
–H
(continued on next page)

B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
1755
integrating to four protons. The compound 11 (mol.
wt.=1290.527), showed (M+Na)+ at 1312.6 and the
base peak at 1132.6 in the ESI MS spectrum.
Table 2( continued)
Compd
R2
R3
The di-Mannich compound 12, showed 1H NMR peaks
similar those of compound 11, with double the integra-
tion for the aminomethyl resonance (ArCH2NRR1) in
aromatic as well as in aliphatic region, and with dis-
0
appearance of the doublet peak due to D8 -H and slight
0
up field shift of the D7 -H. Due to the symmetrical nat-
32
33
34
35
0
ure of disubstituted phenolic ring, the D7-H and D7 -H
–H
–H
have identical electronic environment and appeared as a
singlet at d 7.16. In ESI MS of compound 12 (mol.wt.=
1482.76), the (M+Na)+ peak appeared at 1504.9.
ESI MS (ES+) spectrum of most of the Mannich com-
pounds on Orn-5-benzyloxy MCN showed 1132.5 as the
base peak for the mono-Mannich products and 1144.5
as the base peak for the di-Mannich products. This is
due to the cleavage of C–N bond of aminomethyl group
to generate stable benzylic radical cation.
Use of imidazole, as the secondary amine in the Man-
nich reaction using the same reaction conditions as for
the other amines, with Orn-5-benzyloxy MCN, did not
give any Mannich product. However, when anhydrous
dimethylformamide was used as the solvent instead of
anhydrous dioxane, and Orn-5-benzyloxy MCN, imi-
dazole and paraformaldehyde were reacted a ratio of
1:10:20 at 130–140 ꢁC for 15 h, an unexpected product
36 was obtained in 38% yield (Scheme 3). In addition,
the Mannich product 37 corresponding to compound 36
was also obtained in a relatively low yield of 13%, as
shown in Scheme 3. In the above reaction, the first step
appears to be the replacement of HTyr-4-hydroxyl
group by imidazole giving compound 36 as the major
product, followed by Mannich reaction to give the
mono-Mannich compound 37 as a minor product.
a Small alphabets b–n is used to identify the protons position.
gradient. The product purities were analyzed on a Per-
kin Elmer HPLC using YMC pack RP-18 analytical
column (10 m particle size) and an acetonitrile/water
gradient. The Mannich products were characterized by
spectral comparison with the starting compound. Since
most of Mannich compounds have similar 1H NMR
and ESI MS spectral pattern, all of them were char-
acterized, only a representative example is described for
1
its structure elucidation. H NMR, ESI MS and other
spectroscopic data of all Mannich derivatives are
detailed in experimental section.
Compound 7 on treatment with a secondary amine and
paraformaldehyde in the ratio 1:10:20 in anhydrous
dioxane at 80–120 ꢁC for 2–31 h gave mono- and di-
Mannich products 38–46, in 5–15% yield.
1
The comparison of H NMR spectrum of 11 with that
of Orn-5-benzyloxy MCN, showed the disappearance of
two original doublets in aromatic region and the
appearance of three peaks for one proton each. This is
due to the introduction of one aminomethyl group at
ortho-position to phenolic hydroxyl. A multiplet at d
7.28–7.41 integrating for five aromatic protons was
assigned to benzyloxy group (–OCH2Ph) at Orn-5 posi-
tion, a doublet of doublet at d 7.17 (J=8.11 and 1.86
It has been observed earlier that Orn-5-OMe-MCN (47)
has shown improved stability and antifungal activity (in
vitro and in vivo) against Candida albicans, over
mulundocandin. Mannich products 48–51 were synthe-
sized in 60–70% yield, starting from 47, using the gen-
eral procedure described earlier. Starting compound 47
was prepared in 70% yield from MCN as shown in
Scheme 5 and Table 5. The compounds 48–51 were
characterized by spectral methods described earlier.
0
Hz), integrating for one proton, was assigned to D7 -H.
The other aromatic proton of HTyr ring, that is, D7-H,
was merged with four aromatic protons of 1-(2-
flurophenyl) piperazine. The ortho coupled doublet at d
0
6.8 (J=8.11 Hz) was assigned to D8 -H. The iminol
2.2. Biological activity
proton (B7-H) was observed as a doublet at d 5.32
(J=1.8 Hz). The methylene protons of Orn-5-benzyloxy
group (–OCH2Ph) appeared as a singlet at d 4.67,
another singlet at d 3.85 was assigned to two methylene
protons of aminomethyl group (ArCH2NRR1). The
piperazine protons appeared as a multiplet at d 3.18
integrating to four protons (f and g) and another mul-
tiplet at d 2.82 for methylene protons (e and h) also
In vitro: Mannich products were screened in vitro by
standard agar well method against C. albicans (IV) and
A. fumigatus (AF-1), in Sabouraud agar bioassay
plates.21 The zones produced by the MCN analogues
were compared with the zone of mulundocandin and the
results of the assay are given in Table 6 (see the supple-
mentary material). The Mannich compounds (Schemes

1756
B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
Scheme 3. Mannich reaction on 6 using imidazole as secondary amine.
Scheme 4. Mannich reaction on Orn-5- and HTyr-4-dibenzyloxy Mulundocandin (7).

B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
1757
Table 4 (continued)
Table 4. Structures of Mannich products 38–46
Compd
R2
R3
Compd
R2
R3
38
–H
43
44
39
40
41
–H
–H
45
46
–H
CH2dNðCH2ePhÞ2
42
–H
Note: small alphabets c–l is used to identify the protons position.
Scheme 5. Mannich reaction on Orn-5-methyoxy Mulindocandin (47).

1758
B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
Table 5. Structures of Mannich products 48–51a
mens consisted of drug administrating on D0, D+1,
D+2, D+3 and D+4 by either of the routes mentioned
above. After the treatment was over, the animals were
sacrificed after 2days. Paired kidneys were removed
aseptically and cultured for the presence of C. albicans
in sterile saline. The kidneys of each group were homo-
genized and ten fold serial dilutions were prepared. A 10
mL aliquot from each dilution was plated on Sabouraud
agar plate, incubated at 30 ꢁC for 24–48 h and the
CFU’s were determined. The number of CFU per gram
of paired kidneys was calculated. Activity was inter-
preted by the difference shown in the load of organisms
per gram kidney weights of treated group and untreated
control group (for details, see supplementary material
Tables 7 and 8).
Compd
48
R2
R3
–H
49
50
51
–H
Among the compounds tested 25, 34, 35, 43, 46 and 49,
showed one log reduction in CFU and 9, 21, 22, 24, 28,
39, 41, 45, 50, 51 showed similar activity to MCN (at
lower dose) by oral application in Swiss mice (Table 7).
The results of in vivo testing by intraperitoneal route
showed that most of the compounds retained the activ-
ity of parent and the compounds 18, 19, 29, 39 and 51
showed one log reduction in CFU (Table 8). Compound
20 showed three log reduction in CFU and is with sig-
nificant improvement of activity over mulundocandin
and compares well with fluconazole, however com-
pound did not reach the level of activity that is required
to be drug candidate.
a Small alphabets b–m is used to identify the protons position.
1–5 and Table 4) failed to show promising in vitro
activity, with the exception of compound 5, which has
shown 24 mm and 19hd mm zone of inhibition against
C. albicans and A. fumigatus, respectively Table 6 (refer
to the supplementary material).
3. Conclusion
A single step introduction of various aminomethyl
groups in ortho position(s) of phenolic hydroxyl of
HTyr unit of MCN has been achieved in 7–85% yield.
The resultant Mannich bases can be converted in to the
salt form to increase their aqueous solubility, whenever
required. All Mannich compounds 4–51 were isolated in
2.3. In vivo testing of mulundocandin analogues in
C. albicans infected Swiss mice model22
Antifungal activity of MCN analogues was evaluated in
vivo in C. albicans infected Swiss mice after oral and
intraperitoneal administration. The colony formation
unit (CFU) in kidney homogenate after administrating
MCN derivatives were determined and compared with
MCN, a standard antifungal in clinical use (Flucona-
zole) and an untreated control group. The experimental
details of activity evaluation and results are summarized
in Tables 7 and 8 (see supplementary material).
1
>95% purity and were characterized using H NMR,
13C NMR, DEPT-135, ESI MS, UV and IR spectral
techniques. The in vitro screening of Mannich products
against Candida albicans and Aspergillus fumigatus, did
not show improvement in activity whereas, most of
them retained the in vivo activity of parent by oral and
intraperitoneal route. Compound 20, in which case a
three-log reduction in CFU by intraperitoneal injection
was observed, which indicates significant improvement
over MCN and activity compares well with that of flu-
conazole. In conclusion, we have developed a method
for improving the aqueous solubility of mulundo-
candins involving Mannich reaction and in one case
improved activity over the original compound.
Swiss mice, 4–6 weeks old and 18–22 gms body weights
were used for in vivo studies. The inoculum was pre-
pared by culturing C. albicans on Sabouraud agar and
cell density adjusted to 106 cells/mouse for non-lethal
model. Mice were infected by injecting the conidia of C.
albicans through the lateral tail vein, with 0.25 mL of
the inoculum on D0. Treatment commenced at zero h
after infection and continued from D0 to D+4. Control
groups (untreated) consisted of four mice infected but
not treated as the negative control and mice infected
and treated with the standard antifungal fluconazole as
the positive control. Mulundocandin, MCN analogues
and fluconazole were prepared in Tween 80 and distilled
water and tested at 100 mg/kg, 80 mg/kg and 50 mg/kg,
respectively, administered by the oral route and at 20
mg/kg each by the intraperitoneal route. Dosing regi-
4. Experimental
4.1. General remarks
All reagents were of commercial quality and were used
as supplied unless otherwise stated. Yields reported are
isolated yields of the materials. All synthesized mulun-
docandin analogues were >95% pure based on TLC

B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
1759
and HPLC analysis. Dioxan was freshly distilled from
4.3. Experimental procedures
sodium prior to use. Melting points (MP) were recorded
on a Kofler hot-plate apparatus and are uncorrected. 1H
NMR spectra and 13C NMR spectra were recorded at
300 MHz and 75 MHz, respectively, on a Brucker ACP-
300 spectrometer, using CD3OD as solvent, unless
otherwise mentioned. Chemical shifts values are expres-
sed in scale (parts per million) using tetramethylsilane
(TMS) as the internal standard. Coupling constant (J)
were reported in Hertz (Hz). Small alphabets a to n on
4.3.1. 1-(2-(1-(1-(2-(4,5-dihydroxy-1-(12-methyltetra-
decanoyl)-2-azolanylcarboxamido)-3-hydroxybutanoyl)-4-
hydroxy-2- azolanylcarboxamido) - 2,3- dihydroxy -3-(4-
hydroxyphenyl)propylcarboxamido)-3-hydroxypropanoyl)-
3-hydroxy-4-methyl-2-azolanecarboxamide (2) and 1-(2-
(3-(3-(1-azolanylmethyl)-4-hydroxyphenyl)-1-(1-(2-(4,5-
dihydroxy-1-(12-methyltetradecanoyl)-2-azolanylcarbox-
amido)-3-hydroxybutanoyl)-4-hydroxy-2-azola-nylcarbox-
amido)-2,3-dihydroxypropylcarboxamido)-3-hydroxypro-
panoyl)-3-hydroxy-4-methyl-2-azolanecarboxamide (3).
In a 25 mL oven dried round-bottom flask were placed
mulundocandin (1) (0.15 g, 0.148 mmol), pyrrolidin
(0.105 g, 1.48 mmol), paraformaldehyde (0.089 g, 2.97
mmol), anhydrous dioxane (15 mL) and heated under
reflux for 3 h. Reaction progress was monitored by TLC
(25% MeOH/CHCl3). After being cooled to ambient
temperature, the solvent was evaporated under vacuum
to leave a crude residue, which was diluted with water
(100 mL), extracted in n-BuOH (3ꢂ50 mL), n-BuOH
extract were washed with water (100 mL) followed by
brine (100 mL). Combined organic extracts were dried
over anhydrous Na2SO4, filtered and concentrated
under vacuum to furnish crude product. The crude
product was dissolved in minimum amount of MeOH (5
mL), adsorbed on silica gel (1:1 w/w), and was subjected
to flash chromatography over silica gel column. 0–20%
MeOH/CHCl3 was used as 5% step gradient elution.
Evaporation of the appropriate solvent fractions gave
compound 2 (0.057 g, 38%) and 3 (0.041 g, 25.3%).
0
0
R2 and R3 groups and B7, D7, D7 , D8, & D8 in struc-
tures of compounds 3–51 is used to identify the protons
position (Fig. 1). ESI MS were recorded on a Fisons VG
QUATTRO II instrument. IR spectra were recorded as
KBr wafers on a Perkin–Elmer 782: Infrared spectro-
photometer. UV spectra were recorded on Chemito
2500: UV–vis Recording spectrometer. TLC was per-
formed on precoated silica gel aluminium plates con-
taining a fluorescent indicator (1.05554, Silica gel 60
F254, Merck) technique using appropriate solvent sys-
tem for development. Flash chromatography was per-
formed using Acme’s silica gel (230–400 mesh) and
MeOH–CHCl3 mixtures as eluent. Visualization of the
components on TLC plates was achieved either by
exposure to iodine vapor or UV light or by charring
with 50% water-sulfuric acid spray. Perkin Elmer
HPLC (Diode Array Detector 235, Binary LC pump
250) was used for purification (Knauer Eurosphere 100,
RP C-18 column, 250ꢂ16 mm, 10 mm, CH3CN/H2O
gradient as an eluent, flow rate 5-8 mL/min, l=220 and
270 nm) and for checking purity (YMC-Pack, AQ-313
S-5 120A ODS, RP C-18 column, 250ꢂ6 mm, 5 mm,
70% CH3CN/H2O as an eluent, flow rate 1 mL/
min,=220 & 270 nm) of MCN analogues.
Compound 2: Partial 1H NMR: 7.25 (d, 2H, J=7.33 Hz,
0
0
D7 & D7 ), 6.77 (d, 2H, J=8.55 Hz, D8 & D8 ), 5.31 (br
s, 1H, iminol proton). 13C NMR: d 177.19, 176.96,
175.76, 175.32, 175.04, 174.98, 173.34, 173.03, 172.75,
172.28, 159.11, 133.74, 130.50, 116.95, 89.56, 82.45,
77.74, 77.54, 77.26, 76.18, 72.08, 71.85, 71.74, 69.73,
69.57, 63.96, 62.04, 61.93, 61.59, 59.25, 58.58, 58.02,
57.02, 57.21, 54.79, 53.25, 39.64, 39.36, 38.58, 36.80,
36.46, 35.68, 31.89, 31.56, 31.44, 31.36, 31.22, 28.99,
27.01, 26.79, 20.43, 12.54, 11.95. IR (KBr): nmax 3400–
3300, 2910, 1660, 1630, 1540, 1440, 1320 cmꢀ1. ESI MS
(ES+): for C48H77N7O16; calculated: 1008.167; found:
(M+Na)+=1030.6 (base peak). UV (MeOH) lmax (e
Mꢀ1 cmꢀ1): at neutral pH: 210, 225, 275 nm (14226,
10584, 1223); at acidic pH: 210, 224, 277 nm (13951,
10584, 1168).
4.2. Nomenclature of mulundocandin
N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R,12R)-23-
((1S,2S)-1,2-dihydroxy-2-(4-hydroxyphenyl)ethyl)-2,11,
12,15-tetrahydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxy-
methyl-16-methyl-5,8,14,19,22,25-hexaoxoperhydrodi-
azolo[2,1-c:2,1-/][1,4,7,10,13,16] hexaazacyclohenicosin-
9-yl]-12-methyltetradecanamide.
1
Compound 3: Partial H NMR: 7.21 (dd, 1H, J=8.55
0
Hz & 1.9 Hz, D7 ), 7.1 (d, 1H, J=1.9 Hz, D7), 6.85 (d,
0
1H, J=8.55 Hz, D8 ), 5.31 (br, 1H, iminol proton), 3.15
(s, 2H, a), 2.4 (br, 4H, b & e), 1.6-1.7 (m, 4H, c & d).
ESI MS (ES+): for C53H86N8O16; calculated: 1091.30;
found: (M)+=1091.6, 1061.5 (base peak).
4.3.2. 1-(2-(3-(3-(1-azinanylmethyl)-4-hydroxyphenyl)-1-
(1-(2-(4,5-dihydroxy-1-(12-methyltetr-adecanoyl)-2-azo-
lanylcarboxamido)-3-hydroxybutanoyl)-4-hydroxy-2-azo-
lanylcarboxam-ido)-2,3-dihydroxypropylcarboxamido)-3-
hydroxypropanoyl)-3-hydroxy-4-methyl-2-azol-anecarb-
oxamide (4) and 1-(2-(3-(3,5-di(1-azinanylmethyl)-4-
Figure 1. Mulundicandin (1).

1760
B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
hydroxyphenyl)-1-(1-(2-(4,5-dihydroxy-1-(12-methyltetra-
decanoyl)-2-azolanylcarboxamido)-3-hydroxybutanoyl)-
4-hydroxy-2-azolanylcarboxamido)-2,3-dihydroxypropyl-
carboxamido)-3-hydroxypropa-noyl)-3-hydroxy-4-methyl-
2-azolanecarboxamide (5). In a 25 mL oven dried round-
bottom flask were placed mulundocandin (1) (0.12g,
0.119 mmol), piperidine (0.101 g, 1.19 mmol), para-
formaldehyde (0.071 g, 2.38 mmol), anhydrous dioxane
(15 mL) and heated under reflux for 3 h. Reaction pro-
gress was monitored by TLC (25% MeOH/CHCl3).
After being cooled to ambient temperature, the solvent
was evaporated under vacuum to leave a crude residue,
which was then diluted with water (100 mL), extracted
in n-BuOH (3ꢂ50 mL), n-BuOH extract were washed
with water (100 mL) followed by brine (100 mL). Com-
bined organic extracts were dried over anhydrous
Na2SO4, filtered and concentrated under vacuum to give
impure product. Which was then dissolved in minimum
amount of MeOH (5 mL), adsorbed on silica gel (1:1 w/
w), and was subjected to flash chromatography over
silica gel column. 0-20% MeOH/CHCl3 was used as 5%
step gradient elution. Evaporation of the appropriate
solvent fractions gave white compound 4 (0.038 g,
28.89%) and 5 (0.013 g, 9.09%).
was subjected to flash chromatography over silica gel
column. 0–25% MeOH/CHCl3 was used as 5% step
gradient eluent. Evaporation of the appropriate fractions
gave white compound 8 (0.03 g, 27.57%). Partial H
NMR: d 7.28–7.41 (m, 5H, –OCH2Ph), 7.17 (dd, 1H,
0
J=8.32Hz & 1.8 Hz, D7 ), 7.0 (d, 1H, J=1.8 Hz, D7),
1
0
6.78 (d, 1H, J=8.37 Hz, D8 ), 5.31 (d, 1H, J=1.65 Hz, B7),
4.68 (s, 2H, –OCH2Ph), 4.05 (s, 2H, d), 2.7 (m, 4H, e & i),
1.45–1.7 (m, 6H, f, g & h). IR (KBr): nmax 3300–3400,
2920, 1660, 1630, 1540, 1460, 1260 cmꢀ1. ESI MS (ES+):
for C61H94N8O16; calculated: 1195.451; found: (M+
Na)+=1217.5, 1132.5 (base peak). UV (MeOH) lmax
(e Mꢀ1 cmꢀ1): 210, 232, 276 nm (60,230, 33,362,
4381).
4.4. General procedure for the preparation of compound
9–35, 38–46 and 48–51
To a stirred solution of Orn-5-substituted starting com-
pound (6, 7 and 47) (1 equiv) in anhydrous dioxane (10–
40 mL) was slowly added secondary amine (10 equiv),
paraformaldehyde (20 equiv) and heated under reflux
(80–120 ꢁC) for 2–31 h. Reaction progress was mon-
itored by TLC (20% MeOH/CHCl3). The reaction work
up and purification process were similar to the described
for compound 8. Stoichiometric ratios of the starting
compound, secondary amine, paraformaldehyde and
anhydrous dioxane are given in Table 3. Yield, mp,
reaction time, molecular formula and molecular weight
of the Mannich products 9–35, 38–46 and 48–51 are
also given in Table 3.
Compound 4: Partial 1H NMR: d 7.4 (dd, 1H, J=8.3 Hz
0
& 1.3 Hz, D7 ), 7.22 (d, 1H, J=1.3 Hz, D7), 6.90 (d, 1H,
0
J=8.3 Hz, D8 ), 5.31 (br, 1H, iminol proton), 3.15 (s,
2H, a), 2.6 (br, 4H, b & f), 1.6-1.7 (m, 6H, c, d & e). ESI
MS (ES+): for C54H88N8O16; calculated: 1105.327;
found: (M+Na)+=1127.6, 1105.9 (M+, base peak).
UV (MeOH) lmax (e M-1 cmꢀ1): at neutral pH: 210 nm
(58,261).
4.4.1. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S,2S)-2-(3-(1-azolanylmethyl)-4-hydroxyphenyl)-
1,2-dihydroxyethyl)-12-benzyloxy-2,11,15-trihydroxy-6-
((1R)-1-hydroxyethyl)-20-hydroxymethyl-16-methyl-5,8,
14,19,22,25-hexaoxoperhydrodiazolo [2,1-c:2,1-/][1,4,7,
10,13,16]hexaazacyclohenicosin-9-yl]-12-methyltetradeca-
namide (9). Partial 1H NMR: d 7.3–7.4 (m, 5H,
1
0
Compound 5: Partial H NMR: d 7.2(s, 2H, D7 & D7 ),
5.3 (br, 1H, iminol proton), 3.1–3.2(s, 4H, 2 ꢂa), 2.55–
2.62 (br, 8H, 2ꢂb & f), 1.55–1.85 (2ꢂbr, 12H, 2ꢂc, d &
e). ESI MS (ES+): for C60H99N9O16; calculated:
1202.486; found: (M+Na)+=1224.9, 1202.8 (M+, base
peak).
0
OCH2Ph), 7.25 (dd, 1H, J=8.55 Hz & 1.9 Hz, D7 ), 7.15
0
(d, 1H, J=1.9 Hz, D7), 6.85 (d, 1H, J=8.55 Hz, D8 ),
4.3.3. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S,2S)-2-(3-(1-azinanylmethyl)-4-hydroxyphenyl)-
1,2-dihydroxyethyl)-12-benzyloxy-2,11,15-trihydroxy-6-
((1R)-1-hydroxy-ethyl)-20-hydroxymethyl-16-methyl-5,8,
14,19,22,25-hexaoxoperhydrodiazolo[2,1-c:2,1-/] [1,4,7,
10,13,16]hexaazacyclohenicosin-9-yl]-12-methyltetradeca-
namide (8). In a 25 mL oven dried round-bottom flask
were placed Orn-5-benzyloxy mulundocandin (6) (0.1 g,
0.091 mmol), piperidine (0.077 g, 0.91 mmol), paraf-
ormaldehyde (0.0546 g, 1.82mmol), anhydrous dioxane
(10 mL) and heated under reflux for 2h. TLC (20%
MeOH/CHCl3) analysis after 2h showed disappearance
of starting compound. Reaction mixture was cooled to
ambient temperature, the solvent was evaporated under
vacuum to leave a crude residue, which was then diluted
with water (100 mL), extracted with n-BuOH (3ꢂ50
mL), n-BuOH extracts was washed with water (100
mL), followed by brine (100 mL). Combined organic
extracts were dried over anhydrous Na2SO4, filtered and
concentrated under reduced pressure to give crude pro-
duct. Crude product was dissolved in minimum amount
of MeOH (5 mL), adsorbed on silica gel (1:1 w/w) and
5.33 (d, 1H, J=1.65 Hz, B7), 4.65 (s, 2H, –OCH2Ph),
4.12(s, 2H, d), 3.3 (m, 4H, e & h), 2.05 (m, 4H, f & g).
IR (KBr): nmax 3300–3400, 2930, 1650, 1625, 1530,
1450, 1260 cmꢀ1. ESI MS (ES+): for C60H92N8O16;
calculated: 1181.424; found: (M+Na)+=1204.7,
1132.5 (base peak). UV (MeOH) lmax ( M-1 cmꢀ1): at
neutral pH: 207, 231, 280 nm (49,807, 15,214, 3515); at
acidic pH: 206, 230, 283 nm (47,710, 15,214, 3751).
4.4.2. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-2-(3,5-di(1-azolanylmethyl)-4-
hydroxyphenyl)-1,2-dihydroxyethyl)-2,11,15-trihydroxy-
6-((1-R)-1-hydroxyethyl)-20-hydroxymethyl-16-methyl-
5,8,14,19,22,25-hexaoxoperhydrodiazolo [2,1-c:2,1-/][1,4,
7,10,13,16]hexaazacyclohenicosin-9-yl]-12-methyltetra-
1
decanamide (10). Partial H NMR: d 7.28–7.41 (m, 5H,
0
OCH2Ph), 7.09 (s, 2H, D7 & D7 ), 5.33 (br, 1H, B7), 4.68
(s, 2H, OCH2Ph), 4.13 (s, 4H, 2ꢂb), 3.1 (m, 8H, 2ꢂc &
f), 1.95 (m, 8H, 2ꢂd & e). IR (KBr): nmax 3300–3400
(br), 2930, 1650, 1625, 1530, 1450, 1260 cmꢀ1. ESI MS
(ES+): for C65H101N9O16; calculated: 1264.557; found:
(M+Na)+=1287.6, 1144.5 (base peak).

B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
1761
0
4.4.3. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-2-(3-(4-(2-flurophenyl)-1,4-diazi-
nan-1-ylmethyl)-4-hydroxyphenyl)-1,2-dihydroxyethyl)-
2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxy-
methyl-16-methyl-5,8,14,19,22,25-hexaoxoperhydrodia-
zolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-
OCH2Ph, D7, D7 , 2ꢂg, h, i & j), 5.33 (br, 1H, B7), 4.67
(s, 2H, OCH2Ph), 3.87 (s, 4H, 2ꢂb), 3.18 (br, 8H, 2ꢂd
& e), 2.95 (br, 8H, 2ꢂc & f). IR (KBr): nmax 3350–
3450, 2930, 1645, 1630, 1530, 1450, 1260 cmꢀ1. ESI MS
(ES+): for C77H109Cl2N11O16; calculated: 1515.672;
found: (M+Na)+=1538.7, 1144.3 (base peak).
1
yl]-12-methyltetradecanamide (11). Partial H NMR: d
7.28–7.41 (m, 5H, OCH2Ph), 7.17 (dd, 1H, J=8.11 Hz
4.4.7. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-1,2-dihydroxy-2-(4-hydroxy-3-
(4-(3-trifluromethylphenyl)-1,4-diazinan-1-ylmethyl)phen-
yl)ethyl)-2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-
hydroxymethyl-16-methyl-5,8,14,19,22,25-hexaoxoper-
hydrodiazolo[2,1c:2,1/][1,4,7,10,13,16]hexaazacyclohenic-
osin-9-yl]-12-methyltetradecanamide (15). Partial 1H
NMR: 7.28–7.45 (m, 5H, OCH2Ph), 7.18–7.26 (m, 4H,
0
& 1.86 Hz, D7 ), 7.0-7.15 (m, 5H, D7 , i, j, k, & l), 6.8 (d,
0
0
1H, J=8.11 Hz, D8 ), 5.32(d, 1H, J=1.8 Hz, B7), 4.67
(s, 2H, OCH2Ph), 3.85 (s, 2H, d), 3.18 (m, 4H, f & g),
2.82 (m, 4H, e & h). IR (KBr): nmax 3300–3400, 2910,
. ESI MS (ES+):
1640, 1615, 1515, 1490 cmꢀ1
for C66H96FN9O16; calculated: 1290.527; found:
(M+Na)+=1312.6, 1132.6 (base peak). UV (MeOH)
lmax (e M-1 cmꢀ1): 207, 231, 276 nm (41,469, 14,667,
4107).
0
i, j, k & l), 7.15 (dd, 1H, J=8.13 Hz & 1.86 Hz, D7 ), 7.1
0
(d, 1H, J=1.86 Hz, D7), 6.8 (d, 1H, J=8.13 Hz, D8 ),
5.32(d, 1H, J=1.86 Hz, B7), 4.68 (s, 2H, OCH2Ph), 3.8
(s, 2H, d), 2.85 (br, 8H, e, f, g & h). 13C NMR: 176.82,
174.90, 174.23, 174.09, 173.56, 172.72, 170.74, 159.17,
153.73, 153.65, 140.71, 133.76, 133.35, 133.12, 132.93,
131.67, 130.70, 130.08, 129.66, 129.39, 128.44, 124.11,
123.53, 121.13, 117.53, 113.82, 81.45, 77.57, 76.85,
76.57, 72.22, 71.04, 70.68, 69.04, 64.18, 63.26, 62.07,
60.36, 59.16, 57.88, 56.43, 54.67, 54.28, 53.64, 51.89,
39.84, 39.45, 38.56, 37.64, 36.46, 35.96, 31.89, 31.58,
31.47, 31.36, 31.11, 28.99, 27.85, 20.57, 20.46, 12.56, 12.01.
IR (KBr): nmax 3350–3450, 2930, 1660 (br), 1635, 1540,
1455, 1260 cmꢀ1. ESI MS (ES+): for C67H96F3N9O16;
calculated: 1340.535; found: (M+Na)+=1362.6,
1132.6 (base peak). UV (MeOH) lmax (e M-1 cmꢀ1): at
neutral pH: 208, 240, 255 nm (4902, 904, 1609); at acidic
pH: 207, 230, 249 nm (4490, 2660, 1330); at basic pH:
210, 253, 296 nm (5908, 1968, 505).
4.4.4. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-2-(3,5-di(4-(2-flurophenyl)-1,4-
diazinan-1-ylmethyl)-4-hydroxyphenyl)-1,2-dihydroxy-
ethyl)-2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-
hydroxymethyl-16-methyl-5,8,14,19,22,25-hexaoxoper-
hydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohen-
icosin-9-yl]-12-methyltetradecanamide (12). Partial 1H
NMR: d 7.28–7.41 (m, 5H, OCH2Ph), 7.16 (s, 2H, D7 &
0
D7 ), 7.0–7.15 (m, 8H, 2ꢂg, h, i & j), 5.32(d, 1H, J=1.8
Hz, B7), 4.67 (s, 2H, OCH2Ph), 3.9 (s, 4H, 2ꢂb), 3.2(br,
8H, 2ꢂd & e), 2.9 (br, 8H, 2ꢂc & f). IR (KBr): lmax
3300–3400, 2910, 1660, 1620, 1520, 1490, 1440 cmꢀ1
.
ESI MS (ES+): for C77H109F2N11O16; calculated:
1482.763; found: (M+Na)+=1504.9. UV (MeOH)
lmax ( M-1 cmꢀ1): at neutral pH: 207, 235, 278 nm
(40,426, 11,675, 2626); at acidic pH: 206, 232, 280 nm
(39,200, 12259, 2626); at basic pH: 209, 236, 280 nm
(62,142, 24,810, 3444).
4.4.8. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-2-(3-(4-(1,3-diazin-2-yl)-1,4-
diazinan-1-ylmethyl)-4-hydroxyphenyl-1,2-dihydroxy-
ethyl)-2,11,15-tri-hydroxy-6-((1R)-1-hydroxyethyl)-20-
hydroxymethyl-16-methyl-5,8,14,19,22,25-hexaoxo-per-
hydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohen-
icosin-9-yl]-12-methyltetr-adecanamide (16). Partial 1H
NMR: d 8.36 (d, 2H, J=6.78 Hz, j & k), 7.29–7.41 (m,
5H, OCH2Ph), 7.19 (dd, 1H, J=8.01 Hz & 1.86 Hz,
4.4.5. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-2-(3-(4-(2-chlorophenyl)-1,4-
diazinan-1-ylmethyl)-4-hydroxyphenyl)-1,2-dihydroxy-
ethyl)-2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-
hydroxymethyl-16-methyl-5,8,14,19,22,25-hexaoxoper-
hydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohen-
icosin-9-yl]-12-methyltetradecanamide (13). Partial 1H
NMR: d 7.28–7.40, 7.15–7.21, 7.05–7.12 (3ꢂm, 11H,
0
D7 ), 7.08 (d, 1H, J=1.86 Hz, D7), 6.81 (d, 1H, J=8.01
0
Hz, D8 ), 6.65 (t, 1H, J=9.3 Hz & 4.5 Hz, J), 5.31 (d,
0
OCH2Ph, D7, D7 , i, j, k, & l), 6.81 (d, 1H, J=8.01 Hz,
0
D8 ), 5.31 (d, 1H, J=1.86 Hz, B7), 4.67 (s, 2H,
1H, J=1.53 Hz, B7), 4.68 (s, 2H, OCH2Ph), 3.85 (s, 2H,
d), 3.95 (br. 4H, f & g), 2.75 (br, 4H, e & h). IR (KBr):
lmax 3350–3450, 2940, 1660, 1630, 1590 (s), 1550, 1450,
1390, 1365, 1270, 1075 cmꢀ1. ESI MS (ES+): for
OCH2Ph), 3.88 (s, 2H, d), 3.18 (br, 4H, f & g), 2.9 (br,
4H, e & h). IR (KBr): nmax 3350–3450, 2935, 1650, 1630,
1530, 1450, 1260 cmꢀ1. ESI MS (ES+): for C66H96Cl-
N9O16; calculated: 1306.982; found: (M+Na)+=
1329.6, 1132.7 (base peak). UV (MeOH) lmax (e M-1
cmꢀ1): 209, 249, 276 nm (44,379, 8061, 3572).
C64H95N11O16;
calculated:
1274.512;
found:
(M+Na)+=1296.5, 1132.7 (base peak).
4.4.9. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-2-(3,5-di(4-(1,3-diazin-2-yl)-
1,4-diazinan-1-ylmethyl)-4-hydroxyphenyl-1,2-dihydroxy-
ethyl)-2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-
hydroxymethyl-16-methyl-5,8,14,19,22,25-hexaox-oper-
hydrodiazo-lo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohen-
icosin-9-yl]-12-methylte-tradecanamide (17). Partial 1H
NMR: 8.35 (d, 4H, J=7.8 Hz, 2ꢂg & i), 7.26–7.41 (m,
4.4.6. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-2-(3,5-di(4-(2-chlorophenyl)-
1,4-diazinan-1-ylmethyl)-4-hydroxyphenyl)-1,2-dihydroxy-
ethyl)-2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-
hydroxymethyl-16-methyl-5,8,14,19,22,25-hexaoxoper-
hydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohen-
icosin-9-yl]-12-methyltetradecanamide (14). Partial 1H
NMR: 7.28–7.40, 7.15–7.12, 7.06–7.13 (3ꢂm, 15H,
0
5H, OCH2Ph), 7.13 (s, 2H, D7 & D7 ), 6.63 (t, 2H,
J=9.6 Hz, 4.8 Hz, 2ꢂh), 5.31 (br, 1H, B7), 4.68 (s, 2H,

1762
B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
OCH2Ph), 3.9 (s, 4H, 2ꢂb), 3.95 (br., 8H, 2ꢂd & e),
2.75 (br., 8H, 2ꢂc & f). IR (KBr): nmax 3350–3450, 2925,
16-methyl-5,8,14,19,22,25-hexaoxoperhydro-diazolo[2,1-
c:2,1 - /][1,4,7,10,13,16]hexaazacyclohenicosin - 9 - yl] - 12-
methyltetradecanamide (21). Partial 1H NMR:d 7.25–
1660, 1630, 1590, 1550, 1450, 1390, 1360, 1265, 1080 cmꢀ1
.
0
ESI MS (ES+): for C73H107N15O16; calculated: 1450.773;
found: (M+Na)+=1472.7, 1144.6 (base peak).
7.41 (m, 9H, OCH2Ph, 2ꢂh & j), 7.14 (s, 2H, D7 & D7 ),
7.03 (d, 4H, J=8.70 Hz, 2ꢂg & k), 6.88 (tt, 2H, J=7.5
Hz & 1.2Hz, 2 ꢂi), 5.31 (d, 1H, J=1.53 Hz, B7), 4.68 (s,
2H, OCH2Ph), 3.85 (s, 4H, 2ꢂb), 3.87 (br, 8H, d & e),
2.80 (br, 8H, c & f). IR (KBr): nmax 3300–3400 (br),
2910, 1650 (br), 1625, 1525, 1440, 1220 cmꢀ1. ESI MS
(ES+): for C77H111N11O16; calculated: 1446.782; found:
(M+Na)+=1468.8, 1144.6 (base peak). UV (MeOH)
lmax (e M-1 cmꢀ1): at neutral pH: 208, 248, 282 nm
(65,504, 32,883, 4472); at acidic pH: 208, 242, 284 nm
(60,177, 19,072, 3847); at basic pH: 210, 248, 284 nm
(72,574, 25,649, 4932).
4.4.10. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-1,2-dihydroxy-2-(3-(4-(4-fluro-
phenyl)-1,4-diazinan-1-ylmethyl)-4-hydroxyphenyl)ethyl)-
2,11,15-trihyd-roxy-6-((1R)-1-hydroxyethyl)-20-hydroxy-
methyl-16-methyl-5,8,14,19,22,25-hexaoxoperh-ydrodia-
zolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-yl]-
12-methyltetradec-anamide (18). Partial 1H NMR: d
7.28–7.41 (m, 5H, OCH2Ph), 7.18 (dd, 1H, J=8.40 Hz
0
& 1.53 Hz, D7 ), 7.08 (d, 1H, J=1.53 Hz, D7), 7.0 (d,
0
4H, J=8.16 Hz, i, j, k & l), 6.8 (d, 1H, J=8.40 Hz, D8 ),
5.33 (d, 1H, J=1.5 Hz, B7), 4.68 (s, 2H, OCH2Ph), 3.85
(s, 2H, d), 3.20 (br., 4H, f & g), 2.80 (br., 4H, e & h). IR
(KBr): nmax 3350–3450, 2920, 1645, 1615, 1509, 1430,
1225 cmꢀ1. ESI MS (ES+): for C66H96FN9O16; calcu-
lated: 1290.527; found: (M+Na)+=1312.4, 1132.5
(base peak).
4.4.14. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-2-(3-dibenzyl-aminomethyl-4-
hydroxyphenyl)-1,2-dihydroxyethyl)-2,11,15-trihydroxy-6-
((1R)-1-hydro-xyethyl)-20-hydroxymethyl-16-methyl-5,8,
14,19,22,25-hexaoxoperhydrodiazolo[2,1-c:2,1-/][1,4,7,10,
13,16]hexaazacyclohenicosin-9-yl]-12-methyltetradecan-
amide (22). Partial 1H NMR: d 7.28–7.42 (m, 15H,
OCH2Ph, 2ꢂNCH2Ph), 7.17 (dd, 1H, J=8.64 Hz &
4.4.11. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-2-(3,5-di(4-(4-flurophenyl)-1,4-
diazinan-1-ylmethyl)-4-hydroxyphenyl)-1,2-dihydroxy-
ethyl)-2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-
hydroxymethyl-16-methyl-5,8,14,19,22,25-hexaox-oper-
hydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacycloheni-
cosin-9-yl]-12-methyltet-radecanamide (19). Partial 1H
NMR: 7.28–7.41 (m, 5H, OCH2Ph), 7.14 (s, 2H, D7 &
0
2.16 Hz, D7 ), 7.09 (d, 1H, J=2.16 Hz, D7), 6.79 (d, 1H,
0
J=8.64 Hz, D8 ), 5.31 (d, 1H, J=1.53 Hz, B7), 4.68 (s,
2H, OCH2Ph), 3.63–3.7 (2ꢂs, 6H, d & 2ꢂe). 13C NMR:
d 176.83, 174.96, 174.15, 174.08, 173.5, 172.66, 170.62,
158.97, 140.66, 139.11, 134.0, 131.51, 130.44, 130.02,
129.76, 129.67, 129.57, 129.34, 128.86, 124.07, 117.41,
81.46, 77.39, 76.77, 76.48, 72.21, 72.12, 71.05, 70.63, 69.01,
64.09, 63.15, 59.53, 59.24, 57.88, 56.74, 56.36, 53.55,
51.99, 39.80, 39.38, 38.54, 37.60, 36.43, 35.95, 31.87,
31.55, 31.42, 31.36, 31.06, 28.96, 27.83, 20.42, 12.53,
11.98. IR (KBr): nmax 3300–3400, 2910, 1640, 1615, 1515,
1430, 1240 cmꢀ1. ESI MS (ES+): for C70H98N8O16;
calculated: 1307.582; found: (M+Na)+=1330.7, 1132.6
(base peak). UV (MeOH) lmax (e M-1 cmꢀ1): pH: 206,
225, 279 nm (37,234, 8761, 15,135).
0
D7 ),7.0 (d, 8H, J=7.41 Hz, 2ꢂg, h, i & j), 5.33 (d, 1H,
J=1.8 Hz, B7), 4.68 (s, 2H, OCH2Ph), 3.85 (s, 4H,
2ꢂb), 3.22 (br, 8H, 2ꢂd & e), 2.83 (br, 8H, 2ꢂc & f). IR
(KBr): nmax 3350–3450, 2920, 1645, 1615, 1509, 1430,
1225 cmꢀ1. ESI MS (ES+): for C77H109F2N11O16; cal-
culated: 1482.763; found: (M+Na)+=1504.8, 1144.7.
UV (MeOH) lmax (e M-1 cmꢀ1): 210, 233, 285 nm
(75,574, 36,321, 8063).
4.4.12. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-1,2-dihydroxy-2-(4-hydroxy-3-
(4-phenyl)-1,4-diazinan-1-ylmethyl)phenyl)ethyl)-2,11,15-
trihydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-16-
methyl-5,8,14,19,22,25-hexaoxoperhydrodiaz-olo[2,1-c:2,
1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-yl]-12-methyl-
tetradecanamide (20). Partial 1H NMR: 7.28–7.41(m,
5H, OCH2Ph), 7.21–7.27 (m, 2H, j & l), 7.19 (dd, 1H,
4.4.15. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S,2S)-2-(3-(4-benzyl-1,4-diazinan-1-ylmethyl)-4-
hydroxyphenyl)-1,2-dihydroxyethyl)-12-benzyloxy-2,11,15-
trihydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-
16-methyl-5,8,14,19,22,25-hexaoxoperhydrodiaz-olo[2,1-
c:2,1 - /][1,4,7,10,13,16]hexaazacyclohenicosin - 9 - yl] - 12-
1
methyltetradecanamide (23). Partial H NMR: d 7.28–
7.43 (m, 10H, OCH2Ph, –NCH2Ph), 7.18 (dd, 1H,
0
J=8.40 Hz & 2.16 Hz, D7 ), 7.08 (d, 1H, J=2.16 Hz,
0
J=8.64 Hz & 1.86 Hz, D7 ), 7.03 (d, 1H, J=1.86 Hz,
0
D7), 6.78 (d, 1H, J=8.64 Hz, D8 ), 5.31 (d, 1H, J=2.04
D7), 7.02(d, H2 ,
J=8.40 Hz, i & m), 6.90 (t, 1H,
0
J=7.20 Hz, k), 6.80 (d, 1H, J=8.40 Hz, D8 ), 5.31 (d,
1H, J=2.25 Hz, B7), 4.68 (s, 2H, OCH2Ph), 3.85 (s, 2H,
d), 3.27 (br, 4H, f & g), 2.80 (br, 4H, e & h). IR (KBr):
nmax 3300–3400, 2910, 1645, 1610, 1515, 1430, 1215
cmꢀ1. ESI MS (ES+): for C66H97N9O16; calculated:
1272.537; found: (M+Na)+=1294.7, 1272.4, 1132:5
(base peak). UV (MeOH) lmax (e M-1 cmꢀ1): 207, 230,
246, 279 nm (47,454, 14,338, 12,697, 3314).
Hz, B7), 4.68 (s, 2H, –OCH2Ph), 3.58–3.62(2 ꢂs, 4H, d
& i), 3.18, 2.68 (2ꢂt, 8H, e, f, g & h). IR (KBr): nmax
3300–3400, 2930, 1650, 1625, 1520, 1450, 1390, 1260,
1070 cmꢀ1. ESI MS (ES+): for C67H99N9O16; calcu-
lated: 1286.563; found: (M+Na)+=1309.6, 1132.5
(base peak). UV (MeOH) lmax (e M-1 cmꢀ1): at neutral
pH: 208, 229, 280 nm (42,242, 12,359, 2648); at acidic
pH: 207, 228, 284 nm (39,130, 11,035, 3089); at basic
pH: 208, 230 nm (43,055, 6201).
4.4.13. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-1,2-dihydroxy-2-(4-hydroxy-3,5-
di(4-phenyl)-1,4-diazinan-1-ylmethyl)phenyl)ethyl)-2,11,
15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-
4.4.16. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S,2S)-2-(3-(4-(2-azinyl)-1,4-diazinan-1-ylmethyl)-4-
hydroxyphenyl-1,2-dihydroxyethyl)-12-benzyloxy-2,11,15-

B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
1763
trihydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-
16-methyl - 5,8,14,19,22,25 - hexaoxoperhydrodiazolo[2,1-
c:2,1 - /][1,4,7,10,13,16]hexaazacyclohenicosin - 9 - yl] - 12-
methyltetradecanamide (24). Partial 1H NMR: d 8.1–
8.16 (m, 1H, l), 7.6 (m, 1H, J), 7.3–7.45 (m, 5H, –
0
OCH2Ph), 7.18 (dd, 1H, J=8.37 Hz & 1.41 Hz, D7 ),
7.08 (d, 1H, J=1.41 Hz, D7), 6.89 (m, 1H, i), 6.8 (d, 1H,
0
J=8.37 Hz, D8 ), 6.75 (m, 1H, k), 5.31 (d, 1H, J=1.53
(br, 8H, 2ꢂd & e), 2.73 (br, 8H, 2ꢂc & f). IR (KBr):
max 3350–3450, 2920, 1650, 1610, 1540, 1510, 1440, 1385,
n
1230, 1070 cmꢀ1. ESI MS (ES+): for C75H109N13O16;
calculated: 1448.457; cound: (M+Na)+=1470.6, 1449.6.
UV (MeOH) lmax (e M-1 cmꢀ1): at neutral pH: 208, 237,
262 nm (75379, 10,463, 41,034); at acidic pH: 206, 240,
280 nm (54,780, 3816, 48,050); at basic pH: 212, 235,
258 nm (101,025, 10,340, 40,377).
Hz, B7), 4.68 (s, 2H, –OCH2Ph), 3.8 (s, 2H, d), 3.6 (m,
4H, f & g), 2.72 (m, 4H, e & h). IR (KBr): nmax 3300–
3400, 2930, 1640, 1620, 1520, 1430, 1375, 1235, 1060
cmꢀ1. ESI MS (ES+): for C65H96N10O16; calculated:
1273.524; found: (M+Na)+=1295.7, 1273.7, 1132.5.
UV (MeOH) lmax (e Mꢀ1 cmꢀ1): 208, 248, 299 nm
(43,844, 27,725, 5899).
4.4.20. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S,2S)-2-(3-(4-(1-azinanyl)-1-azin-anylmethyl)-4-
hydroxyphenyl)-1,2-dihydroxyethyl)-12-benzyloxy-2,11,15-
trihydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-
16-methyl-5,8,14,19,22,25-hexaoxoperhydrodiaz-olo[2,1-
c:2,1 - /][1,4,7,10,13,16]hexaazacyclohenicosin - 9 - yl] - 12-
methyltetradecanamide (28). Partial 1H NMR: 7.28–
7.45 (m, 5H, –OCH2Ph), 7.18 (dd, 1H, J=8.64 Hz &
4.4.17. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-1,2-dihydroxy-2-(4-hydroxy-3-
(4-(4-methylphenyl)-1,4-diazinan-1-ylmethyl)phenyl)ethyl)-
2,11,15-trih-ydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxy-
methyl-16-methyl-5,8,14,19,22,25-hexaoxoperhydrodia-
zolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-
0
1.86 Hz, D7 ), 7.06 (d, 1H, J=1.86 Hz, D7), 6.8 (d, 1H,
0
J=8.64 Hz, D8 ), 5.02(d, 1H, J=1.86 Hz, B7), 4.68 (s,
2H, –OCH2Ph), 3.78 (s, 2H, d), 2.89–3.28 (m, 9H, e, i, g,
j & n), 1.7–1.9 (m, 10H, f, h, k, l & m). IR (KBr): nmax
3300-3400, 2940, 1660, 1635, 1518, 1460, 1370 cmꢀ. 1 ESI
MS (ES+): for C66H103N9O16; calculated: 1278.584;
found: (M+Na)+=1300.5, 1132.4 (base peak). UV
(MeOH) lmax (e M-1 cmꢀ1): 208, 225, 279 nm (46,029,
13,780, 1619).
1
yl]-12-methyltetrad-ecanamide (25). Partial H NMR: d
7.29–7.43 (m, 5H, –OCH2Ph), 7.18 (dd, 1H, J=8.64 Hz
0
& 1.53 Hz, D7 ), 7.06–7.12(m, 3H, D7, j & k), 6.93 (d,
0
2H, J=8.64 Hz, i & l), 6.79 (d, 1H, J=8.64 Hz, D8 ),
5.31 (d, 1H, J=1.53 Hz, B7), 4.68 (s, 2H, –OCH2Ph),
3.81 (s, 2H, d), 3.2(br, 4H, f & g), 2.78 (br, 4H, e & h),
2.38 (s, 3H, Ar-CH3). IR (KBr): nmax 3300–3400, 2930,
1640, 1620, 1520, 1430, 1375, 1235, 1060 cmꢀ1. ESI MS
(ES+): for C67H99N9O16; calculated: 1286.583; found:
(M+Na)+=1309.6, 1132.5. UV (MeOH) lmax (e Mꢀ1
cmꢀ1): 209, 230, 247, 279 nm (71,176, 61,764, 20,808,
5147).
4.4.21. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-1,2-dihydroxy-2-(3-(4-(2,6-di-
methylphenyl)-1,4-diazinan-1-ylmethyl)-4-hydroxyphenyl)-
ethyl)-2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-
hydroxymethyl-16-methyl-5,8,14,19,22,25-hexaoxoper-
hydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohen-
icosin-9-yl]-12-methyltetradecanamide (29). Partial 1H
NMR: d 7.29–7.42 (m, 5H, –OCH2Ph), 7.18 (dd, 1H,
0
4.4.18. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-1,2-dihydroxy-2-(4-hydroxy-
3,5-di(4-(4-methylphenyl)-1,4-diazinan-1-ylmethyl)phenyl)-
ethyl)-2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-
hydroxymethyl - 16 - methyl - 5,8,14,19,22,25 - hexaoxoper
hydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohen-
icosin-9-yl]-12-methyltet-radecanamide (26). Partial 1H
NMR: d 7.29–7.43 (m, 5H, –OCH2Ph), 7.14 (s, 2H, D7
J=8.55 Hz & 1.32Hz, D7 ), 7.09 (d, 1H, J=1.32Hz,
D7), 6.9-7.03 (m, 3H, i, j & k), 6.81 (d, 1H, J=8.55 Hz,
0
D8 ), 5.31 (br, 1H, B7), 4.68 (s, 2H, –OCH2Ph), 3.91 (s,
2H, d), 3.2(br, 4H, f & g), 2.82 (br, 4H, e & h), 2.38 (s,
6H, 2ꢂAr-CH3). 13C NMR: d 176.82, 174.95, 174.20,
174.03, 173.53, 172.67, 170.63, 159.28, 149.74, 140.71,
138.70, 133.76, 130.98, 130.84, 130.06, 129.64, 129.36,
127.85, 127.35, 122.66, 117.51, 81.42, 77.57, 76.79,
76.54, 72.22, 71.04, 70.74, 69.04, 64.16, 63.24, 62.09,
59.25, 57.91, 56.32, 55.62, 54.98, 54.73, 53.59, 51.94,
51.11, 39.81, 39.45, 38.56, 37.61, 36.46, 35.93, 31.89,
31.58, 31.47, 31.36, 31.11, 28.99, 27.85, 20.65, 20.51,
20.46, 12.56, 11.98. IR (KBr): nmax 3300–3400, 2935,
1660, 1625, 1530, 1450, 1385, 1260, 1070 cmꢀ1. ESI MS
(ES+): for C68H101N9O16; calculated: 1300.590; found:
(M+Na)+=1322.5, 1132.5 (base peak). UV (MeOH)
lmax ( M-1 cmꢀ1): 208, 226, 267 nm (37,979, 14,394,
2709).
0
& D7 ), 7.1 (d, 4H, J=8.64 Hz, 2ꢂh & i), 6.92(d, 4H,
J=8.64 Hz, 2ꢂg & j), 5.33 (d, 1H, J=1.86 Hz, B7), 4.68
(s, 2H, –OCH2Ph), 3.82(s, 4H, 2 ꢂb), 3.21 (br, 8H, 2ꢂd
& e), 2.73 (br, 8H, 2ꢂc & f), 2.29 (s, 6H, 2ꢂAr-CH3).
IR (KBr): nmax 3350-3450, 2940, 1655, 1630, 1519, 1450,
1385, 1060 cmꢀ1. ESI MS (ES+): for C79H115N11O16;
calculated: 1474.835; found: (M+Na)+=1496.8, 1474.6,
1144.4 (base peak). UV (MeOH) lmax ( M-1 cmꢀ1): 210,
242, 284 nm (62,037, 26,909, 5900).
4.4.19. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-2-(3,5-di(4-(4-azinyl)-1,4-diazi-
nan-1-ylmethyl)-4-hydroxyphenyl-1,2-dihydroxyethyl)-
2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxy-
methyl-16-methyl-5,8,14,19,22,25-hexaoxoperhydrodia-
zolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-
yl]-12-methyltetradecanamide (27). Partial 1H NMR:
8.15–8.22 (m, 4H, 2ꢂh & i), 7.25–7.43 (m, 5H, –OCH2Ph),
4.4.22. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-1,2-dihydroxy-2-(3,5-di(4-(2,6-
dimethylphenyl)-1,4-diazinan-1-ylmethyl)-4-hydroxyphen-
yl)ethyl)-2,11, 15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-
hydroxymethyl-16-methyl-5,8,14,19,22,25-hexaoxoperhy-
drodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohenico-
sin-9-yl]-12-methyltetradecanamide (30). Partial 1H
NMR: d 7.28–7.42 (m, 5H, –OCH2Ph), 7.21 (s, 2H, D7
0
7.14 (s, 2H, D7 & D7 ), 7.0 (m, 4H, 2ꢂg & j), 5.31 (br,
0
1H, B7), 4.68 (s, 2H, –OCH2Ph), 3.81 (s, 4H, 2ꢂb), 3.65
& D7 ), 6.98–7.2(m, 6H, 2 ꢂg, h & i), 5.33 (br, 1H, B7),

1764
B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
4.68 (s, 2H, –OCH2Ph), 4.11 (s, 4H, 2ꢂb), 3.29 (br, 8H,
2ꢂd & e), 3.05 (br, 8H, 2ꢂc & f), 2.40 (s, 12H, 4ꢂAr-
CH3). IR (KBr): nmax 3350–3450 (br), 2920, 1670, 1630,
1535, 1460, 1390, 1220, 1070 cmꢀ1. ESI MS (ES+):
for C81H119N11O16; calculated: 1502.889; found:
(M+Na)+=1525.6, 1144.6 (base peak). UV (MeOH)
10H, –OCH2Ph & –NCH2Ph), 7.05 (dd, 1H, J=8.37 Hz
0
& 1.41 Hz, D7 ), 6.95 (d, 1H, J=1.41 Hz, D7), 6.55 (d,
0
1H, J=8.37 Hz, D8 ), 5.32(d, 1H, J=2.1 Hz, B7), 4.68
(s, 2H, –OCH2Ph), 4.09 (s, 2H, d), 3.89 (s, 2H, f), 1.42
(s, 9H, 3ꢂe or –C(CH3)3). IR (KBr): nmax 3300–3400,
2920, 1660, 1625, 1525, 1440, 1375, 1250, 1070 cmꢀ1
.
l
8513, 5187).
max ( M-1 cmꢀ1): 211, 226, 257, 282 nm (58,787, 26,424,
ESI MS (ES+): for C67H100N8O16; calculated:
1273.565; found: (M+Na)+=1296.6, 1132.5 (base
peak). UV (MeOH) lmax ( M-1 cmꢀ1): 210, 226, 280 nm
(76,304, 28,418, 4257).
4.4.23. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-1,2-dihydroxy-2-(4-hydroxy-3-
(4-(1-phenylethyl)-1,4-diazinan-1-ylmethyl)phenyl)ethyl)-
2,11,15-trihy-droxy-6-((1R)-1-hydroxyethyl)-20-hydroxy-
methyl-16-methyl-5,8,14,19,22,25-hexaoxoper-hydrodia-
zolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-
4.4.26. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S,2S)-2-(3-benzyl(isopropyl)amin-omethyl-4-hydroxy-
phenyl)-1,2-dihydroxyethyl)-12-benzyloxy-2,11,15-tri-
hydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-16-
methyl-5,8,14,19,22,25-hexaoxoperhydrodiazolo[2,1-c:2,1-/]
[1,4,7,10,13,16]hexaazacyclohenicosin- 9-yl]- 12-methyl-
1
yl]-12-methyltetradecanamide (31). Partial H NMR: d
7.28–7.45 (m, 10H, –OCH2Ph & –CH(CH3)Ph), 7.17
1
0
(dd, 1H, J=8.55 Hz & 1.32Hz, D7 ), 7.03 (d, 1H,
0
J=1.32Hz, D7), 6.77 (d, 1H, J=8.55 Hz, D8 ), 5.31 (d,
tetradecanamide (34). Partial H NMR: d 7.28–7.45 (m,
10H, –OCH2Ph & –NCH2Ph), 7.16 (dd, 1H, J=8.55 Hz
0
& 1.98 Hz, D7 ), 7.05 (d, 1H, J=1.98 Hz, D7), 6.74 (d,
0
1H, J=8.55 Hz, D8 ), 5.32(br, 1H, B7), 4.68 (s, 2H,
1H, J=1.98 Hz, B7), 4.68 (s, 2H, –OCH2Ph), 3.75 (s,
2H, d), 3.8 (q, 1H, J=7.89 Hz, i), 2.6-2.79 (m, 8H, e, f,
g & h), 1.45 (d, 3H, J=7.89 Hz, j). 13C NMR: 176.80,
174.92, 174.08, 173.50, 172.66, 170.65, 159.20, 144.93,
144.51, 140.70, 133.68, 130.41, 130.18, 130.05, 129.63,
129.34, 129.15, 129.08, 123.63, 117.41, 81.43, 77.49,
76.81, 76.55, 72.18, 72.12, 71.02, 70.66, 69.01, 67.13,
64.13, 63.19, 62.09, 59.21, 57.85, 56.43, 54.68, 54.29,
53.58, 52.38, 51.93, 51.41, 50.99, 46.62, 39.80, 39.41,
38.54, 37.60, 36.43, 35.95, 31.87, 31.55, 31.45, 31.36,
31.10, 28.96, 27.83, 20.94, 20.45, 12.56, 11.98. IR (KBr):
nmax 3300–z3400, 2920, 1660, 1625, 1530, 1455, 1390,
1260, 1070 cmꢀ1. ESI MS (ES+): for C68H101N9O16;
calculated: 1300.590; found: (M+Na)+=1323.6, 1300.6,
1132.5. UV (MeOH) lmax (e M-1 cmꢀ1): 206, 223, 279
nm (47,065, 14,834, 1881).
OCH2Ph), 3.9, 3.65 (2ꢂs, 4H, d & h), 3.1 (m, 1H, e),
1.22 (m, 6H, f & g). IR (KBr): nmax 3300–3400, 2935,
1680–1625, 1540, 1450, 1385, 1260, 1075 cmꢀ1. ESI MS
(ES+): for C66H98N8O16; calculated: 1259.538; found:
(M+Na)+=1281.8, 1132.4 (base peak). UV (MeOH)
lmax (e M-1 cmꢀ1): 207, 231, 280 nm (58,232, 10,790,
2997).
4.4.27. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-2-(3,5-di (benzyl(iso-propyl)-
aminomethyl-4-hydroxyphenyl)-1,2-dihydroxyethyl)-12-
benzyloxy-2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-
20-hydroxymethyl-16-methyl-5,8,14,19,22,25-hexaoxoper-
hydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohen-
icosin-9-yl]-12-methyltetradecanamide (35). Partial 1H
4.4.24. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S,2S)-1,2-dihydroxy-2-(3,5-di(4-(1-
phenylethyl)-1,4-diazinan-1-ylmethyl)-4-hydroxyphenyl)-
ethyl)-2,11,15-tri-hydroxy-6-((1R)-1-hydroxyethyl)-20-
hydroxymethyl-16-methyl-5,8,14,19,22,25-hexaoxo-per-
hydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohen-
icosin-9-yl]-12-methyltetradecanamide (32). Partial 1H
NMR:
d
7.28–7.43 (m, 15H, –OCH2Ph
&
2ꢂ–
0
NCH2Ph), 7.03 (s, 2H, D7 & D7 ), 5.33 (br, 1H, B7), 4.68
(s, 2H, –OCH2Ph), 3.87, 3.63 (2ꢂs, 8H, 2ꢂb & f), 3.0
(m, 2H, 2ꢂc), 1.2–1.3 (m, 12H, 2ꢂe & d). IR (KBr):
nmax 3400–3500, 2945, 1680–1630, 1540, 1460, 1385,
1260, 1080 cmꢀ1. ESI MS (ES+): for C77H113N9O16;
Calculated: 1420.784; Found: (M)+=1420.9, 1144.9
(base peak). UV (MeOH) lmax (e M-1 cmꢀ1): at neutral
pH: 207, 227, 282 nm (67,687, 10,661, 1465); at acidic
pH: 206, 225, 284 nm (55,658, 11,973,1995); at basic
pH: 210 nm (118,206).
NMR:
d
7.22–7.40 (m, 15H, –OCH2Ph
&
0
2ꢂ-CH(CH3)Ph), 6.84 (s, 2H, D7 & D7 ), 5.33 (br, 1H,
B7), 4.45 (s, 2H, –OCH2Ph), 3.52(s, 4H, 2 ꢂb), 3.42(q,
2H, 7.8 Hz, 2ꢂg), 2.3–2.55 (m, 16H, 2ꢂc, d, e & f), 1.28
(d, 6H, 7.8 Hz, 2ꢂh). IR (KBr): nmax 3300–3450, 2920,
1655, 1625, 1525, 1450, 1385, 1255, 1070 cmꢀ1
.
4.5. Procedure for the preparation of compounds
36 and 37
ESI MS (ES+): for C81H119N11O16; calculated:
1502.889; found: (M+Na)+=1525.7, 1502.8, 1144.4.
UV (MeOH) lmax (e M-1 cmꢀ1): at neutral pH: 205, 219,
284 nm (50,300, 7314, 1833); at acidic pH: 205, 225, 286
nm (=41,733, 9769, 1803); at basic pH: 211 nm
(110,020).
To a stirred solution of Orn-5-benzyloxymulundocandin
(6) (0.2g, 0.182mmol) in anhydrous N,N-dimethylfor-
mamide (10 mL) was added imidazole (0.122 g, 1.8
mmol), paraformaldehyde (0.108 g, 3.6 mmol) and
heated under reflux for 15 h. Reaction progress was
monitored by TLC (20% MeOH/CHCl3). The reaction
work up and purification procedure were similar to that
of compound 8. Yield of the white solid 36 (0.08 g,
38.25%) and 37 (0.03 g, 13.42%).
4.4.25. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S,2S)-2-(3-benzyl(ter.butyl)aminomethyl-4-hydroxy-
phenyl)-1,2-dihydroxyethyl)-12-benzyloxy-2,11,15-tri-
hydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-16-
methyl-5,8,14,19,22,25-hexaoxoperhydrodiazolo[2,1-c:2,1-/]
[1,4,7,10,13,16]hexaazacyclohenicosin-9-yl]-12-methyl-
4.5.1. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S)-2-(1H-1,3-diazol-1-yl)-1-hydroxy-
1
tetradecanamide (33). Partial H NMR: d 7.15–7.45 (m,

B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
1765
2-(4-hydroxyphenyl)ethyl)-2,11,15-trihydroxy-6-((1R)-1-
hydroxyethyl)-20-hydroxymethyl-16-methyl-5,8,14,19,
22,25-hexaoxoperhydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]
2.98 (m, 8H, 2ꢂe & i), 1.98 (m, 12H, 2ꢂf, g & h). IR
(KBr): nmax 3300–3400, 2910, 1650, 1625, 1530, 1440,
1250 cmꢀ1. ESI MS (ES+): for C74H111N9O16; calcu-
lated: 1382.735; found: (M+Na)+=1404.8 (base peak),
1382.6. UV (MeOH) lmax ( M-1 cmꢀ1): 209, 234, 290 nm
(46,021, 9127, 3989).
hexaazacyclohenicosin-9-yl]-12-methyltetradecanamide
1
(36). Partial H NMR: d 7.89 (br, 1H, g), 7.72(s, 2H,
e
& f), 7.22 (m, 5H, –OCH2Ph), 7.18, 6.98 (2ꢂd, 2H,
0
J=8.55 Hz, D7 & D7 ), 6.8 (2ꢂd, 2H, J=8.55 Hz, D8 &
0
D8 ), 5.35 (d, 1H, J=1.9 Hz, B7), 4.63 (s, 2H,
4.5.5. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S)-2-(3-(1-azolanylmethyl)-4-hydroxyphenyl)-2-
benzyloxy - 1 - hydroxyethyl) - 12 - benzyloxy - 2,11,15 - tri-
hydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-16-
methyl-5,8,14,19,22,25-hexaoxoperhydrodiazolo[2,1-c:2,1-/]
[1,4,7,10,13,16]hexaazacyclohenicosin-9-yl]-12-methyltetra-
decanamide (40). Partial 1H NMR: d 7.25–7.41 (m, 10H,
–OCH2Ph). 13C NMR: d 176.85, 174.95, 174.56, 174.09,
173.70, 172.28, 170.49, 159.87, 159.62, 140.71, 139.26,
138.98, 138.11, 131.81, 131.23, 130.56, 130.06, 129.64,
129.36, 121.38, 120.43, 120.07, 117.65, 117.28, 81.39,
76.82, 74.17, 72.22, 71.04, 70.71, 69.15, 66.30, 64.74,
64.02, 63.12, 62.99, 59.33, 57.88, 56.85, 56.49, 54.81,
53.59, 52.00, 45.36, 39.87, 39.53, 38.56, 37.61, 36.46,
35.88, 31.89, 31.58, 31.44, 31.36, 31.08, 28.99, 27.85,
20.43, 20.29, 12.56, 12.01. IR (KBr):nmax 3300–3400, 2930,
1650, 1625, 1520, 1455, 1390, 1225, 1070 cmꢀ1. ESI MS
(ES+): for C58H85N9O15; calculated: 1148.355; found:
(M+Na)+=1170.6 (base peak), 1148.5. UV (MeOH)
0
2ꢂ–OCH2Ph), 7.25 (dd, 1H, J=8.5 Hz & 1.9 Hz, D7 ),
7.14 (d, 1H, J=1.9 Hz, D7), 6.87 (d, 1H, J=8.5 Hz,
0
D8 ), 5.31 (br, 1H, B7), 4.67 (s, 4H, 2ꢂ–OCH2Ph), 4.13
(s, 2H, d), 3.35 (m, 4H, e & h), 2.1 (m, 4H, f & g). IR
(KBr): nmax 3300–3400, 2925, 1650, 1620, 1535, 1450,
1250 cmꢀ1. ESI MS (ES+): for C67H98N8O16; calcu-
lated: 1271.549, found: (M+Na)+=1293.6 (base peak).
UV (MeOH) lmax (Mꢀ1 cmꢀ1): 211, 230, 278 nm
(64,015, 27056, 6845).
l
max (e M-1 cmꢀ1): 205, 226, 273 nm (23152, 8180, 1929).
4.5.2. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S)-2-(1H-1,3-diazol-1-yl)-2-(3-(1H-
1,3-diazol-1-ylmethyl)-4-hydroxyphenyl)-1-hydroxyethyl)-
2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxy-
methyl-16-ethyl-5,8,14,19,22,25-hexaoxoperhy-drodia-
zolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-
4.5.6. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S)-2-benzyloxy-2-(3,5-di(1-azolanyl-
methyl)-4-hydroxyphenyl)-1-hydroxyethyl)-2,11,15-tri-
hydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-16-
methyl-5,8,14,19,22,25-hexaoxoperhydrodiazolo[2,1-c:2,
1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-yl]-12-methyl-
1
yl]-12-methyltetradecanamide (37). Partial H NMR: d
7.8–7.7 (m, 2H, 2ꢂb), 7.42–7.28 (m, 5H, OCH2Ph),
1
0
6.99–7.1, 7.19 (2ꢂbr, 6H, 2ꢂc & d+D7 & D7 ), 6.82(d,
tetradecanamide (41). Partial H NMR: d 7.28–7.41 (m,
0
1H, J=8.13 Hz, D8 ), 5.32(s, 1H, B 7), 4.67 (s, 2H,
0
10H, 2ꢂ–OCH2Ph), 7.10, 7.14 (2ꢂs, 2H, D7 & D7 ),
OCH2Ph), 3.8 (s, 2H, a). ESI MS (ES+): for
C62H89N11O16; calculated: 1228.444; found: (M+Na)+=
1250.4. UV (MeOH) lmax ( M-1 cmꢀ1): 210, 271 nm
(53,232, 2538).
5.33 (br, 1H, B7), 4.68 (s, 4H, 2ꢂ–OCH2Ph), 4.18 (m,
4H, c & d), 3.12(m, 8H, 2 ꢂe & h), 2.05 (m, 8H, 2ꢂf &
g). IR (KBr): nmax 3320–3420, 2920, 1660–1630, 1530,
1465, 1080 cmꢀ1. ESI MS (ES+): for C72H107N9O16;
calculated: 1354.682; found: (M+Na)+=1376.6 (base
peak), 1354.5, 1305.6. UV (MeOH) lmax ( M-1 cmꢀ1):
208, 230, 289 nm (64,738, 12888, 5155).
4.5.3. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S)-2-(3-(1-azinanylmethyl)-4-hydroxyphenyl)-2-
benzyloxy - 1 - hydroxyethyl) - 12 - benzyloxy - 2,11,15 - tri
hydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-16-
methyl-5,8,14,19,22,25-hexaoxoperhydrodiazolo[2,1-c:2,
1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-yl]-12-methyl-
4.5.7. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy- 23- ((1 S) - 2- benzyloxy- 1- hydroxy-2 - (4-
hydroxy-3-(4-methyl-1-azinanylmethyl)phenyl)ethyl)-2,
11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxyme-
thyl - 16 - methyl - 5,8,14,19,22,25 - hexaoxoperhydrodia-
zolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-
1
tetradecanamide (38). Partial H NMR: d 7.25–7.41 (m,
10H, 2ꢂOCH2Ph), 7.2(dd, 1H, J=8.5 Hz & 1.85 Hz,
0
D7 ), 7.14 (d, 1H, 1 J=0.85 Hz, D7), 6.87 (d, 1H, J=8.5
1
0
Hz, D8 ), 5.35 (br, 1H, B7), 4.6 (s, 4H, 2ꢂ–OCH2Ph),
yl]-12-methyltetradecanamide (42). Partial H NMR: d
4.14 (s, 2H, d), 3.12(m, 4H, e & i), 2.04 (m, 6H, f, g &
h). IR (KBr): nmax 3300–3400, 2915, 1650, 1620, 1530,
1440, 1250 cmꢀ1. ESI MS (ES+): for C68H100N8O16;
calculated: 1285.576; found: (M+Na)+=1308.6 (base
peak). UV (MeOH) lmax (e M-1 cmꢀ1): 211, 255, 288 nm
(73,984, 20,087, 5142).
7.2–7.41 (m, 10H, 2ꢂ–OCH2Ph), 7.17 (dd, 1H, J=8.32
0
Hz & 1.8 Hz, D7 ), 7.0 (d, 1H, J=1.8 Hz, D7), 6.78 (d,
0
1H, J=8.32Hz, D8 ), 5.31 (br, 1H, B7), 4.68 (s, 4H, 2ꢂ–
OCH2Ph), 4.1 (s, 2H, d), 2.65 (m, 4H, e & i), 1.85 (m,
4H, f & h), 1.28 (m, 1H, g), 1.06 (m, 3H, CHCH3). IR
(KBr, acetate salt): n
3330–3400, 2950, 1717, 1635,
max
1530, 1450, 1250 cmꢀ1. ESI MS (ES+): for C69H102N8O16;
calculated: 1299.602; found: (M+Na)+=1321.7 (base
peak). UV (MeOH) lmax (e M-1 cmꢀ1): 208, 230, 284 nm
(49233, 17260, 3249).
4.5.4. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S)-2-benzyloxy-2-(3,5-di(1-azinanyl-
methyl)-4-hydroxyphenyl)-1-hydroxyethyl)-2,11,15-tri-
hydroxy-6-(1R)-1-hydroxyethyl)-20-hydroxymethyl-16-
methyl-5,8,14,19,22,25-hexaoxoperhydrodiazolo[2,1-c:2,1-/]
[1,4,7,10,13,16]hexaazacyclohenicosin-9-yl]-12-methyl-
4.5.8. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy - 23- ((1S) - 2 - benzyloxy - 1 - hydroxy- 2 - (4-
hydroxy-3,5-di(4-methyl-1-azinanylmethyl)phenyl)ethyl)-
2,11,15-trihydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxy-
methyl - 16 - methyl - 5,8,14,19,22,25 - hexaoxoperhydrodi-
1
tetradecanamide (39). Partial H NMR: d 7.28–7.45 (m,
0
10H, 2ꢂ–OCH2Ph), 7.21 (2ꢂs, 2H, D7 & D7 ), 5.32(br,
1H, B7), 4.65 (s, 4H, 2ꢂ-OCH2Ph), 4.11 (m, 4H, c & d),

1766
B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
azolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-
1
4.5.12. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S,2S)-1,2-dihydroxy-2-(3-(4-(4-fluorophenyl)-1,4-
diazinan-1-ylmethyl)-4-hydroxyphenyl)ethyl)-2,11,15-tri-
hydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-12-
methoxy-16-methyl-5,8,14,19,22,25-hexaoxoperhydrodi-
azolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-
yl]-12-methyltetradecanamide (48). Partial H NMR: d
0
7.18 (dd, 1H, J=8.40 Hz & 1.53 Hz, D7 ), 7.08 (d, 1H,
J=1.53 Hz, D7), 7.02(d, 4H, J=8.25 Hz, i, j, k & l), 6.8
yl]-12-methyltetradecana-mide (43). Partial H NMR: d
7.25–7.41 (m, 10H, 2ꢂ–OCH2Ph), 7.09, 7.21 (2ꢂs, 2H,
0
D7 & D7 ), 5.33 (br, 1H, B7), 4.68 (s, 4H, 2ꢂOCH2Ph),
4.11 (s, 4H, c & d), 2.7 (m, 8H, 2ꢂe & i), 1.85 (m, 8H,
2ꢂh & f), 1.25 (m, 2H, 2ꢂg), 1.06 (m, 6H, 2ꢂj). IR
(KBr, acetate salt): nmax 3350–3450, 2960, 1715,
1635, 1530, 1455, 1060 cmꢀ1. ESI MS (ES+): for
C76H115N9O16; calculated: 1410.789; found: (M+Na)+=
1432.9, 1411.6. UV (MeOH) lmax (e Mꢀ1 cmꢀ1): at
neutral pH: 206, 237, 288 nm (1463, 153, 29); at acidic
pH: 205, 229, 286 nm (1339, 197, 28).
1
0
(d, 1H, 8.40 Hz, D8 ), 5.12(d, 1H, J=1.5 Hz, B7), 3.83
(s, 2H, d), 3.38 (s, 3H, OCH3), 3.2(br, 4H, f & g), 2.79
(br, 4H, e & h). IR (KBr): nmax 3300–3400, 2930,
1645, 1620, 1510, 1440, 1380 cmꢀ1. ESI MS (ES+):
for C60H92FN9O16; calculated: 1214.429; found:
(M+Na)+=1236.7, 1056.4 (base peak). UV (MeOH)
lmax (e M-1 cmꢀ1): at neutral pH: 205, 230, 282 nm
(35,278, 16251, 1477); at acidic pH: 205, 228, 286 nm
(33,298, 11,592, 2255); at basic pH: 210, 246, 294 nm
(58,067, 11,328, 2062).
4.5.9. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S)-2-benzyloxy-1-hydroxy-2-(4-
hydroxy-3-(4-(3-trifluromethylphenyl)-1,4-diazinan-1-yl-
methyl)phenyl) ethyl)-2,11,15-trihydroxy-6-((1R)-1-hydro-
xyethyl)-20-hydroxymethyl-16-methyl-5,8,14,19,22,25-
hexaoxoperhydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexa-
azacyclohenicosin-9-yl]-12-methyltetradecanamide (44).
1
Partial H NMR: d 7.28–7.5 (m, 10H, 2ꢂ–OCH2Ph),
4.5.13. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S,2S)-2-(3,5-di(4-(4-fluorophenyl)-1,4-diazinan-1-
ylmethyl)-4-hydroxyphenyl)-1,2-dihydroxyethyl)-2,11,15-
trihydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-
12-methoxy-16-methyl-5,8,14,19,22,25-hexaoxoperhydro-
diazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexaazacyclohenicosin-
9-yl]-12-methyltetradecanamide (49). Partial 1H NMR: d
7.15–7.27 (m, 4H, i, j, k & l), 7.12(dd, 1H, J=8.22 Hz,
0
& 1.38 Hz, D7 ), 7.05 (d, 1H, J=1.38 Hz, D7), 6.85 (d,
0
1H, J=8.22 Hz, D8 ), 5.32(br, 1H, B7), 4.68 (s, 4H, 2ꢂ-
OCH2Ph), 3.85 (s, 2H, d), 2.81 (m, 8H, e, f, g & h). IR
(KBr): nmax 3300–3400, 2910, 2330, 1640 br, 1610,
1515, 1430, 1300, 1220, 1065 cmꢀ1. ESI MS (ES+):
for C74H102F3N9O16; calculated: 1430.659; found:
(M+Na)+=1452.7, 1222.2 (base peak).
0
7.13 (s, 2H, D7 & D7 ), 7.0–7.1 (m, 8H, 2ꢂg, h, i & j),
5.12(br, 1H, B7), 3.82(s, 4H, 2 ꢂb), 3.38 (s, 3H, OCH3),
3.21 (br, 8H, 2ꢂd & e), 2.78 (br, 8H, 2ꢂc & f). IR
(KBr): nmax 3300–3400, 2930, 1645, 1620, 1510, 1440,
1380 cmꢀ1. ESI MS (ES+): for C71H105F2N11O16; cal-
culated: 1406.665; found: (M+Na)+=1428.9, 1068.4
(base peak). UV (MeOH) lmax (e M-1 cmꢀ1): 207, 215,
234, 284 nm (46,370, 30,669, 14,068, 2900).
4.5.10. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S)-2-benzyloxy-1-hydroxy-2-(4-hydr-
oxy-3,5-di(4-(3-trifluromethylphenyl)-1,4-diazinan-1-yl-
methyl)phenyl) ethyl)-2,11,15-trihydroxy-6-((1R)-1-hydr-
oxyethyl)-20-hydroxymethyl-16-methyl-5,8,14, 19,22,25-
hexaoxoperhydrodiazolo[2,1-c:2,1-/][1,4,7,10,13,16]hexa-
azacyclohenicosin-9-yl]-12-methyltetradecanamide (45).
4.5.14. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S,2S)-1,2-dihydroxy-2-(4-hydroxy-3-(4-phenyl-1,4-
diazinan-1-ylmethyl)phenyl)ethyl)-2,11,15-trihydroxy-6-
((1R)-1-hydroxyethyl)-20-hydroxymethyl-12-methoxy-
16-methyl-5,8,14,19,22,25-hexaoxoperhydrodiaz-olo[2,1-
c:2,1 - /][1,4,7,10,13,16]hexaazacyclohenicosin - 9 - yl] - 12-
1
Partial H NMR: d 7.25–7.45 (m, 10H, 2ꢂ-OCH2Ph),
0
7.02-7.2 (m, 10H, D7, D7 , 2ꢂi, j, k & l), 5.33 (br, 1H,
B7), 4.68 (s, 4H, 2ꢂ-OCH2Ph), 3.8 (s, 4H, c & d), 2.75-
2.9 (m, 16H, 2ꢂe, f, g & h). IR (KBr): nmax 3300–3400,
2925, 1660, 1610, 1540, 1455, 1330, 1260, 1075 cmꢀ1
.
1
ESI MS (ES+): for C86H115F6N11O16; calculated:
1672.903; found: (M+Na)+=1695.5, 1222.6. UV
(MeOH) lmax (e M-1 cmꢀ1): 212, 255, 282, 305 nm
(41827, 20244, 4567, 2018).
methyltetradecanamide (50). Partial H NMR: d 7.22–
7.35 (m, 2H, j & l), 7.2(dd, 1H, J=8.22 Hz & 1.98 Hz,
0
D7 ), 7.1 (d, 1H, J=1.98 Hz, D7), 7.02(m, 2H, i & m),
0
6.9 (m, 1H, k), 6.81 (d, 1H, J=8.22 Hz, D8 ), 5.13 (d,
1H, 1.5 Hz, B7), 3.9 (s, 2H, d), 3.42(s, 3H, OC H3), 3.2-
3.3 (br, 4H, f & g), 2.85–2.95 (br, 4H, e & h). IR (KBr):
4.5.11. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
12-benzyloxy-23-((1S)-2-benzyloxy-2-(3-dibenzylamino-
methyl-4-hydroxyphenyl)-1-hydroxyethyl)-2,11,15-tri-
hydroxy-6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-16-
methyl-5,8,14,19,22,25-hexaoxoperhydrodiazolo[2,1-c:2,
1-/][1,4,7,10,13,16]hexaazacyclohenicosin-9-yl]-12-methyl-
3350–3450, 2920, 1650, 1620, 1530, 1435, 1375,
max
1220, 1070 cmꢀ1. ESI MS (ES+): for C60H93N9O16;
calculated: 1196.439; found: (M+Na)+=1218.2,
1056.4 (base peak). UV (MeOH) lmax (e Mꢀ1 cmꢀ1):
207, 232, 248, 279 nm (44,536, 15,767, 15,368, 3562).
1
tetradecanamide (46). Partial H NMR: d 7.22–7.44 (m,
20H, 2ꢂ–OCH2Ph & –N(CH2Ph)2), 7.11 (dd, 1H,
4.5.15. N1-[(6S,9S,14aS,15S,16S,20S,23S,25aS,2R,11R)-
23-((1S,2S)-1,2-dihydroxy-2-(3,5-di(4-phenyl-1,4-diazinan-
1-ylmethyl)-4-hydroxyphenyl)ethyl)-2,11,15-trihydroxy-
6-((1R)-1-hydroxyethyl)-20-hydroxymethyl-12-methoxy-
16-methyl-5,8,14,19,22,25-hexaoxoperhy-drodiazolo[2,1-
c:2,1 - /][1,4,7,10,13,16]hexaazacyclohenicosin - 9 - yl] - 12-
0
J=8.6 Hz & 2.2 Hz, D7 ), 7.08 (d, 1H, J=2.2 Hz, D7),
0
6.81 (d, 1H, J=8.6 Hz, D8 ), 5.3 (br, 1H, B7), 4.68 (s,
4H, 2ꢂ–OCH2Ph), 3.6-3.7 (s, 4H, 2ꢂe), 3.79 (s, 2H, d).
IR (KBr): nmax 3300–3400, 2930, 1650, 1615, 1516, 1435
cmꢀ1. ESIMS (ES+): for C77H104N8O16; calculated:
1397.706; found: (M+Na)+=1421.6, 1222.8 (base
peak). UV (MeOH) lmax (e M-1 cmꢀ1): 210, 228, 280 nm
(61484, 15835, 2697).
1
methyltetrade-canamide (51). Partial H NMR: d 7.24–
0
7.41 (m, 4H, 2ꢂh & j), 7.15 (s, 2H, D7 & D7 ), 7.0 (m,
4H, 2ꢂg & k), 6.89 (m, 2H, 2ꢂi), 5.1 (br, 1H, B7), 3.83

B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
1767
(s, 4H, 2ꢂb), 3.4 (s, 3H, CH3), 3.12–3.21 (br, 8H, 2ꢂd &
e), 2.68–2.95 (br, 8H, 2ꢂc & f). IR (KBr): nmax 3350–
3450, 2920, 1650, 1620, 1530, 1435, 1375, 1220, 1070
cmꢀ1. ESI MS (ES+): for C71H107N11O16; calculated:
1370.684; found: (M+Na)+=1393.0, 1054.3 (base
peak). UV (MeOH) lmax (e M-1 cmꢀ1): 205, 248, 279 nm
(29,408, 8099, 1557).
9. Fujisawa launches Funguard (Micafungin) in Japan,
Fujisawa Pharmaceutical Co Ltd; Press Release Posted
on: 10 December 2002.
10. (a) Kurtz, M. B.; Douglas, C. M. J. Med. Vet. Mycol.
1997, 35, 79. (b) Onishi, J.; Meinz, M.; Thompson, J.;
Curotto, J.; Dreikorn, S.; Rosenbach, M.; Douglas, C.;
Abruzzo, G.; Flattery, A.; Kong, L.; Cabello, A.; Vicente,
F.; Pelaez, F.; Diez, M. T.; Martin, I.; Bills, G.; Giacobbe,
R.; Dombrowski, A.; Schwartz, R.; Morris, S.; Harris, G.;
Tsipouras, A.; Wilson, K.; Kurtz, M. B. Antimicrob.
Agents Chemother. 2000, 44, 368. (c) Balani, S. K.; Xu, X.;
Arison, B. H.; Silva, M. V.; Gries, A.; DeLuna, F. A.;
Cui, D.; Kari, P. H.; Ly, T.; Hop, C. E.; Singh, R.; Wal-
lace, M. A.; Dean, D. C.; Lin, J. H.; Pearson, P. G.;
Baillie, T. A. Drug Metab. Dispos. 2000, 28, 1274.
11. (a) Ablordeppey, S. Y.; Fan, P.; Ablordeppey, J. H.;
Mardenborough, L. Curr. Med. Chem. 1999, 6, 1151. (b)
Green, L. J.; Marder, P.; Mann, L. L.; Chio, L.-C.; Cur-
rent, W. L. Antimicrob. Agents Chemother. 1999, 43, 830.
(c) Bartizal, K. Antimicrob. Agents Chemother. 1997, 41,
2326. (d) Tomishima, M.; Ohki, H.; Yanada, A.; Taka-
sugi, H.; Maki, K.; Tawara, S.; Tanaka, H. J. Antibiot.
1999, 52, 674.
12. (a) Roy, K.; Mukhopadhyay, T.; Reddy, G. C. S.; Desi-
kan, K. R.; Ganguli, B. N. J. Antibiotics 1987, 40, 275. (b)
Mukhopadhay, T.; Ganguli, B. N.; Fehlhaber, H. W.;
Kogler, H.; Vertesy, L. J. Antibiot. 1987, 40, 281.
13. (a) Hawser, S.; Borgonovi, M.; Markus, A.; Isert, D. J.
Antibiot. 1999, 52, 305. (b) Hawser, S.; Islam, K. J. Anti-
microb. Chemother. 1999, 43, 411.
14. (a) Balkovec, J. M.; Black, R. M.; Hammond, M. L.;
Heck, J. V.; Zambras, R. A.; Abruzzo, G.; Bartizal, K.;
Kropp, H.; Trainor, C.; Schwartz, R. E.; McFadden,
D. C.; Nollstadt, K. H.; Pittarelli, L. A.; Powels, M. A.;
Schmatz, D. M. J. Med. Chem. 1992, 35, 194. (b) Grutsch,
J. L., Jr.; Hansen, M. M.; Harkness, A. R.; Udodong, U.
E.; Verral, D. E. PCT Int. Appl. WO 9906062, 11 Feb.
1999.
Note: Details of reaction conditions, yields and related
data along with biological activity is available in Tables
3, 6–8 as supplementary material are not cited in the
text.
Acknowledgements
The authors wish to thank Antiinfective Research
Department, Hoechst AG, Germany for the constant
supply of Mulundocandin and our microbiology group
at Mulund for testing our compounds for antifungal
activity. Prof. S. Durani of IIT Mumbai for helpful
suggestions. We also thank Dr. Ramakrishna, N. V. S.
and his group for providing spectra.
References and notes
1. (a) Pfaller, M. A. Clin. Infect. Dis. 1994, 19 (suppl. 1), S8.
(b) Rees, J. R.; Pinner, R. W.; Hajjesh, R. A.; Brandt,
M. E.; Reingold, A. L. Clin. Infect. Dis. 1998, 27, 1138. (c)
Pfaller, M. A.; Wenzel, R. Eur. J. Clin. Microbiol. Infect.
Dis. 1992, 11, 287. (d) Beck-Sague, C. M.; Jarvis, W. R. J.
Infect. Dis 1993, 167, 1247. (e) Jarvis, W. R. Clin. Infect.
Dis 1995, 20, 1526. (f) Lewis, R. E.; Klepser, M. E. Am. J.
Health-Syst. Pharm. 1999, 56, 525.
2. (a) Walsh, T. J.; Jarrosinski, P. F.; Fromtling, R. A.
Diagn. Microbiol. Infect. Dis. 1990, 13, 37. (b) Samonis,
G.; Bafaloukos, D. In vivo 1992, 6, 183. (c) Pizzo, P. A.;
Young, L. S. Am. J. Med. 1984, 76, 101. (d) Walsh, T. J.;
Gonzalez, C.; Lyman, C. A.; Chanock, S. J.; Pizzo, P. A.
Adv. Pediatr. Infect. Dis. 1996, 11, 187.
15. (a) Lal, B.; Gund, V. G.; Gangopadhyay, A. K. PCT Int.
Appl., WO 0107468 A2 200110201, 2001 (b) Gund,
B. L. V. G.; Gangopadhyay, A. K.; Nadkarni, S. R.;
Dikshit, V.; Chatterjee, D. K.; Shirvaikar, R. Bioorg. Org.
Med. Chem. 2003, 11, 5189.
3. Gallis, H. A.; Drew, R. H.; Pickard, W. W. Rev. Inf. Dis
1990, 12, 308.
4. Bailey, E. M.; Drakovsky, D. J.; Ryback, M. J. Pharma-
cotherapy 1990, 10, 146.
16. Black, R. M.; Balkovec, J. M.; Nollstadt, K. M.; Drei-
korn, S.; Bartizal, K. F.; Abruzzo, G. K. Bioorg. Med.
Chem. Lett. 1997, 7, 2879.
17. (a) Flick, K.; Frankus, E.; Friderichs, E. Arzneim. Forsch.
1978, 28, 107. (b) Werner, W.; Jungstand, W.; Gutsche,
W.; Wohlrabe, K. Pharmazie 1977, 32, 341. (c) Cagniant,
P.; Kirsch, G.; Wierzbicki, M.; Lepage, F.; Cagniant, D.;
Lobenberg, D.; Parmegiani, R.; Sherlock, M. Eur. J. Med.
Chem. 1980, 15, 439. (d) de Narvaez, R.; Josefina, A.;
Elizabeth, F. Igne. Quim. Nova 1985, 8, 38. (e) Thiele, K.;
Posselt, K.; Offermanns, H.; Thiemer, K. Arzneim. -
Forsch 1980, 30, 747. (f) Poplevskaya, I. A.; Kondaurov,
G. N.; Abdullin, K. A.; Shipunova, L. K.; Chermanova,
G. B.; Kabiev, O. K. Tr. Inst. Khim. Nauk, Akad. Kaz.
SSR 1980, 52, 52. (g) Dimmock, J. R.; Raghavan, S. K.;
Logan, B. M.; Bigam, G. E. Eur. J. Med. Chem. Chim.
Ther. 1983, 18, 248. (h) Bundgaard, H. Methods in Enzy-
mology 1985, 112, 347.
18. (a) Fowler, J. S. J. Org. Chem. 1977, 42, 2637. (b)
Masuda, K.; Toga, T.; Hayashi, N. J. Labelled Compound
1975, 11, 301. (c) Nakatsuka, I.; Kawahara, K.; Yoshi-
tate, A. J. Labelled Comp. Radiopharm 1981, 18, 495. (d)
Schreier von, E. Helv. Chim. Acta 1976, 59, 585.
19. (a) Tramontini, M. Synthesis 1973, 703. (b) Tramontini,
M.; Angiolini, L. Tetrahydron 1990, 46, 1791. (c) Thomp-
son, B. B. J. Pharm. Sci. 1968, 57, 715.
5. (a) Fox, R.; Neal, K. R.; Leen, C. L. S.; Ellis, M. E.;
Mandal, B. K. J. Infect. 1991, 22, 201. (b) Smith, D.;
Boag, F.; Midgley, J.; Gazzard, B. J. Infect 1991, 23, 345.
(c) Hitchcock, C. A.; Pye, G. W.; Trok, F. J.; Johnson,
E. M.; Warnock, D. W. Antimicrob. Agents Chemother.
1993, 37, 1962. (d) Sanglard, D.; Kuchler, K.; Ischer, F.;
Pagani, J. L.; Monod, M.; Bille, J. Antimicrob. Agents
Chemother. 1995, 39, 2378. (e) Frosco, M.; Barrett, J. F.
Expert Opin. Invest. Drugs 1998, 7, 175. (f) Fanos, V.;
Cataldi, L. J. Chemother. 2000, 12, 463.
6. (a) Debono, M.; Turner, W. W.; LaGrandeur, L.; Bur-
khardt, F. J.; Nissen, J. S.; Nichols, K. K.; Rodriguez,
M. J.; Zweifel, M. J.; Zekner, D. J.; Gordee, R. S.; Tang,
J.; Parr, T. R., Jr. J. Med. Chem. 1995, 38, 3271. (b)
Masurekar, P. S.; Fountoulakis, J. M.; Hallada, T. C.;
Sosa, M. S.; Kaplan, L. J. Antibiot. 1992, 45, 1867. (c)
Iwamoto, T.; Fujie, A.; Nitta, K.; Hashimoto, S.; Oku-
hara, M.; Kohsaka, M. J. Antibiot. 1994, 47, 1092.
7. Debono, M.; Gordee, R. S. Ann. Rev. Microbiol. 1994, 48,
471.
8. Pound, M. W.; Drew, R. H.; Perfect, J. R. Curr. Opin.
Infect. Dis. 2002, 15, 183.

1768
B. Lal et al. / Bioorg. Med. Chem. 12 (2004) 1751–1768
20. Results will be published elsewhere.
1321. (b) Nawada, R.; Amitani, R.; Tanaka, E.; Niimi,
A.; Suzuki, K.; Murayama, T.; Kuze, F. J. Clin. Microb
1996, 34, 1433. (c) Valentin, A.; Guennec, R. L.; Rodri-
guez, E.; Reynes, J.; Mallie, M.; Bastide, J. M. Anti-
microb. Agents Chemother. 1996, 40, 1342. (d) Bartizal,
K.; Abruzzo, G.; Trainor, C.; Krupa, D.; Nollstadt, K.;
Schmatz, D.; Hammond, M.; Balkovec, J.; Van Middles-
worth, F. Antimicrob. Agents Chemother. 1992, 36, 1648.
21. (a) Antibiotics in Laboratory Medicine, 3rd ed.; Lorian,
V.; Williams and Wilkins: 1991; p 16. (b) Denning, D. W.;
Radford, S. A.; Oakley, K. L.; Hall, L.; Johnson, E. M.;
Warnock, D. W. J. Antimicrob. Chemother. 1997, 40, 401.
22. (a) Walsh, T. J.; Lee, J. W.; Kelly, P.; Bacher, J.; Lec-
ciones, J.; Thomas, V.; Lyman, C.; Coleman, D.; Gordee,
R.; Pizzo, P. A. Antimicrob. Agents Chemother. 1991, 35,