Aminoparthenolides as novel anti-leukemic agents: Discovery of the NF-κB inhibitor, DMAPT (LC-1)

Article abstract of DOI:10.1016/j.bmcl.2009.05.092

A series of aminoparthenolide analogs (6-37) were synthesized and evaluated for their anti-leukemic activity. Eight compounds exhibited good anti-leukemic activity with LD₅₀'s in the low μM range (1.5-3.0 μM). Compounds 16, 24 and 30 were the m

Full text of DOI:10.1016/j.bmcl.2009.05.092

Bioorganic & Medicinal Chemistry Letters 19 (2009) 4346–4349
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Aminoparthenolides as novel anti-leukemic agents: Discovery of the NF-
inhibitor, DMAPT (LC-1)
jB
Sundar Neelakantan a, Shama Nasim a, Monica L. Guzman b, Craig T. Jordan b, Peter A. Crooks a,
*
a Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, Lexington, KY 40536, USA
b James P. Wilmot Cancer Center, University of Rochester, Rochester, NY 14642, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of aminoparthenolide analogs (6–37) were synthesized and evaluated for their anti-leukemic
activity. Eight compounds exhibited good anti-leukemic activity with LD50’s in the low M range (1.5–
3.0 M). Compounds 16, 24 and 30 were the most potent compounds in the series, causing greater than
90% cell death at 10
and 1.6 M.
Received 31 March 2009
Revised 18 May 2009
Accepted 20 May 2009
Available online 27 May 2009
l
l
l
M concentration against primary AML cells in culture, with LD50 values of 1.7, 1.8
l
Ó 2009 Elsevier Ltd. All rights reserved.
Keywords:
Sesquiterpene lactone
Parthenolide
Aminoparthenolide analogs
NF-jB inhibitor
DMAPT (LC-1)
Anti-leukemic activity
Leukemia
Parthenolide (PTL, 1, Fig. 1) isolated from Tanacetum parthenium
(commonly referred as feverfew), is a naturally occurring sesqui-
terpene lactone used in the treatment of fever, migraine head-
aches, rheumatoid arthritis, and also as an anti-inflammatory
agent.1–3 PTL is a neutral, lipophilic lactone with low polarity and
water-solubility (0.169 l
M/mL maximum solubility in serum),20
thus limiting its potential as a promising clinical agent.
Initial structure–activity studies carried out on the PTL mole-
cule involved reduction of the exocyclic double bond by catalytic
hydrogenation over palladium-on-charcoal to afford compound
2,21 which was inactive when evaluated for anti-leukemic activity
contains an
a-methylene-c-lactone ring and an epoxide moiety
that interact with nucleophilic sites on biological macromole-
cules.4 The PTL molecule and several structurally related analogs
have become topics of recent interest because of their potent
anti-tumor and cytotoxic properties.5–13
against primary AML cells in culture (Table 1). Thus, the
a-methy-
lene- -lactone moiety in PTL appears to be a critical functionality
c
for its cytotoxic effect. Interestingly, introduction of an additional
epoxide moiety at the C1–C10 double bond with m-chloroperoxy
benzoic acid, afforded compound 3, which was also inactive in
PTL and its analogs have been shown to promote apoptosis by
inhibiting the activity of the NF-
jB transcription factor complex,
the 10 lV probe cytotoxicity assay (Table 1). Oxidation of the
and thereby down-regulating anti-apoptotic genes under NF-
j
B
C14 methyl group of PTL via selenium dioxide/t-butyl hydroperox-
ide afforded a mixture of aldehyde 4 and the corresponding alco-
hol, 5,22 both of which were inactive as anti-leukemic agents
(Table 1).
Our laboratory has recently been successful in overcoming the
poor water-solubility of PTL without loss of its anti-leukemic activ-
ity, by derivatizing PTL into several aralkylamino analogs, which
can then be converted into water-soluble organic salts.23 We have
now carried out the synthesis, structure–activity relationships
(SAR) and anti-leukemic activity of a novel series of second gener-
ation water-soluble aminoparthenolide analogs derived from a
variety of aliphatic primary and secondary amines.
control.4,6,14–17 Recently, it has been shown that parthenolide in-
duces robust apoptosis of primary acute myeloid leukemic (AML)
cells.18,19 Notably, PTL causes cell death of AML stem and progen-
itor cells in vitro, with no observed toxicity towards normal hema-
topoietic cells. The apoptosis induced by PTL is not solely due to
NF-jB inhibition, but rather arises from a broad set of biological re-
sponses, which include activation of p53 and an increase in reac-
tive oxygen species. However, despite promising in vitro activity,
this potent natural product has a major limitation which precludes
its further development as a therapeutic agent, that is, its poor
To obtain a sub-library of these water-soluble PTL analogs, we
exploited the highly electrophilic a-methylene-c-lactone ring nat-
* Corresponding author. Tel.: +1 859 257 1718; fax: +1 859 257 7585.
E-mail address: pcrooks@email.uky.edu (P.A. Crooks).
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.05.092

S. Neelakantan et al. / Bioorg. Med. Chem. Lett. 19 (2009) 4346–4349
4347
14
1
4
9
6
2
3
8
10
5
7
13
11
12
O
15
O
Parthenolide (1) O
Figure 1. The structure of parthenolide (PTL, 1).
Table 1
Anti-leukemic activity of parthenolide (1) and analogs 2-5
Compound #
Structure
Anti-leukemic activity
(% primary AML cell
death at 10
lM)
1
84
O
O
O
O
Figure 3. Single crystal structure of 13-(N,N-dimethyl)amino-4
a
,5b-epoxy-4,10-
dimethyl)-6a-hydroxy-12-oic acid-c-lactone-germacra-1(10)-ene (16, DMAPT, LC-
1).
2
3
4
5
0
0
0
0
O
O
the protonation of the enolate formed during Michael addition,
which occurs exclusively from the exo face of the molecule, result-
ing in an R configuration at C-11 in the product. The above obser-
vation is consistent with the structure of Michael addition
products from other structurally related sesquiterpene lactones re-
ported in the literature.25,26
The aminoparthenolide analogs (6–37) were synthesized by
reacting PTL with a variety of aliphatic primary and secondary
amines. PTL (25 mg, 0.1 mmol) was dissolved in 8 mL of methanol,
the appropriate amine (0.15 mmol) added, and the mixture was
stirred under ambient conditions for 6–8 h. The crude product
was subjected to flash silica gel column chromatography to afford
the pure aminoparthenolide. The compounds synthesized and their
O
O
O
O
O
O
CHO
O
O
OH
anti-leukemic activities (10 lM concentration) in the probe assay
O
O
are shown in Table 2. Selected analogs were also evaluated in full
dose-response assays to generate LD50 values, which are also given
in Table 2.
The 10 lM probe assays were carried out as follows: one mil-
urally present in the molecule. Compounds containing an
a-meth-
lion primary acute myelogenous leukemia (AML) cells, obtained
from volunteer donors with informed consent, were cultured in
serum free medium (SFM).27 Cells were treated for 24 h in the
presence or absence of the appropriate PTL analog at a probe con-
ylene- -lactone ring are excellent Michael acceptors, and undergo
c
facile Michael addition. Michael addition reactions were performed
using a variety of primary and secondary aliphatic amines, and the
amino parthenolide analogs thus obtained (6–37) were converted
into water-soluble organic acid salts, thereby increasing their
water solubility and bioavailability (see Fig. 2).
The Michael addition of aliphatic amines with PTL was found to
be highly stereospecific and yielded exclusively a single stereoiso-
mer with the R-configuration at the newly formed C-11 chiral car-
bon. None of the alternative diastereomer with the ‘S’ configuration
at C-11 was observed in the reaction mixture. We have confirmed
the C-11 R-configuration in many of our synthesized aminoparthe-
nolide analogs by single crystal X-ray analysis. The ORTEP struc-
centration of 10
lV. Selected analogs were then evaluated in a
dose–response assay over the concentration range 2.5–20
lM, to
obtain LD50 values. Cell viability was determined as previously de-
scribed;28 briefly, cells were washed with cold PBS and resus-
pended in 200 lL of Annexin binding buffer (10 mM HEPES/
NaOH pH 7.4; 140 mM NaCl; 2.5 mM CaCl2). Annexin-V and 7-ami-
no-actinomycin (7-AAD) were added and the tubes were incubated
at ambient temperature in the dark for 15 min. Cells were then di-
luted with 200 lL of Annexin binding buffer and analyzed immedi-
ately by flow cytometry. Viable cells were scored as Annexin V
negative/7-AAD negative. Data provided are normalized to un-
treated control specimens. LD50 values were determined using Cal-
cusyn software (Biosoft).
From the structure–activity data shown in Table 2, it is evi-
dent that several of the aminoparthenolide analogs obtained
from the reaction of secondary aliphatic amines with PTL (com-
pounds 16–36) exhibit significant anti-leukemic activity in the
AML assay compared to those analogs obtained from primary
amines (compounds 6–15). In the acyclic aliphatic secondary
amine series, the methylpropylamino analog 21 was the most
ture of
a
representative compound, 13-(N,N-dimethyl)amino-
-hydroxy-12-oic acid- -lactone-
4a,5b-epoxy-4,10-dimethyl-6
a
c
germacra-1(10)-ene mono-fumarate (16), is illustrated in Figure
3.24 The exclusive formation of the C-11 R diastereomer is due to
1) 1º or 2º Amine
COOH
MeOH, rt
R
O
O
2) Fumaric Acid
Diethyl Ether
O
O
HOOC
O
O
6-37
1
potent analog in the 10 lM probe assay, and exhibited an LD50
Figure 2. Synthesis of aminoparthenolide analogs.

4348
S. Neelakantan et al. / Bioorg. Med. Chem. Lett. 19 (2009) 4346–4349
Table 2
of 3.2
potent analog in the series (93% cell death at 10
assay; LD50 = 1.7 M). In the symmetrical secondary aliphatic
l
M. The dimethylamino analog 16 was the second most
Anti-leukemic activity of amino parthenolide analogs against AML cells in culture
l
M in the probe
a
l
Compound
R
Anti-leukemic activity
LD50
M)
(% cell death at 10
lM)
(
l
amine series, as one moves from dimethylamino (16) to higher
dialkylamino analogs, that is, diethylamino (18), diisopropyl-
amino (20), dipropylamino (25), and diallylamino (26), the anti-
leukemic activity gradually decreases. It was also observed that
aminoparthenolide analogs with at least one N-methyl group
exhibited better antileukemic activity than secondary aminopar-
thenolide analogs lacking an N-methyl group. In secondary
aminoparthenolide analogs that contained an N-methyl group,
increasing the chain length of the other alkyl moiety resulted
in a gradual decrease in antileukemic activity. Thus, the N-
methyl, N-propyl analog (21), the N-methyl, N-butyl analog
(22), and the N-methyl, N-pentyl analog (23) exhibited 95%,
68% and 46% cell death in the AML probe assay, respectively.
The N-methyl, N-2-hydroxyethyl analog (24) exhibited 86% cell
1
6
—
84
4.0
1.4
H3C–NH
18
31
—
H3C
NH
7
8
2.0
6.0
7.0
0
NH
NH
9
—
NH
NH
NH
10
11
—
HO
HO
0
—
12
0
—
H2N
13
14
0
0
—
—
H2N
N
NH
NH
NH
15
16
17
18
19
0
93
80
60
78
—
death in the probe assay and afforded an LD50 of 1.8
l
M, while
1.7
2.5
N
no observable cell death was observed at 10
lM for the structur-
N
N
ally related N-demethylated analog 10.
CH3
Among the aliphatic cyclic amino analogs examined, ring size
appeared to play an important role in antileukemic activity. The
optimum ring size was found to be between five and six; analogs
with smaller or greater ring size had significantly reduced antileu-
—
—
—
H3C
N
kemic activity in the 10
log (28) and the piperidino analog (29) afforded 82% and 76% cell
death in the 10 M probe assay, with LD50’s of 2.7 and 2.4 M,
lM probe assay. Thus, the pyrrolidino ana-
20
21
22
20
95
68
N
l
l
H3C
H3C
CH3
3.2
N
H
respectively, whereas the 3-, 7- and 8-membered cyclic amino ana-
logs 27, 32 and 33 exhibited values in the range 12–22% cell death
in the probe assay.
Introduction of a heteroatom, such as oxygen, into the six-
membered cyclic amine moiety (e.g., analog 29, 76% cell death at
2.8
N
H
CH3
H3C
H3C
H3C
CH3
23
24
25
26
27
28
29
30
46
86
0
4.6
1.8
N
N
10
l
M; LD50 = 2.4
l
M) to afford a morpholino analog (analog 31,
M) resulted in a significant decrease in cyto-
OH
20% cell death at 10
l
CH3
CH2
toxicity, whereas introduction of a methyl group at C-4 of the
piperidine ring in analog 29 to afford 30 (83% cell death at
10 lM; LD50 = 1.6 lM), increased anti-leukemic activity (Table 2).
Thus, compounds 16–17, 21–22, 24, 28–30 exhibited the most
—
N
H2C
N
0
—
—
N
22
82
76
83
promising anti-leukemic activities, with LD50 values in the low
2.7
l
M range (1.6–2.7 lM); full characterization data on four of the
N
N
more potent analogs are provided.29
2.4
1.6
In summary, 31 aminoparthenolide analogs have been synthe-
sized and evaluated for anti-leukemic activity against primary
AML cells in culture. All of the synthesized compounds were more
water-soluble than PTL; some analogs exhibited 100-fold greater
water-solubility than PTL. A range of antileukemic activities was
observed, with several analogs exhibiting LD50 values between 1
N
H3C
O
31
32
20
15
14
N
N
—
—
and 2
Compounds 16, 24, and 30 were the most potent compounds in
the series causing 80–90% cell death of AML cells in the 10
probe assay, and affording LD50 values of 1.7, 1.8 and 1.6
respectively. 13-(N,N-Dimethyl)amino-4 ,5b-epoxy-4,10-di-
methyl-6 -hydroxy-12-oic-acid- -lactone-germacra-1(10)-ene
lM.
N
N
33
34
35
13
12
0
lV
lM,
a
—
a
c
CH3
N
(16; DMAPT, LC-1) was selected as a lead compound, based on
favourable pharmacokinetic and pharmacodynamic properties
(unpublished results).
15
HO
HO
OH
Aminoparthenolides have been reported to undergo retro-Mi-
chael degradation to parthenolide and the parent amine in the
presence of nucleophiles.30,21 Thus, we were concerned that the
aminoparthenolides might be degrading in the cell culture buffer
during the AML cell assay. Stability studies on DMAPT in the HEPES
buffer utilized in the antileukemic cell assay by HPLC over the
time-course of the experiment indicated that <3% of DMAPT had
degraded to parthenolide at 24 h. This indicates that DMAPT is
not degrading and releasing significant amounts of parthenolide
into the culture media over time.
36
0
—
O
HN
OH
OH
HO
HO
O
37
0
—
N
OH
H3C
a
Concentration of analog causing death of 50% of the cell population in primary
cultures of AML cells.

S. Neelakantan et al. / Bioorg. Med. Chem. Lett. 19 (2009) 4346–4349
4349
21. Hwang, D. R.; Wu, Y. S.; Chang, C. W.; Lien, T. W.; Chen, W. C.; Tan, U. K.; John,
T. A.; Hsu, J. T.; Hsieh, H. P. Bioorg. Med. Chem. 2006, 14, 83.
22. El-Feraly, F. S. Phytochemistry 1984, 23, 2372.
23. Nasim, S.; Crooks, P. A. Bioorg. Med. Chem. Lett. 2008, 18, 3870.
24. Neelakantan, S.; Parkin, S.; Crooks, P. A. Acta Crystallogr., Sect. E, in press.
25. Lawrence, N. J.; McGown, A. T.; Nduka, J.; Hadfield, J. A.; Pritchard, R. G. Bioorg.
Med. Chem. Lett. 2001, 11, 429.
26. Matsuda, H.; Kagerura, T.; Toguchida, I.; Ueda, H.; Morikawa, T.; Yoshikawa, M.
Life Sci. 2000, 66, 2151.
27. Lansdorp, P. M.; Dragowska, W. J. Exp. Med. 1992, 175, 1501.
28. Guzman, M. L.; Neering, S. J.; Upchurch, D.; Grimes, B.; Howard, D. S.; Rizzieri,
D. A.; Luger, S. M.; Jordan, C. T. Blood 2001, 98, 2301.
This stability data for DMAPT is consistent with the variable SAR
obtained for the 31 aminoparthenolide analogs examined. Further-
more, pharmacokinetic studies in the rat indicate that DMAPT has
an oral bioavailability of around 70%, and is biotransformed into a
major metabolite: 13-(N-methyl)amino-4
a,5b-epoxy-4,10-di-
methyl-6 -hydroxy-12-oic-acid- -lactone-germacra-1(10)-ene via
a
c
oxidative N-demethylation (unpublished data). Only very small
amounts of parthenolide were detected in plasma over 8 h post oral
dosing.
Recently, more detailed investigations into DMAPT’s anti-leuke-
mic activity have been carried out against cultured AML, CML and
CLL cells, as well as in in vivo studies in dogs with canine leuke-
mia.31,32 In addition, we have also shown that DMAPT causes a
dose-dependent decrease in the binding of the Rel-A subunit of
29. Representative characterization data for four of the most active compounds 16,
17, 24, and 30 are provided below: 13-(N,N-dimethyl)-amino-4
a,5b-epoxy-4,10-
di methyl-6 -hydroxy-12-oic acid- -lactone-germacra-1(10)-ene monofumarate
a
c
(16): mp: 183–184 °C; yield, 95%. 1H NMR (300 MHz, D2O): d 6.67 (2H, s,
(HOOC–CH)2–), 5.24 (1H, dd, J = 2.1, 12.0 Hz, 1-CH), 4.30 (1H, t, J = 9.0 Hz, 6-
CH), 2.98 (6H, s, N–(CH3)2), 1.88–3.60 (13H, m, 2-CH2, 3-CH2, 8-CH2, 9-CH2, 13-
CH2, 5-CH, 7-CH, 11-CH), 1.70 (3H, s, 14-CH3), 1.34 (3H, s, 15-CH3) ppm. 13C
NMR (75 MHz, D2O): d 179.8 ((HOOC–CH)2–), 173.8 (12-C@O), 138.2 ((HOOC–
CH)2–),137.2 (10-C), 127.3 (1-C), 85.8 (6-C), 69.2 (N–(CH3)2), 67.1 (5-C), 58.3
(4-C), 49.6 (7-C), 44.9 (11-C), 42.5 (9-C), 38.1 (3-C), 30.9 (8-C), 26.1(2-C), 18.7
(15-C), 18.6 (14-C) ppm. Elemental Anal. Calcd for C21H31NO7: C, 61.60; H,
7.63; N, 3.42. Found: C, 61.81; H, 7.64; N, 3.47. 13-(N-ethyl-N-methyl)-amino-
NF-
of three NF-
j
B to DNA, thereby triggering apoptosis.33 Also, transcription
jB-regulated genes, CFLAR, BCL2, and BIRC5, which
have been implicated in CLL cell survival and resistance to chemo-
therapy, were all significantly suppressed by DMAPT.32 DMAPT is
currently being evaluated in AML patients in a first-in-man study
in the United Kingdom.
4a,5b-epoxy-4,10-dimethyl}-6a-hydroxy-12-oicacidc-lactone-germacra-1(10)-
ene monofumarate (17): mp: 156–157 °C; yield, 80%. 1H NMR (400 MHz, D2O) d
6.70 (2H, s, (HOOC–CH)2–), 5.17 (1H, app d, J = 12.0 Hz, 1-CH), 4.33 (1H, t,
J = 12.0 Hz, 6-CH), 2.95 (3H, s, N-CH3), 1.71 (3H, s, 14-CH3), 1.36 (3H, s, 15-CH3),
1.27–3.75 (15H, m, 2-CH2, 3-CH2, 8-CH2, 9-CH2, 13-CH2, N-CH2-CH3 5-CH, 7-CH,
11-CH) 1.24 (3H, t, J = 12.0 Hz, N–CH2–CH3) ppm; 13C NMR (75 MHz, D2O) d
179.1 ((HOOC-CH)2-), 173.0 (12-C@O), 137.3 ((HOOC–CH)2–), 136.3 (10-C),
126.5 (1-C), 85.2 (6-C), 68.4 (N-CH3), 66.6 (5-C), 55.5 (4-C), 55.2 (13-C), 52.2
(N–CH2–CH3), 49.1 (7-C), 44.2 (11-C), 42.0 (9-C), 37.6 (3-C), 30.4 (8-C), 25.6 (2-
C), 18.1 (15-C), 18.0 (14-C), 10.7 (N-CH2-CH3) ppm. MALDI-TOFMS: 308 m/z
[M+]: Calcd. C18H29NO3 (307.2142) observed (307.2147), EI MS m/z 307.
Elemental Anal. Calcd for C22H33NO7: C, 62.39; H, 7.85; N, 3.31. Found: C,
Acknowledgement
This work was supported by a grant from the Kentucky Lung
Cancer Research Program.
References and notes
61.98; H, 7.85; N, 3.30. 13-(N-(2-hydroxyethyl)-N-methyl)-amino-4
a,5b-epoxy-
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Biol. 2006, 26, 1864.
4,10-dimethyl-6 -hydroxy-12-oicacid- -lactone germacra-1(10)-ene
a
c
monofumarate (24): mp: 122–124 °C; yield, 78%. 1H NMR (400 MHz, D2O) d
6.68 (2H, s, (HOOC–CH)2–), 5.10 (1H, app. d, J = 10.0 Hz, 1-CH), 4.34 (1H, t,
J = 9.2 Hz, 6-CH), 3.95 (2H, t, J = 5.2 Hz, N–CH2–CH2–OH), 3.01 (3H, s, N-CH3),
1.34 (3H, s, 14-CH3), 1.69 (3H, s, 15-CH3), 1.89–3.70 (15H, m, 2-CH2, 3-CH2, 8-
CH2, 9-CH2, 13-CH2, N–CH2, 5-CH, 7-CH, 11-CH) ppm; 13C NMR (75 MHz, D2O):
d 179.5 ((HOOC–CH)2–), 173.0 (12-C@O), 137.7 ((HOOC–CH)2–), 136.5 (10-C),
126.8 (1-C), 85.3 (6-C), 68.7 (N-CH3), 67.0 (5-C), 58.4 (4-C), 57.0 (N–CH2–CH2–
OH) 56.5 (13-C), 52.5 (N–CH2–CH2–OH), 49.3 (7-C), 44.3 (11-C), 42.0 (9-C), 37.6
(3-C), 30.3 (8-C), 25.6 (2-C), 18.2 (15-C), 18.0 (14-C). MALDI-TOFMS: 324 m/z
[M+]: Calcd. C18H29NO4 (323.2091) observed (323.2090). EI MS (M+) m/z 323.
Elemental Anal. Calcd for C22H33NO8ꢀH2O: C, 57.75; H, 7.71; N, 3.06. Found: C,
57.73; H, 7.32; N, 3.42. 13-(4-Methylpiperidin-1-yl)-amino-4
a,5b-epo xy-4,10-
dimethyl-6 -hydroxy-12-oic acid- -lactone-germacr-a-1(10)-ene monofumarate
a
c
(30): mp: 132–134 °C; yield, 70%. 1H NMR (400 MHz, CD3OD) d 6.60 (2H, s,
(HOOC–CH)2–), 5.19 (1H, d, J = 10.0 Hz, 1-CH), 4.21 (1H, t, J = 9.2 Hz, 6-CH), 1.29
(3H, s, 14-CH3), 1.65 (3H, s, 15-CH3), 1.12–3.60 (22H, m, 2-CH2, 3-CH2, 8-CH2, 9-
CH2, 13-CH2, 2 ꢁ N–CH2, 2 ꢁ N–CH2–CH2–, piperidino 40-CH, 5-CH, 7-CH, 11-
CH), 0.97 (3H, d, J = 6.3 Hz, piperidino 4-CH3) ppm. 13C NMR (75 MHz, CD3OD) d
177.3 ((HOOC-CH)2-), 170.9 (12-C@O), 136.1 ((HOOC–CH)2–), 135.7 (10-C),
126.2 (1-C), 84.2 (6-C), 67.3 (5-C), 63.3 (4-C), 55.1 (2 ꢁ piperidino N-CH2), 56.2
(13-C), 49.5 (7-C), 44.5 (11-C), 41.9 (9-C), 37.6 (3-C), 32.5 (2 ꢁ piperidino N–
CH2–CH2), 30.0 (piperidino H3C–CH), 29.9 (8-C), 25.1 (2-C), 21.4 (piperidino-4-
CH3), 17.5 (15-C), 17.1 (14-C) ppm. MALDI-TOFMS: m/z 348 [M+]: Elemental
Anal. Calcd for C25H37NO7: C, 64.77; H, 8.05; N, 3.02. Found: C, 64.73; H, 8.29;
N, 3.27. EI MS (M+) m/z 347.
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