Pitipeptolides A and B, new cyclodepsipeptides from the marine cyanobacterium Lyngbya majuscula

Article abstract of DOI:10.1021/np000456u

Two new cyclodepsipeptides have been isolated from a population of the marine cyanobacterium Lyngbya majuscula collected at Piti Bomb Holes, Guam. They appear to be unique to this particular Guamanian collection and have been named pitipeptolides A (1) and B (2). Their structures have been elucidated by spectroscopic techniques and by characterization of degradation products. Distinctive features include the presence of a 2,2-dimethyl-3-hydroxy-7-octynoic acid residue in 1 and a 2,2-dimethyl-3-hydroxy-7-octenoic acid residue in 2, previously shown to be biosynthetic signatures of cyanobacterial metabolites. Pitipeptolides A (1) and B (2) exhibit weak cytotoxicity against LoVo cancer cells, but possess moderate antimycobacterial activity and stimulate elastase activity.

Full text of DOI:10.1021/np000456u

304
J . Nat. Prod. 2001, 64, 304-307
P itip ep tolid es A a n d B, New Cyclod ep sip ep tid es fr om th e Ma r in e
Cya n oba cter iu m Lyn gbya m a ju scu la
Hendrik Luesch,† Ronald Pangilinan,‡ Wesley Y. Yoshida,† Richard E. Moore,*,† and Valerie J . Paul*,‡
Department of Chemistry, University of Hawaii at Manoa, Honolulu, Hawaii 96822, and University of Guam Marine
Laboratory, UOG Station, Mangilao, Guam 96923
Received September 28, 2000
Two new cyclodepsipeptides have been isolated from a population of the marine cyanobacterium Lyngbya
majuscula collected at Piti Bomb Holes, Guam. They appear to be unique to this particular Guamanian
collection and have been named pitipeptolides A (1) and B (2). Their structures have been elucidated by
spectroscopic techniques and by characterization of degradation products. Distinctive features include
the presence of a 2,2-dimethyl-3-hydroxy-7-octynoic acid residue in 1 and a 2,2-dimethyl-3-hydroxy-7-
octenoic acid residue in 2, previously shown to be biosynthetic signatures of cyanobacterial metabolites.
Pitipeptolides A (1) and B (2) exhibit weak cytotoxicity against LoVo cancer cells, but possess moderate
antimycobacterial activity and stimulate elastase activity.
Cyanobacteria produce a wide array of bioactive second-
ary metabolites, and therefore, are valuable as a potential
source of novel pharmaceuticals.1 Nevertheless, cyanobac-
teria may represent a health threat if potent toxins are
being produced, especially under bloom conditions.2 In-
creasing attention has focused on both the biomedical and
ecological aspects of these organisms.3 We recently initiated
chemical investigations of Guamanian varieties of marine
cyanobacteria, predominantly strains of Lyngbya majus-
cula Harvey ex Gomont, affording several new cytotoxins
whose anticancer activities have been assessed.4 L. ma-
juscula occurs abundantly in many habitats around the
island of Guam. Strains that differ morphologically based
on width of cells and trichomes and in secondary metabolite
production are sometimes quite similar based on 16S rDNA
analysis. For example, a population of L. majuscula in
Cocos Lagoon produces majusculamides A and B5 and
diagnostic 13C NMR signals at δ 83.6 for a quaternary
carbon and at δ 69.1 for a methine carbon. Seven ester/
amide carbonyl resonances were observed in the 13C NMR
spectrum. The presence of both functionalities could be
deduced from the IR spectrum displaying strong absorption
bands centered at 1725 and 1645 cm-1. Oxygenated sp3
carbons were suspected because of signals at δ 78.1 and
77.2 in the 13C NMR spectrum, suggesting the presence of
two hydroxy acids in addition to amino acids. Seven partial
structures could be assembled by analysis of 1D and 2D
NMR data (Table 1), i.e., N-methylphenylalanine (N-Me-
Phe), glycine (Gly), proline (Pro), valine (Val), isoleucine
(Ile), 2-hydroxy-3-methylpentanoic acid (Hmp), and a 2,2-
dimethyl-3-hydroxy-7-octynoic acid (Dhoya) unit that has
been identified previously in cyanobacterial metabolites.8
The sequential relationship of the different spin systems
could be established from HMBC experiments (Table 1),
leading to the gross structure as depicted for 1, except for
the linkage between Hmp and Pro cyclizing the molecule,
which at this point had only been assessed to account for
the molecular formula and the last degree of unsaturation.
Evidence for this linkage was found by evaluating NOE
data. The R-H of the Hmp residue (H-27) did not show a
ROESY correlation to the δ-H’s of Pro (H-36), but to a
signal at δ 4.62. However, this could not be unambiguously
assigned to the R-H of Pro (H-33) since its doublet overlaps
with the doublet of doublets of one of the glycine methylene
protons (H-44) (Table 1). 1D NOE experiments were more
malyngamides A and B,6 while a population at Piti Bomb
Holes produces the compounds reported here (<1.5%
sequence divergence).7 A large collection of L. majuscula
was made in J anuary 2000 at Piti Bomb Holes, Guam.
Pitipeptolides A (1) and B (2) were the major secondary
metabolites produced by this organism (UOG strain VP627).
Resu lts a n d Discu ssion
The organic extract of the cyanobacterial collection
afforded a mixture of compounds 1 and 2 after solvent
partition and chromatography on silica. Their separation
was achieved by normal-phase HPLC, yielding both com-
pounds as colorless amorphous solids.
HRFABMS data for the major compound, pitipeptolide
A (1), suggested a molecular formula of C44H65N5O9. The
1H NMR spectrum indicated the peptide nature of 1 by
showing signals for exchangeable protons at δ 6.09, 6.38,
and 7.92 and an N-Me singlet at δ 2.78, arising from
secondary and tertiary amide functionalities, respectively,
in the molecule. It also revealed the presence of a mono-
substituted phenyl group as well as a terminal alkyne
moiety (t at δ 1.96, J ≈ 2.5 Hz) consistent with the
* To whom correspondence should be addressed. (R.E.M.) Tel: (808) 956-
7232. Fax: (808) 956-5908. E-mail: moore@gold.chem.hawaii.edu. (V.J .P.)
Tel: (671) 735-2186. Fax: (671) 734-6767. E-mail: vpaul@uog9.uog.edu.
† University of Hawaii at Manoa.
‡ University of Guam Marine Laboratory.
10.1021/np000456u CCC: $20.00
© 2001 American Chemical Society and American Society of Pharmacognosy
Published on Web 02/13/2001

Pitipeptolides A and B
J ournal of Natural Products, 2001, Vol. 64, No. 3 305
Ta ble 1. NMR Spectral Data for Pitipeptolide A (1) and B (2) at 400 MHz (1H) and 100 MHz (13C) in CDCl3
pitipeptolide A (1)
pitipeptolide B (2)
unit
C/H no.
δH (J in Hz)
δC
HMBCa,b
δH (J in Hz)
δC
Dhoyac/Dhoead
1
2
3
4
5
6
7
8
175.4, s
45.5, s
77.2, d
28.8, t
24.4, t
18.0, t
83.6, s
69.1, d
H-3, H-9, H-10, NH-(1), H-12e
H-9, H-10
H-9, H-10
175.5, s
45.5, s
77.3, d
29.4, t
25.0, t
33.4, t
4.95, dd (9.6, 2.9)
1.58, m, 1.83, m
1.43, m
4.94, dd (10.4, 3.3)
1.47, m, 1.59, m
1.32, m
2.22, m
H-4, H-8
2.05, m
5.75, ddt (17.1, 10.2, 6.9)
4.96, br dd (10.2, 1.7),
5.00, br dd (17.1, 1.7)
1.28, s
138.1, d
115.2, t
1.96, ∼t (∼2.5)
9
10
11
12
13
14
15
NH-(1)
16
1.31, s
1.16, s
19.4, q
22.9, q
171.9, s
53.3, d
29.5, d
16.0, q
20.3, q
H-3, H-10
H-9
H-12, H-25
H-14
H-14
H-12
19.4, q
22.9, q
171.9, s
53.2, d
29.6, d
16.0, q
20.3, q
1.12, s
Val
4.70, dd (9.2, 2.1)
1.76, m
0.90, d (7.1)
0.88, d (6.9)
6.09, d (9.2)
4.70, dd (9.2, 2.3)
1.76, m
0.90, d (7.1)
0.89, d (6.9)
6.09, d (9.2)
N-Me-Phe
172.5, s
65.7, d
33.9, t
H-17, H-18, H-27
H-18, H-25
H-17
172.5, s
65.8, d
33.9, t
17
18
3.85, dd (11.2, 3.8)
3.11, dd (14.2, 11.2),
3.21, dd (14.2, 3.8)
3.84, dd (11.2, 3.9)
3.11, dd (14.3, 11.2),
3.21, dd (14.3, 3.9)
19
137.5, s
128.9, d
128.6, d
126.9, d
39.2, q
169.5, s
78.1, d
37.1, d
24.9, t
11.6, q
14.5, q
170.3, s
61.1, d
31.2, t
H-17, H-18, H-21/23
H-18, H-24/20
H-23/21
137.6, s
128.9, d
128.6, d
126.9, d
39.2, q
169.5, s
78.2, d
37.2, d
24.9, t
11.6, q
14.5, q
170.2, s
61.2, d
31.2, t
20/24
21/23
22
25
26
27
28
29
30
31
32
33
34
35
36
7.11, d (6.9)
7.28, m
7.24, m
7.11, d (6.9)
7.28, m
7.24, m
H-20/24
2.78, s
H-17
2.78, s
Hmp
Pro
Ile
H-27,e H-28e
H-29, H-31
4.93, d (7.1)
1.81, m
1.14, m, 1.58, m
0.89, t (7.4)
0.93, d (7.1)
4.94, d (7.1)
1.81, m
1.15, m, 1.58, m
0.89, t (7.4)
0.93, d (6.9)
H-27, H-30, H-31
H-27, H-30, H-31
H-27, H-29
H-33, H-34, H-38, NH-(4)
4.62, d (7.1)
4.62, d (7.3)
1.97, m, 2.64, m
1.76, m, 1.97, m
3.55, dd (11.9, 9.6),
3.70, m
H-33
H-33
1.96, m, 2.66, m
1.77, m, 1.96, m
3.55, dd (11.7, 9.4),
3.70, m
21.7, t
46.3, t
21.7, t
46.3, t
37
38
39
40
41
42
NH-(4)
43
44
171.8, s
61.0, d
35.1, d
25.8, t
10.8, q
15.8, q
H-38, NH-(5), H-44
171.6, s
60.9, d
35.1, d
25.8, t
10.8, q
15.8, q
4.19, dd (8.9, 8.1)
2.03, m
1.24, m, 1.58, m
0.875, t (7.4)
1.00, d (6.9)
7.92, d (8.1)
4.21, dd (8.9, 8.3)
2.05, m
1.25, m, 1.59, m
0.88, t (7.4)
1.00, d (6.9)
7.95, d (8.3)
H-38, H-41
H-38, H-41, H-42
H-40
H-38, H-40
Gly
170.2, s
41.0, t
H-44, H-3
170.1, s
41.0, t
3.98, d (17.5),
4.62, dd (17.5, 8.9)
6.38, d (8.9)
3.98, d (17.5),
4.61, dd (17.5, 8.9)
6.38, d (8.9)
NH-(5)
a
Protons showing long-range correlation with indicated carbon. b If not indicated otherwise, correlations were observed after optimization
for J CH ) 7 Hz. c Refers to compound 1. Refers to compound 2. e Correlation observed after optimization for J CH ) 4 Hz.
n
d
n
conclusive. Irradiation at δ 4.93 for H-27 resulted in
enhancement of the doublet at δ 4.62 corresponding to
H-33. The NOE between H-27 and H-33 not only supports
the Hmp-Pro linkage, confirming the gross structure for
1, but also discloses the cis geometry of the propyl amide
bond between these two units.9 Upon methanolysis of 1,
methyl esters 3 and 4 were formed, providing additional
support for the gross structure.
HRMS. The presence of all other units was confirmed by
TOCSY experiments and the sequence by HMBC and
ROESY experiments.
The HRMS for pitipeptolide B (2) indicated that the
molecular weight was two mass units higher than that of
pitipeptolide A (1). The NMR spectra revealed that both
1
compounds are closely related. The H NMR spectrum of
2 lacked the triplet at δ 1.96 for H-8 and showed additional
resonances in the olefinic region (δ 4.96, 5.00, and 5.75).
Signals in the 13C NMR spectrum of 2 for sp2 carbons at δ
115.2 and 138.1 instead of acetylenic signals supported that
the Dhoya unit had been substituted by a 2,2-dimethyl-3-
hydroxy-7-octenoic acid (Dhoea) unit in 2, accounting for
the molecular formula of C44H67N5O9 derived from the

306 J ournal of Natural Products, 2001, Vol. 64, No. 3
Luesch et al.
Ta ble 2. Antimycobacterial Disk Diffusion Susceptibility
Assay Results for Compounds 1 and 2
isolation of the chlorinated lipids pitiamide A and B from
cyanobacteria tufts overgrowing coral tips has been re-
ported earlier.15
Diameter (mm) of zone
of growth inhibition for
Exp er im en ta l Section
M. tuberculosis strains
amounta
sample
(µg)
ATTC 25177
ATTC 35818
Gen er a l Exp er im en ta l P r oced u r es. Unless otherwise
stated, 1H and 13C NMR spectra were recorded in CDCl3 at
400 and 100 MHz, respectively. The HSQC experiments were
streptomycin
25
5
1
100
25
100
25
50
15
0
25
10
30
15
55
33
10
15
9
1
optimized for J CH ) 140 Hz, and the HMBC experiments for
nJ CH ) 7 or 4 Hz. HRMS were obtained by FAB, MALDI, or
CI as indicated.
pitipeptolide A (1)
pitipeptolide B (2)
15
10
Biologica l Ma t er ia l. Cyanobacterium VP627 was a L.
majuscula collected at Piti Bomb Holes, Guam, in J anuary
2000. A specimen preserved in formalin has been deposited
at the University of Guam Marine Laboratory.
a
Sample was dissolved in ethanol, applied to each 6 mm disk,
and air-dried prior to placement on the plate.
Extr a ction a n d Isola tion . The freeze-dried organism
VP627 (∼35 g) was extracted with EtOAc-MeOH (1:1) three
times to afford the crude organic extract VP627L (3.69 g).
VP627L was partitioned between EtOAc and H2O and the
concentrated organic phase applied to a Si Bond-Elut column.
After washing with hexane, compounds 1 and 2 were eluted
with hexane-EtOAc (1:3). The fraction was dried and the
mixture separated by normal-phase HPLC (Econosil Silica, 10
µm, 250 × 10 mm, 3.0 mL/min; detection at 250 nm) using an
isocratic system of hexane-EtOAc (3:7) to afford pitipeptolide
B (2) (9.5 mg, tR 23.5 min) and pitipeptolide A (1) (46.9 mg, tR
25.0 min).
Chiral HPLC analysis of the acid hydrolyzates of 1 and
2 revealed the L-configuration of all amino acids and
detected the presence of (2S,3S)-Hmp in both cases. Since
conversion of 3 to the corresponding Mosher esters failed,10
the stereocenter in 1 at C-3 was assigned after hydrogena-
tion of 1 on Pd/C, acid hydrolysis of the resulting product,
tetrahydropitipeptolide A, and isolation of 2,2-dimethyl-3-
hydroxyoctanoic acid (5, Dhoaa). Its negative optical rota-
P itip ep tolid e A (1): colorless amorphous solid; [R]25D -109°
(c 1.0, MeOH); UV (MeOH) λmax (log ꢀ) 202 (4.48) nm; IR (film)
ν
max 3401, 2965, 2872, 1725, 1643 (br), 1514, 1448, 1407, 1373,
1
1284, 1185, 1026 cm-1; H NMR, 13C NMR, and HMBC data,
see Table 1; HRFABMS m/z [M + H]+ 808.4867 (calcd for
C
44H66N5O9, 808.4861).
tion revealed 3S configuration by comparison with data for
the known (S)-Dhoaa (5).11 Closely matching NMR data
suggested the same relative stereochemistry of 1 and 2,
and their identical optical rotations indicated identical
absolute configuration. Furthermore, since the hydrogena-
tion products of 1 and 2 (tetrahydropitipeptolide A and
dihydropitipeptolide B, respectively) were the same, the
configuration of the Dhoea unit in 2 was S as well.
P itip ep tolid e B (2): colorless amorphous solid; [R]25D -109°
(c 1.0, MeOH); UV (MeOH) λmax (log ꢀ) 202 (4.48) nm; IR (film)
ν
max 3401, 2966, 2870, 1731, 1643 (br), 1514, 1455, 1414, 1367,
1279, 1185, 1026 cm-1; H NMR, 13C NMR data, see Table 1;
HRMS (MALDI) m/z [M + H]+ 810.5005 (calcd for C44H68N5O9,
810.5012).
1
Meth a n olysis of 1. Compound 1 (10.0 mg) was dissolved
in 1 mL of MeOH. A 0.1 M solution of NaOMe in MeOH (100
µL) was added to the mixture, which was then stirred at room
temperature for 72 h. The solvent was evaporated and the
residue partitioned between diluted HCl (1 mL, pH 4) and
EtOAc (3 × 2 mL). The organic phase was concentrated and
applied to a SiO2 Sep Pak. Elution was initiated with hexane
followed by hexane solutions containing progressively increas-
ing amounts of EtOAc and final washing with MeOH. Methyl
ester 3 (2.7 mg) eluted first (hexane-EtOAc, 2:1 to 1:1)
followed by unreacted 1 (2.3 mg) (hexane-EtOAc, 1:2 to 1:3),
and methyl ester 4 (3.3 mg) was encountered in the MeOH
fraction.
Pitipeptolides A (1) and B (2) exhibit weak cytotoxicity
against LoVo cells with IC50 values of 2.25 and 1.95 µg/
mL, respectively. Both compounds show activity in the
antimycobacterial diffusion susceptibility assay. Magni-
tudes of growth inhibition for Mycobacterium tuberculosis
strains ATCC 25177 and ATCC 35818 are listed in Table
2 in comparison with a standard drug, streptomycin, which
possesses superior activity. No other antibacterial, anti-
fungal, or protease-inhibiting activity (elastase, papain,
thrombin, trypsin, plasmin, chymotrypsin) was detected,
but both compounds increased elastase activity signifi-
cantly (2.76-fold and 2.55-fold, respectively, at 50 µg/mL).
We had made a similar observation for the lipopeptide
apramide A,12 indicating a possible correlation between the
stimulation of elastase activity and the presence of hydro-
phobic portions in the molecule. This result is not surpris-
ing; other studies have shown that certain unsaturated
long-chain alcohols markedly enhance elastase activity.13
The most distinctive features of 1 and 2, the Dhoya and
Dhoea units, are found in some other marine natural
products such as the kulolides11,14 (Dhoya and Dhoea),
isolated from marine mollusks known for sequestering
cyanobacterial metabolites via their diet, and yanucamides
A and B8 (Dhoya), directly obtained from a cyanobacterial
source. The isolation of compounds 1 and 2 provides more
evidence that Dhoya and Dhoea units are diagnostic for
metabolites of cyanobacterial origin and that they can be
considered biosynthetic signatures. Pitipeptolides A (1) and
B (2) represent the second group of novel secondary
cyanobacterial metabolites from this collection site. The
3: colorless oil; [R]25 -85° (c 0.5, MeOH); UV (MeOH) λmax
D
(log ꢀ) 202 (4.33) nm; IR (film) νmax 3413 (br), 2955, 2872, 1737,
1631, 1525, 1455, 1261, 1214, 1167, 1085, 1008 cm-1; HRMS
(MALDI) m/z [M + Na]+ 481.2674 (calcd for C26H38N2O5 + Na,
481.2673).
4: colorless oil; [R]25 -75° (c 1.0, MeOH); UV (MeOH) λmax
D
(log ꢀ) 202 (4.25) nm; IR (film) νmax 3410 (br), 2955, 2870, 1743,
1637 (br), 1537, 1455, 1385, 1202, 1132, 1038 cm-1; HRMS
(MALDI) m/z [M + Na]+ 436.2424 (calcd for C20H35N3O6 + Na,
436.2418).
Acid Hyd r olysis of 1 a n d 2. Compounds 1 and 2 (0.7 mg
each) were treated with 0.5 mL of 6 N HCl and the suspensions
heated at 110 °C for 12 h. The product mixtures were dried
and analyzed by chiral HPLC for their amino acid contents
[column, Chirex phase 3126 (D) (4.6 × 250 mm), Phenomenex;
solvent, 2 mM CuSO4-MeCN (95:5), except for N-Me-Phe, 2
mM CuSO4-MeCN (85:15); flow rate, 0.8 mL/min; detection
at 254 nm]. The retention times (tR, min) of the authentic
amino acids were Gly (7.1), L-Pro (11.5), D-Pro (22.2), L-Val
(19.3), D-Val (26.1), L-Ile (47.6), D-Ile, (63.0), L-allo-Ile (40.1),
D-allo-Ile (52.0), N-Me-L-Phe (34.0), and N-Me-D-Phe (37.2).
The retention times of the amino acids in the hydrolyzates

Pitipeptolides A and B
J ournal of Natural Products, 2001, Vol. 64, No. 3 307
were 7.1, 11.5, 19.3, 47.6 (solvent mixture 95:5), and 34.0
(solvent mixture 85:15), indicating the presence of Gly, L-Pro,
L-Val, L-Ile, and N-Me-L-Phe. The HPLC traces of both
hydrolyzates were identical.
0.90 (3H, t, J ) 6.7 Hz), 1.20 (3H, s), 1.25 (3H, s), 1.26-1.42
(6H, m), 1.51 (1H, m), 1.58 (1H, m), 3.64 (1H, dd, J ) 10, 2
Hz); CIMS (NH3) m/z 206 [M + NH4]+ 206 (96), 188 [M + NH4
- H2O]+ (40); HRCIMS (NH3) m/z [M + NH4 - H2O]+ 188.1645
(calcd for C10H20O3 + NH4 - H2O, 188.1651).
The stereochemistry of the Hmp unit in 1 and 2 was
determined in a similar manner by chiral HPLC analysis of
the acid hydrolyzates, but using a different stationary phase
[column, CHIRALPAK MA(+) (4.6 × 50 mm), Daicel Chemical
Industries, Ltd.; solvent, 2 mM CuSO4-MeCN (90:10); flow
rate, 1.0 mL/min; detection at 254 nm]. Authentic standards
of the hydroxy acid eluted as follows (tR, min): (2R,3S)-Hmp
(38.2), (2R,3R)-Hmp (46.0), (2S,3R)-Hmp (60.0), and (2S,3S)-
Hmp (75.0). The HPLC profiles of both hydrolyzates showed
a peak at 75.0 min, corresponding to (2S,3S)-Hmp. The amino
acids eluted within the first 15 min under these conditions.
Hyd r ogen a tion of 1 a n d 2. Compounds 1 (10.0 mg) and 2
(2.0 mg) were hydrogenated on 10% Pd/C (catalytic amount)
in MeOH at room temperature for 6 and 3 h, respectively, by
bubbling hydrogen through the solution. Each solution was
filtered through a pad of Celite and concentrated to dryness,
and the residues were applied to a SiO2 Sep Pak. Elution with
hexane-EtOAc (1:1) afforded tetrahydropitipeptolide A (9.5
mg) and dihydropitipeptolide B (1.8 mg), respectively.
Tetr a h yd r op itip ep tolid e A: colorless amorphous solid; Rf
0.30 [hexanes-EtOAc (1:1)]; [R]25D -104° (c 0.15, MeOH); UV
(MeOH) λmax (log ꢀ) 202 (4.48) nm; IR (film) νmax 3401, 2955,
Ack n ow led gm en t. Funding was provided by NCNPDDG
grant CA53001 from the National Cancer Institute. NIH grant
GM 44796 (MBRS program) financially supported one of us
(R.P.). Bioassays were performed by Dr. M. Lieberman,
University of Hawaii at Manoa, Department of Chemistry.
Mass spectral analyses were conducted by the UCR Mass
Spectrometry Facility, Department of Chemistry, University
of California at Riverside.
Refer en ces a n d Notes
(1) J aspars, M.; Lawton, L. A. Curr. Opin. Drug Discovery Dev. 1998, 1,
77-84.
(2) (a) Yasumoto, T.; Murata, M. Chem. Rev. 1993, 93, 1897-1909. (b)
Moore, R. E.; Ohtani, I.; Moore, B. S.; de Koning, C. B.; Yoshida, W.
Y.; Runnegar, M. T. C.; Carmichael, W. W. Gazz. Chim. Ital. 1993,
123, 329-336.
(3) Compilation of Abstracts, 4th International Conference on Toxic
Cyanobacteria, September 27-October 1, 1998, Beaufort, NC.
(4) (a) Harrigan, G. G.; Yoshida, W. Y.; Moore, R. E.; Nagle, D. G.; Park,
P. U.; Biggs, J .; Paul, V. J .; Mooberry, S. L.; Corbett, T. H.; Valeriote,
F. A. J . Nat. Prod. 1998, 61, 1221-1225. (b) Luesch, H.; Yoshida, W.
Y.; Moore, R. E.; Paul, V. J . J . Nat. Prod. 1999, 62, 1702-1706. (c)
Luesch, H.; Yoshida, W. Y.; Moore, R. E.; Paul, V. J .; Mooberry, S. L.
2000, 63, 611-615.
(5) Marner, F.-J .; Moore, R. E.; Hirotsu, K.; Clardy, J . J . Org. Chem.
1977, 42, 2815-2819.
(6) (a) Cardellina, J . H., II; Marner, F.-J .; Moore, R. E. J . Am. Chem.
Soc. 1979, 101, 240-242. (b) Cardellina, J . H., II; Dalietos, D.;
Marner, F.-J .; Mynderse, J . S.; Moore, R. E. Phytochemistry 1978,
17, 2091-2095.
(7) Thacker, R. W.; Paul, V. J ., work in progress.
(8) Sitachitta, N.; Williamson, R. T.; Gerwick, W. H. J . Nat. Prod. 2000,
63, 197-200.
2872, 1731, 1648 (br), 1514, 1455, 1367, 1190, 1026 cm-1
;
HRMS (MALDI) m/z [M + H]+ 812.5169 (calcd for C44H70N5O9,
812.5168).
Dih yd r op itip ep tolid e B: colorless amorphous solid; Rf
0.30 [hexane-EtOAc (1:1)]; [R]25 -106° (c 0.65, MeOH); UV
D
(MeOH) λmax (log ꢀ) 202 (4.48) nm; IR (film) νmax 3408, 2961,
2870, 1729, 1644 (br), 1504, 1455, 1371, 1190, 1030 cm-1
;
HRMS (MALDI) m/z [M + H]+ 812.5161 (calcd for C44H70N5O9,
812.5168).
The 1H NMR spectra of tetrahydropitipeptolide A and
dihydropitipeptolide B were identical.
(9) Kofron, J . L.; Kuzmicˇ, P.; Kishore, V.; Gemmecker, G.; Fesik, S. W.;
Rich, D. H. J . Am. Chem. Soc. 1992, 114, 2670-2675.
(10) Ohtani, I.; Kusumi, T.; Kashman, Y.; Kakisawa, H. J . Am. Chem.
Soc. 1991, 113, 4092-4096.
Acid Hyd r olysis of Tetr a h yd r op itip ep tolid e A. The
hydrogenation product of 1 (9.0 mg) was subjected to acid
hydrolysis (6 N HCl, 110 °C, 12 h). The hydrolyzate was dried
under N2 and the residue partitioned between 1 N HCl and
EtOAc. The organic phase was concentrated and the remainder
purified by reversed-phase HPLC (YMC-Pack ODS-AQ-323,
5 µm, 250 × 10 mm, 2.0 mL/min; PDA detection from 190 to
540 nm) using a MeCN-0.02 N TFA linear gradient (0-100%
over 55 min after a 5 min period at 0% MeCN) for elution.
This afforded (2S,3S)-Hmp (1.1 mg) at tR 28.1 min and (S)-
Dhoaa (5) (1.0 mg) at tR 40.8 min.
(11) Nakao, Y.; Yoshida, W. Y.; Szabo, C. M.; Baker, B. J .; Scheuer, P. J .
J . Org. Chem. 1998, 63, 3272-3280.
(12) Luesch, H.; Yoshida, W. Y.; Moore, R. E.; Paul, V. J . J . Nat. Prod.
2000, 63, 1106-1112.
(13) Ashe, B. M.; Zimmerman, M. Biochem. Biophys. Res. Commun. 1977,
75, 194-199.
(14) Reese, M. T.; Gulavita, N. K.; Nakao, Y.; Hamann, M. T.; Yoshida,
W. Y.; Coval, S. J .; Scheuer, P. J . J . Am. Chem. Soc. 1996, 118,
11081-11084.
(15) Nagle, D. G.; Park, P. U.; Paul, V. J . Tetrahedron Lett. 1997, 38,
6969-6972.
(S)-2,2-Dim eth yl-3-h yd r oxyocta n oic a cid (5): colorless
1
oil; [R]25 -24° (c 0.33, MeOH); H NMR (300 MHz, CDCl3) δ
NP000456U
D