Complete structure elucidation of shishididemniols, complex lipids with tyramine-derived tether and two serinol units, from a marine tunicate of the family Didemnidae

Article abstract of DOI:10.1021/jo062013m

(Chemical Equation Presented) Two new serinolipid derivatives, shishididemniols A (1) and B (2), were isolated as antibacterial constituents of a tunicate of the family Didemnidae. The structure of 1 was elucidated by interpretation of spectral data and t

Full text of DOI:10.1021/jo062013m

Complete Structure Elucidation of Shishididemniols, Complex
Lipids with Tyramine-Derived Tether and Two Serinol Units, from
a Marine Tunicate of the Family Didemnidae
Hirotsugu Kobayashi,† Jun’ichiro Ohashi,† Tsuyoshi Fujita,‡ Takashi Iwashita,‡
Yoichi Nakao,† Shigeki Matsunaga,*,† and Nobuhiro Fusetani†
Laboratory of Aquatic Natural Products Chemistry, Graduate School of Agricultural and Life Sciences,
The UniVersity of Tokyo, Bunkyo-ku, Tokyo 113-8657, Japan, and Suntory Institute for Bioorganic
Research, Wakayamadai, Shimamoto-cho, Mishima-gun, Osaka 618-8503, Japan
assmats@mail.ecc.u-tokyo.ac.jp
ReceiVed September 29, 2006
Two new serinolipid derivatives, shishididemniols A (1) and B (2), were isolated as antibacterial
constituents of a tunicate of the family Didemnidae. The structure of 1 was elucidated by interpretation
of spectral data and the application of the modified Mosher method to 1 and its suitable degradation
products. Compound 2 was the chlorohydrin of 1. Compounds 1 and 2 exhibited antibacterial activity
against fish pathogenic bacterium Vibrio anguillarum.
Introduction
This paper describes the isolation, structure elucidation and
antimicrobial activitiy of these compounds.
Fish and shellfish diseases are the most serious concerns for
aquaculture today.1 The popular action taken in fish farms is
chemotherapy using chemical substances such as antibiotics and
synthetic organic compounds to control these diseases.1 However
these chemical substances pose environmental and hygienic
problems.1 We have been searching for antimicrobial metabolites
against fish pathogenic bacteria from marine natural products
that are expected to be environmentally benign. As a part of
the study, we have discovered novel bromotyrosine derivatives
from the marine sponge Hexadella sp.2 that exhibited antibacte-
rial activity against the fish pathogenic bacterium Aeromonas
hydrophila. Subsequently, we found that the extract of a tunicate
of the family Didemnidae collected in southern Japan exhibited
activity against the bacterium Vibrio anguillarum, which causes
vibriosis in fish.3 Bioassay-guided fractionation afforded two
novel serinolipid derivatives, shishididemniols A (1) and B (2).
Results and Discussion
† University of Tokyo.
‡ Suntory Institute.
The EtOH extract of the tunicates (780 g) was partitioned
between CHCl3 and H2O, and the aqueous layer was further
extracted with n-BuOH. The n-BuOH fraction was separated
by ODS flash chromatography followed by reversed-phase
HPLC to afford shishididemniol A (1) and B (2) in yields of
(1) Shao, Z. J. AdV. Drug. DeliV. ReV. 2001, 50, 229-243.
(2) Matsunaga, S.; Kobayashi, H.; van Soest, R. W. M.; Fusetani, N. J.
Org. Chem. 2005, 70, 1893-1896.
(3) Roberts, R. J. In Bacterial Disease of Fish; Inglis, V., Roberts, R. J.,
Bromage, N. R., Eds.; Blackwell Science: Oxford, 1993; pp 109-121.
10.1021/jo062013m CCC: $37.00 © 2007 American Chemical Society
Published on Web 01/25/2007
1218
J. Org. Chem. 2007, 72, 1218-1225

Structure Elucidation of Shishididemniols
TABLE 1. 1H and 13C NMR Data for Shishididemniols A (1) and
395 mg (5.1 × 10-2 % wet weight) and 152 mg (1.9 × 10-2
%
a
B (2) in DMSO-d6
wet weight), respectively.
1
2
The molecular formula of shishididemniol A (1) was estab-
position δH (mult, J Hz)
δ
C (multb)
δ
H (mult, J Hz)
δ
C (multb)
lished as C45H81N3O11 on the basis of NMR and HRESIMS
1
data. Interpretation of the H NMR spectrum (in DMSO-d6)
1
2
173.9 (C)
70.8 (CH)
34.3 (CH2) 1.43 (m)
1.57 (m)
20.4 (CH2) 1.37 (m)
1.44 (m)
33.7 (CH2) 1.31 (m)
1.38 (m)
173.9 (C)
70.9 (CH)
34.4 (CH2)
together with HSQC data showed the presence of an N-
substituted methylene (δH 3.12 and 3.31, δC 46.2), two
nitrogenous methines (δH 3.23, δC 52.0; δH 4.02, δC 50.2), five
oxygenated methylenes (δH 3.40 (2H), δC 70.7; δH 3.44 and
3.50, δC 67.5; δH 3.48 and 3.56, δC 58.7; δH 3.93 and 3.95, δC
66.5; δH 3.46 and 3.49, δC 60.1), four oxymethines (δH 3.80,
δC 70.8; δH 3.19, δC 68.7; δH 3.33, δC 69.5; δH 4.55, δC 70.8),
two epoxide methines (δH 2.69, δC 60.5; δH 2.87, δC 56.2), a
triplet methyl (δH 0.97, δC 9.90), a methylene (δH 2.10, δC 28.4),
two aromatic signals [δH 6.89 (2H), δC 114.1 (2C); δH 7.22
(2H), δC 127.1 (2C)], two amide protons (δ 7.51 and 7.74),
and seven exchangeable protons (δ 4.19, 4.83, 4.85, 5.24, 5.41,
3.80 (br)
1.42 (m)
1.55 (m)
1.31 (m)
1.42 (m)
1.29 (m)
1.40 (m)
3.19 (br)
2.69 (dd, 4.1,
8.2)
2.87 (ddd, 4.1,
4.1, 8.2)
1.28 (m)
1.54 (m)
3.81 (br)
3a
3b
4a
4b
5a
5b
6
21.0 (CH2)
32.6 (CH2)
68.7 (CH)
60.5 (CH)
3.46 (m)
3.27 (t, 4.4)
70.9 (CH)
75.6 (CH)
7
8
56.2 (CH)
3.99 (dt, 8.8,
4.4)
66.2 (CH)
34.4 (CH2)
9a
9b
10-14
15
16
17
18-25
26
27.9 (CH2) 1.67 (m)
1.77 (m)
1.15-1.45 (m)
1.21-1.34 (m)
3.33 (m)
1.21-1.34 (m)
1.15-1.45 (m)
1.27 (m)
28.9-29.3 1.17-1.31 (m) 29.4-29.8
37.2 (CH2) 1.22-1.34 (m) 37.7 (CH2)
1
5.41, and 7.80). The remainder of the H NMR signals were
attributed to long methylene chains (δ 1.15-1.45). In addition,
two non-protonated sp2 carbon signals (δ 135.6 and δC 157.7)
and two carbonyl carbon signals (δ 173.0 and 173.9) were
observed in the 13C NMR spectrum (Table 1).
69.5 (CH)
3.33 (m)
69.5 (CH)
37.2 (CH2) 1.22-1.34 (m) 37.7 (CH2)
28.9-29.3 1.17-1.31 (m) 29.4-29.8
25.5 (CH2) 1.27 (m)
28.9 (CH2) 1.49 (m)
25.5 (CH2)
28.6 (CH2)
27
1.49 (tt, 7.6,
7.6)
Interpretation of 2D NMR data led to substructures a-c
(Figure 1A). In substructure a, the propionamide moiety (C-
12′ to C-14′) was identified from the COSY and HMBC data.
The COSY spectrum revealed spin systems for C-9′-C10′(-NH-
10′)-C-11′-OH-11′ and NH-1′-C-1′-C-2′-OH. The connectivity
between N-10′ and C-12′ was determined by HMBC cross-
peaks, NH-10′/C-12′ and H-10′/C-12′. 1H and 13C NMR
chemical shift values clearly indicated the presence of a para-
substituted phenyl ether moiety. The ether linkage was estab-
lished by the HMBC cross-peak, H-9′/C-6′, while further HMBC
correlations, H-2′/C-3′, H-2′/C-4′(8′), and H-4′(8′)/C-2′, dem-
onstrated the connectivity between C-2′ and C-3′. The connec-
tivities from OH-2 to H2-4 and from OH-6 to H2-9 were assigned
on the basis of the COSY data. Further analysis of the COSY
and TOCSY data indicated that the C-6 oxymethine proton was
adjacent to H2-5, which was in turn correlated with H2-4. The
1H-1H coupling constant between H-7 and H-8 (J ) 4.1 Hz)
revealed that C-7 and C-8 were a part of a cis-epoxide. HMBC
cross-peaks, NH-1′/C-1, H-2/C-1, and OH-2/C-1, showed the
connectivity between C-1 and N-1′.
Substructure b comprises the terminus of an alkyl chain
attached to a serinol ether.4,5 The serinol unit (O-C-29-C-30-
C-31-OH-31) was clearly defined by the COSY and HSQC data.
The chemical shift value of C-30 at δC 52.0 and the ninhydrin-
positive property of 1 implied that C-30 was substituted by a
free amino group. The ether linkage was established by HMBC
cross-peaks from H2-28 to C-29.
Substructure c contains an oxymethine group placed in the
middle of a long methylene chain. The COSY spectrum showed
that C-16 oxymethine proton was coupled to OH-16 and two
pairs of methylene protons (H2-15 and H2-17). Therefore,
substructure c should be inserted between substructure a and b
via methylene chains. The location of substructure c in the alkyl
chain was determined by analysis of FAB-MS/MS data of 1.
Notable fragment ions observed at m/z 258 and 288 allowed us
to locate the hydroxyl group at C-16 (Figure 1B).
28
29a
29b
3.40 (t, 7.6)
3.44 (m)
3.50 (dd, -10.5,
4.8)
3.23 (m)
3.48 (m)
3.56 (m)
70.7 (CH2) 3.39 (m)
67.5 (CH2) 3.43 (m)
3.49 (m)
70.7 (CH2)
67.7 (CH2)
30
31a
31b
52.0 (CH)
58.7 (CH2) 3.48 (m)
3.55 (dd, 4.7,
3.20 (m)
52.0 (CH)
59.0 (CH2)
10.9)
46.2 (CH2) 3.13 (m)
3.31 (m)
1′a
1′b
2′
3.12 (m)
3.31 (m)
4.55 (br)
46.1 (CH2)
70.8 (CH)
135.6 (C)
4.55 (br)
70.9 (CH)
135.6 (C)
127.1 (CH)
114.1 (CH)
157.7 (C)
3′
4′, 8′
5′, 7′
6′
7.22 (d, 8.7)
6.89 (d, 8.7)
127.1 (CH) 7.22 (d, 8.8)
114.1 (CH) 6.89 (d, 8.8)
157.7 (C)
9′a
3.93 (dd, -9.9,
5.9)
3.95 (dd, -9.9,
5.7)
4.02 (m)
3.46 (m)
3.49 (m)
66.5 (CH2) 3.93 (dd, -9.8, 66.4 (CH2)
6.0)
3.95 (dd, -9.8,
5.4)
9′b
10′
50.2 (CH)
4.03 (m)
50.2 (CH)
60.1 (CH2)
11′a
11′b
12′
13′
14′
OH-2
OH-6
OH-7
OH-16
60.1 (CH2) 3.46 (m)
3.50 (m)
173.0 (C)
28.4 (CH2) 2.10 (q, 7.7)
9.90 (CH3) 0.97 (t, 7.7)
5.48 (br)
173.1 (C)
28.4 (CH2)
9.91 (CH3)
2.10 (q, 7.2)
0.97 (t, 7.2)
5.41 (d, 4.1)
4.85 (br)
4.19 (br)
4.82 (br)
4.19 (br)
7.61 (br)
4.83 (br)
7.50 (t, 5.8)
5.42 (br)
7.75 (d, 8.3)
4.82 (br)
4.19 (br)
NH2-30 7.80 (br)
OH-31
NH-1′
OH-2′
NH-10′ 7.74 (d, 7.8)
OH-11′ 4.83 (br)
5.24 (br)
7.51 (t, 5.9)
5.41 (d, 4.1)
a 600 MHz for 1H NMR and 150 MHz for 13C NMR. b Multiplicity from
HSQC experiment.
The stereochemistry of C-6 with respected to that of C-7 in
1 could not be determined by analysis of NMR data.6,7 This
(6) Sugiyama, T.; Yamashita, K. Agric. Biol. Chem. 1980, 44, 1983-
1984.
(4) Gonza´lez, N.; Rodriguez, J.; Jime´nez, C. J. Org. Chem. 1999, 64,
5705-5707.
(5) Mitchell, S. S.; Rhodes, D.; Bushman, F. D.; Faulkner, D. J. Org.
Lett. 2000, 2, 1605-1607.
(7) Graham, S. M.; Prestwich, G. D. J. Org. Chem. 1994, 59, 2956-
2966.
(8) Righi, G.; Ronconi, S.; Bonini, C. Eur. J. Org. Chem. 2002, 1573-
1577.
J. Org. Chem, Vol. 72, No. 4, 2007 1219

Kobayashi et al.
FIGURE 1. (A) Partial structures of shishididemniol A (1). (B) FAB-MS/MS analysis of 1.
SCHEME 1 a
a Reagents and conditions: (a) Cbz-Cl, Et3N, CHCl3/MeOH (3:1), rt, 16 h; (b) MgBr2 OEt2, Et2O, rt, 3 h; (c) 2,2-dimethoxpropane, PPTS, CH2Cl2, rt, 1
h; (d) HCl/MeOH, 100 °C, 4 h; (e) hydrazine, 120 °C, 12 h.
was shown by converting 1 to the 6,7-O-isopropylidene deriva-
tives 5, via the N-Cbz-protected bromohydrin 4 (Scheme 1).8
NOESY cross-peaks observed between one acetonide methyl
signal (δH 1.40) and H-6 and between another methyl signal
(δH 1.37) and H-7 showed the trans-relationship for H-6 and
H-7. Therefore, the relative stereochemistry from C-6 to C-8
in 1 was assigned as threo-cis-2,3-epoxy alcohol (6S*,7R*,8S*).
The absolute stereochemistry of C-2′ and C-10′ in 1 was
envisaged to be determined by application of the modified
Mother’s method9,10 to compound 6, which was expected to be
obtained by acidic hydrolysis of 1. We used (R)-2-amino-1-
phenylethanol, which was commercially available, and (S)-2-
amino-3-phenoxy-1-propanol (9) as model compounds to assign
the stereochemistry of C-2′ and C-10′ in 6. Compound 9 was
(9) Ohtani, I.; Kusumi, T.; Kashman, Y.; Kakisawa, H. J. Am. Chem.
Soc. 1991, 113, 4092-4096.
(10) Seco, J. M.; Latypov, Sh. K.; Quin˜oa´, E.; Riguera, R. J. Org. Chem.
1997, 62, 7569-7574.
1220 J. Org. Chem., Vol. 72, No. 4, 2007

Structure Elucidation of Shishididemniols
FIGURE 2. 1H NMR chemical shift values for 10a/10b, 11a/11b, and 12a/12b.
SCHEME 2 a
The absolute stereochemistry of C-2′ and C-10′ in 6 was
determined by comparison of 1H NMR spectra of 10a and 10b
with those of 11a and 11b and those of (R)- and (S)-MTPA
derivatives of (R)-2-amino-1-phenylethanol (12a and 12b,
respectively) (Figure 2). 1H NMR chemical shift values of H2-
1′ and H-2′ of 10a and 10b were in remarkable agreement with
those of the corresponding signals of 12a and 12b, respectively,
whereas the chemical shift values of H2-9′, H-10′, and H2-11′
agreed well with those of the corresponding signals of 12a and
12b, respectively. Therefore, the absolute stereochemistry of
C-2′ and C-10′ was assigned as R and S, respectively (Figure
2).
a Reagents and conditions: (a) phenol, DEAD, PPh3, toluene, 80 °C, 18
h; (b) CH2Cl2/TFA (1:1), rt, 2 h; (c) (S)-or (R)-MTPA-Cl, pyridine, rt, 30
min.
The absolute stereochemistry of C-2, C-6, and C-30 in 1 was
determined by application of the modified Mosher method to 1
and 5. Treatment of 1 with (S)-MTPACl and (R)-MTPACl
yielded the (R)-MTPA derivative (13a) and (S)-MTPA deriva-
tive (13b), respectively. Analysis of ∆δS-R values between 13a
and 13b allowed the assignment of 6S and 30S (Figure 3). The
negative ∆δS-R values observed for H2-3, H2-4, and H2-5
implied the 2S-stereochemistry. In order to exclude the effect
of the C-6 MTPA ester, the acetonide 5 was converted to the
(R)-MTPA derivative (14a) and (S)-MTPA derivative (14b). The
negative ∆δS-R values observed for H2-3 to H2-8 signals
confirmed the 2S-stereochemistry (Figure 3).
Because C-16 was too far from the nearest functionalized
carbon, it was not possible to detect the effects of MTPA esters
on chemical shifts of assigned proton signals, e.g., H-8 and H2-
9. This problem was hampered by esterification with 2NMA
(2-naphthylmethoxyacetic acid), which was reported to exhibit
anisotropic effects to more distant protons.15 In order to avoid
confusion in assignments, we intended to introduce one 2NMA
group to a derivative of 1. For that purpose the bromohydrin 4
was treated with NaIO4 followed by reduction with NaBH4 to
obtain 15, whose two primary alcohols were protected with
TBDPS groups16,17 to yield 16 (Scheme 3). The secondary
alcohol in 16 was esterified with (R)-2NMA or (S)-2NMA in
the presence of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide
synthesized as outlined in Scheme 2. N-Boc-L-serine methyl
ester was converted to a chiral alcohol 7 in two steps according
to Garner’s procedure.12 Treatment of 7 with triphenylphosphine
and diethyl azodicarboxylate at 80 °C in toluene with phenol
provided the phenyl ether 8,13,14 which was deprotected with
TFA to yield the expected compound 9. The (R)- and (S)-MTPA
derivatives of (R)-2-amino-1-phenylethanol and those of com-
1
pound 9 gave characteristic H NMR profiles, indicating that
the stereochemistry at C-2′ and C-10′ in 6 could be determined
by the modified Mosher method. Acidic methanolysis product
of 1 was separated by ODS column chromatography to afford
a fraction that contained UV-absorbing polar material. This
1
material, which gave a single set of H NMR signals, was
converted to the (R)- and (S)-MTPA derivatives. To our
disappointment H2-1′ and H-2′ were doubled, implying that the
benzyl carbon (C-2′) in 6 was racemized by acid and what we
isolated was a 1:1 mixture of 6 and 6a. As a method to cleave
amide bonds under a milder condition, we chose hydrazinolysis.
Compound 1 was hydrazinolyzed11 to yield a fraction that
contained 6, which was then converted to the (R)- and (S)-
MTPA derivatives (10a and 10b, respectively). Only one set
1
of signals were observed in H NMR spectra of 10a and 10b.
(11) Akabori, S.; Ohno, K.; Narita, K. Bull. Chem. Soc. Jap. 1952, 25,
214-218.
(15) Kusumi, T.; Takahashi, H.; Hashimoto, T.; Kan, Y.; Asakawa, Y.
Chem. Lett. 1994, 1093-1094.
(16) Green, T. W.; Wuts, P. G. M. Protecting Groups in Organic
Synthesis, 3rd ed.; Wiley-Interscience: New York, 1999.
(17) Matsunaga, S.; Liu, P.; Celatka, C. A.; Panek, J. S.; Fusetani, N. J.
Am. Chem. Soc. 1999, 121, 5605-5606.
(12) Garner, P.; Park, J. M. J. Org. Chem. 1987, 52, 2361-2364.
(13) Mitsunobu, O. Synthesis 1981, 1-28.
(14) Pave´, G.; Usse-Versluys, S.; Viaud-Massuard, M.; Guillaumet, G.
Org. Lett. 2003, 5, 4253-4256.
J. Org. Chem, Vol. 72, No. 4, 2007 1221

Kobayashi et al.
FIGURE 3. (A) Distribution of ∆δS-R values for the MTPA derivatives 13a/13b. (B) Distribution of ∆δS-R values for the MTPA derivatives
14a/14b. (C) Distribution of the 2NMA esters of ∆δR-S values for 17a/17b.
SCHEME 3 a
1H NMR. Therefore, the absolute stereochemistry of 2 was
determined to be 2S,6S,7R,8R,16R,30S,2′R,10′S.
Two classes of related serinolipid derivatives, didemniser-
inolipids4 and cyclodidemniserinol trisulfate,5 have been reported
from tunicates of the genus Didemnum. The structure of
didemniserinolipid B, whose serinol moiety was in the S
configuration, was determined by total synthesis.20 Shishidi-
demniols, didemniserinolipids, and cyclodidemniserinol trisul-
fate all possess C28-polyketide acid in the central part of the
molecule. Unlike didemniserionlipids and cyclodidemniserinol
trisulfate, shishididemniols A and B had two serinol and an
oxygenated tyramine moieties and were not sulfated.
a Reagents and conditions: (a) NaIO4, MeOH/H2O (4:1), rt, 1 h; then
NaBH4, rt, 15 min; (b) TBDPSCl, pyridine, rt, overnight; (c) (R)- or (S)-
2NMA, EDC, DMAP, DMAP‚Cl, CH2Cl2, rt, overnight.
Shishididemniols A and B showed antibacterial activity in
disk agar diffusion assay against the fish pathogenic bacterium
V. anguillarum (20 µg/ 6.5 mm φ disk zone of inhibition; 8
and 7 mm for 1 and 2, respectively).
hydrochloride (EDC), DMAP, and DMAP‚HCl18 in CH2Cl2 to
give (R)- or (S)-2NMA derivatives (17a and 17b, respectively).
1
Comparison of H NMR data of 17a and 17b showed that the
absolute configuration at C-16 was R (Figure 3). Therefore, the
stereochemistry of 1 was determined to be 2S,6S,7R,8S,16R,-
30S,2′R,10′S.
Experimental Section
Animal Material. The tunicate was collected by hand using
SCUBA at depths of 5-15 m off Kushizaki in Shishijima (32° 15′
N, 130° 14′ E) of the Amakusa Islands and was kept frozen at
-20 °C until processed. The tunicate was identified as a member
of the family Didemnidae on the basis of the presence of
characteristic spicules.21
Shishididemniol B (2) had a molecular formula of C45H82-
ClN3O11 as established by HRESIMS, which implied the
presence of one less unsaturation equivalent than 1. The 1H and
13C NMR spectra of 2 were almost superimposable on those of
1 except for the replacement of the epoxide group by an
oxymethine (δH 3.27, δC 75.6) and a chlorinated methine (δH
3.99, δC 66.2), suggesting that 2 was a HCl adduct of 1.
Interpretation of NMR and FAB-MS/MS data of 2 confirmed
that 2 was the 7-hydroxy-8-chloro-derivative of 1.
Extraction and Isolation. The frozen tunicates (780 g wet
weight) were extracted four times with EtOH, and the combined
extracts were concentrated and partitioned between water and
CHCl3. The aqueous layer was further extracted with n-BuOH. The
n-BuOH fraction was separated by ODS flash chromatography with
aqueous MeOH. The fraction eluted with 70% MeOH was separated
by ODS HPLC (72% MeOH containing 0.2 M NaClO4) to afford
shishididemniols A (1, 395.4 mg, 5.1 × 10-2 % based on wet
weight) and B (2, 151.6 mg, 1.9 × 10-2 % based on wet weight).
In order to determine the stereochemical relationship between
C-6 and C-7 in 2, its N-Cbz derivative 18 was converted to the
acetonide 19 (Scheme 4). On the basis of NOESY correlations
observed between one singlet methyl signal (δH 1.38) and H-6
and between another singlet methyl signal (δH 1.37) and H-7,
the relative stereochemistry of C-6 and C-7 in 2 was assigned
as identical with that in 1. Treatment of 18 with potassium
Shishididemniol A (1). Colorless oil; [R]20.5 -33.7 (c 1.00,
D
MeOH); UV (MeOH) λmax 275 nm (ꢀ 1195), 281 (1017); IR (film)
ν
max 3428, 1655 cm-1; HRESIMS m/z 840.59682 (M + H)+ (calcd
for C45H82N3O11, ∆ +1.89 mmu). 1H and 13C NMR data, see Table
1. HMBC correlations (DMSO-d6) H-2/C-1; H-7/C-6, 8; H-8/C-7,
9; H-27/C-26, 28; H-28/C-27, 29; H-29a/C-28, 30, 31; H-29b/C-
carbonate19 in MeOH afforded the epoxide 3 ([R]20 -17.2 (c
D
0.10 MeOH)), which was indistinguishable from compound 3
derived from 1 ([R]20D -17.6 (c 0.10 MeOH)) in the [R]D and
(20) Kiyota, H.; Dixon, D. J.; Luscombe, C. K.; Hettstedt, S.; Ley, S.
V. Org. Lett. 2002, 4, 3223-3226.
(21) Nishikawa, T. In Guide to Seashore Animals of Japan with Color
Plates and Keys, II; Nishimura, S., Ed; Hoiku-sha: Osaka, 1995; pp 573-
610.
(18) Boden, E. P.; Keck, G. E. J. Org. Chem. 1985, 50, 2394-2395.
(19) Norman, B. H.; Hemsheidt, T.; Schults, R. M.; Andis, S. L. J. Org.
Chem. 1998, 63, 5288-5294.
1222 J. Org. Chem., Vol. 72, No. 4, 2007

Structure Elucidation of Shishididemniols
SCHEME 4 a
a Reagents and conditions: (a) Cbz-Cl, Et3N, CHCl3/MeOH (3:1), rt, 16 h; (b) 2,2-dimethoxpropane, PPTS, CH2Cl2, rt, 1 h; (c) K2CO3, MeOH, rt, 1.5 h.
28, 30, 31; H-1′a/C-1, 2′, 3′; H-1′b/C-1, 2′, 3′; H-2′/C-3′, 4′, 8′;
H-4′, 8′/C-2′, 4′, 5′, 6′, 7′, 8′; H-5′, 7′/C-3′, 5′, 6′, 7′; H-9′a/C-6′,
10′, 11′; H-9′b/C-6′, 10′, 11′; H-10′/C-9′, 11′, 12′; H-13′/C-12′, 14′;
H-14′/C-12′, 13′; NH-1′/C-1, 1′; OH-2′/C-1′, 2′, 3′; NH-10′/C-10′,
11′, 12′.
Preparation of Acetonide 5. To the mixture of 4 (10 mg) and
PPTS (pyridinium p-toluenesulfonate) (2.4 mg) in CH2Cl2 (100 µL)
was added 2,2-dimethoxypropane (20 µL). The reaction mixture
was stirred at room temperature for 1 h and was then evaporated.
The residue was partition between EtOAc and H2O. The organic
layer was purified by ODS HPLC (gradient elution of 80-100%
aqueous MeOH) to give 5 (6 mg): ESIMS m/z 1094.33124 and
Shishididemniol B (2). Colorless oil; [R]20.5 -19.4 (c 1.00,
D
MeOH); UV (MeOH) λmax 275 nm (ꢀ 1222), 281 (1043); IR (film)
1
1096.36362 (M + H)+. H NMR (600 MHz, CD3OD) 7.39-7.25
ν
max 3429, 1642 cm-1; HRESIMS m/z 876.57161 (M + H)+ (calcd
(m, 5H), 7.30 (d, J ) 8.7, 2H), 6.93 (d, J ) 8.7, 2H), 5.08 (d, J )
-12.6, 1H), 5.06 (d, J ) -12.6, 1H), 4.70 (dd, J ) 7.8, 4.6, 1H),
4.22 (m, 1H), 4.07 (br, 1H), 4.05 (dd, J ) 5.5, 2.7, 2H), 3.99 (m,
2H), 3.78 (m, 1H), 3.69 (d, J ) 5.5, 2H), 3.64 (dd, J ) 7.7, 2.7,
1H), 3.57 (m, 2H), 3.52-3.40 (m, 6H), 3.35 (m, 1H), 2.24 (q, J )
7.5, 2H), 1.90-1.24 (m, 42H), 1.40 (s, 3H), 1.37 (s, 3H), 1.12 (t,
J ) 7.5, 3H).
for C45H8335ClN3O11, ∆ -1.21 mmu). H and 13C NMR data, see
Table 1. HMBC correlations (DMSO-d6) H-2/C-1, 3, 4; H-8/C-7,
9; H-27/C-26, 28; H-28/C-26, 27, 29; H-29a/C-28, 30, 31; H-29b/
C-28, 30, 31; H-30/C-29, 31; H-31a/C-29, 30; H-1′a/C-1, 2′, 3′;
H-1′b/C-1, 2′, 3′; H-2′/C-1′, 3′, 4′, 8′; H-4′, 8′/C-2′, 4′, 5′, 6′, 7′,
8′; H-5′, H-8′/C-3′, 4′, 5′, 6′, 7′, 8′; H-9′a/C-6′, 10′, 11′; H-9′b/C-
6′, 10′, 11′; H-10′/C-9′, 11′, 12′; H-11′a/C-9′, 10′; H-11′b/C-9′, 10′;
H-13′/C-12′, 14′; H-14′/C-12′, 13′.
1
Methanolysis of 1. A 60 mg portion of 1 was dissolved in 5 N
HCl/MeOH (1:4), the solution was heated at 100 °C for 4 h, and
the reaction mixture was cooled and dried. The residue was
separated by ODS flash chromatography with H2O, 20%, 60%, 80%
Preparation of N-Cbz Derivative 3. To a solution of 1 (50 mg,
0.06 mmol) in CHCl3/MeOH (3:1, 0.4 mL) were added triethy-
lamine (66.4 µL) and Cbz-Cl (17 µL). After stirring for 16 h at
room temperature, the reaction mixture was evaporated, suspended
in H2O, and extracted with EtOAc to give 3 (45 mg, 77.1%):
1
and MeOH to give the mixture of 6 and 6a (11.1 mg). The H
NMR data of 6a was identical with that of 6 (vide infra).
Hydrazinolysis of 1. A 20 mg portion of 1 was dissolved in
hydrazine (100 µL), the solution was heated at 120 °C for 15 h,
and the reaction mixture was cooled and freeze-dried. The residue
was separated by ODS flash chromatography with 20% aqueous
MeOH and MeOH to give crude 6 (5.3 mg): 1H NMR (600 MHz,
CD3OD) 7.34 (d, J ) 8.7, 2H), 7.07 (d, J ) 8.7, 2H), 4.41 (dd, J
) 8.0, 5.7, 1H), 4.27 (dd, J ) -10.1, 4.2, 1H), 2.52 (dd, J ) -10.1,
7.1, 1H), 3.88 (dd, J ) -11.5, 4.6, 1H), 3.81 (dd, J ) -11.5, 6.0,
1H), 3.66 (m, 1H), 3.06 (d, 7.8, 2H).
1
ESIMS m/z 974.37505 (M + H)+. H NMR (600 MHz, CD3OD)
7.35-7.27 (m, 5H), 7.29 (d, J ) 8.7, 2H), 6.93 (d, J ) 8.7, 2H),
5.09 (d, J ) -12.3, 1H), 5.06 (d, J ) -12.3, 1H), 4.71 (dd, J )
7.7, 4.6, 1H), 4.22 (m, 1H), 4.05 (dd, J ) 4.8, 3.3, 2H), 3.99 (m,
1H), 3.78 (m, 1H), 3.69 (d, J ) 6.0, 2H), 3.57 (m, 2H), 3.51-3.40
(m, 6H), 3.39-3.33 (m, 2H), 2.98 (ddd, J ) 7.8, 4.4, 4.4 1H),
2.82 (dt, J ) 4.4, 8.5, 1H), 2.24 (q, J ) 7.8, 2H), 1.73-1.25 (m,
42H), 1.12 (t, J ) 7.8, 3H).
Preparation of Bromohydrin 4. To a solution of 3 (40 mg) in
dry Et2O (2 mL) was added MgBr2‚OEt2 (90 mg). The solution
was stirred at room temperature for 4 h and was then evaporated.
The residue was separated by ODS HPLC (gradient elution of 50-
60% aqueous MeCN) to afford 4 (21 mg): ESIMS m/z 1054.38330
Preparation of (R)- and (S)-MTPA Derivatives of 6 (10a and
10b). To a solution of crude 6 (1.5 mg) in pyridine (30 µL) was
added (S)-MTPACl (20 µL). The residue was partitioned between
EtOAc and H2O with 0.1 M Na2CO3.The organic layer was purified
by ODS HPLC (gradient elution of 75-100% aqueous MeOH) to
yield (R)-MTPA derivative 10a (0.3 mg). The (S)-MTPA derivative
10b (0.5 mg) was prepared in the same way. 10a: 1H NMR (600
MHz, CD3OD) 7.62-7.20 (m, 20H; aromatic), 7.08 (d, J ) 8.5,
2H; H-4′, H-8′), 6.73 (d, J ) 8.5, 2H; H-3′, H-5′), 6.06 (dd, J )
-9.0, 4.8, 1H; H-2′), 4.62 (m, 1H; H-10′), 4.62 (m, 1H; H-11′a),
4.52 (m, 1H; H-11′b), 4.01 (dd, -10.3, 5.2, 1H; H-9′a), 3.97 (dd,
-10.3, 5.9, 1H; H-9′b), 3.74 (m, 1H; H-1′a), 3.54 (m, 1H; H-1′b),
1
and 1056.35926 (M + H)+. H NMR (600 MHz, CD3OD) 7.36-
7.27 (m, 5H), 7.29 (d, J ) 8.3, 2H), 6.93 (d, J ) 8.3, 2H), 5.08 (d,
J ) -12.9, 1H), 5.05 (d, J ) -12.9, 1H), 4.71 (dd, J ) 7.6, 4.9,
1H), 4.23 (m, 1H), 4.12 (m, 1H), 4.05 (dd, J ) 5.5, 1.8, 2H), 4.00
(m, 1H), 3.77 (m, 1H), 3.69 (d, J ) 5.5, 2H), 3.67 (m, 1H), 3.57
(m, 2H), 3.52-3.40 (m, 6H), 3.38-3.31 (m, 2H), 2.24 (q, J ) 7.6,
2H), 1.94-1.25 (m, 42H), 1.12 (t, J ) 7.6, 3H).
J. Org. Chem, Vol. 72, No. 4, 2007 1223

Kobayashi et al.
3.54 (s, 3H; OMe in MTPA), 3.50 (s, 3H; OMe in MTPA), 3.37
(s, 3H; OMe in MTPA), 3.30 (s, 3H; OMe in MTPA). 10b: 1H
NMR (600 MHz, CD3OD) 7.64-7.26 (m, 22H; aromatic in MTPA,
H-4′, H-8′), 6.87 (d, J ) 8.2, 2H; H-5′, H-7′), 6.13 (t, J ) 6.9, 1H;
H-2′), 4.62 (m, 1H; H-10′), 4.58 (m, 1H; H-11′a), 4.49 (dd, J )
-11.0, 6.2, 1H; H-11′b), 4.02 (m, 2H; H2-9′), 3.68 (d, 6.2, 2H;
H2-1′), 3.39 (s, 3H; OMe in MTPA), 3.35 (s, 3H; OMe in MTPA),
3.34 (s, 3H; OMe in MTPA), 3.20 (s, 3H; OMe in MTPA).
Synthesis of 7. To a solution of N-Boc-L-serine methyl ester
(550 mg, 2.5 mmol) and p-toluenesulfonic acid monohydrate (6.5
mg, 0.03 mmol) in dry toluene (6 mL) was added 2,2-dimethox-
ypropane (0.6 mL). The reaction mixture was refluxed for 30 min
with stirring and evaporated. The residue was subjected to silica
gel flash chromatography (n-hexane/EtOAc ) 4:1) to yield 3-(1,1-
dimethylethyl)-4-methyl-S-2,2-dimethyl-3,4-oxazolidinedicarboxy-
late (556 mg, 2.15 mmol, 85.9% yield). To a solution of 3-(1,1-
dimethylethyl)-4-methyl-S-2,2-dimethyl-3,4-oxazolidinedicarbox-
ylate (556 mg, 2.15 mmol) in THF/MeOH (95:5, 13.2 mL) was
added LiBH4 (93 mg, 4.26 mmol). The reaction mixture was stirred
at room temperature for 2 h and then evaporated. The residue was
purified by silica gel flash chromatography (n-hexane/EtOAc )
1:1) to obtain 7 (416 mg, 1.82 mmol, 84.5% yield). 3-(1,1-
Dimethylethyl)-4-methyl-S-2,2-dimethyl-3,4- oxazolidinedicar-
boxylate: 1H NMR (600 MHz, CDCl3) 4.46-3.96 (m, 3H; CH2-
O, CH-N), 3.71 (s, 3H; OMe), 1.66-1.34 (m, 15H; five methyls).
7: 1H NMR (600 MHz, CDCl3) 4.14-3.56 (m, 5H; CH2-O × 2,
CH-N), 1.60-1.41 (m, 15H; five methyls).
Preparation of 9. To a mixture of 7 (277 mg, 1.20 mmol),
phenol (113 mg, 1.20 mmol), and triphenylphosphine (314 mg, 1.20
mmol) in toluene (3 mL) was added diethyl azodicarboxylate (628
µL (40% solution in toluene), 1.21 mmol). The reaction mixture
was stirred for 18 h at 80 °C and evaporated. The residue was
subjected to silica gel flash chromatography (n-hexane/EtOAc )
4:1) to yield 8 (226 mg, 0.74 mmol, 61.4% yield). To a solution of
8 150 mg (0.49 mmol) in CH2Cl2 (2 mL) was added TFA (1 mL).
The reaction mixture was stirred at room temperature for 2 h and
then evaporated to yield 9 as a TFA salt (135 mg, 0.48 mmol, 98.0%
yield). 8: 1H NMR (600 MHz, CDCl3) 7.28-6.89 (m, 5H;
aromatic), 4.30-3.79 (m, 5H; CH2-O × 2, CH-N), 1.63-1.44 (m,
15H; five methyls). 9: 1H NMR (600 MHz, CD3OD) 7.29 (t, J )
7.7, 2H; aromatic), 7.01-6.95 (m, 3H; aromatic), 4.22 (dd, J )
-10.5, 3.9, 1H; CH2a-O), 4.16 (dd, J ) -10.5, 6.9, 1H; CH2b-O),
3.88 (dd, J ) -11.6, 4.6, 1H; CH2a-O), 3.81 (dd, J ) -11.6, 5.6,
1H; CH2b-O), 3.63 (m, 1H; CH-N).
(R)- and (S)-MTPA Derivatives of 9 (11a and 11b). To a
solution of 9 (8 mg) in pyridine (50 µL) was added (S)-MTPACl
(40 µL). The reaction mixture was stirred at room temperature for
30 min and then evaporated. The residue was partitioned between
EtOAc and H2O with 0.1 M Na2CO3.The organic layer was
subjected to silica gel flash chromatography with CHCl3 to yield
(R)-MTPA derivative 11a (10.9 mg). The (S)-MTPA derivative 11b
(11.0 mg) was prepared in the same way. 11a: 1H NMR (600 MHz,
CD3OD) 7.58-6.77 (m, 15H; aromatic), 4.06 (m, 1H; CH2a-OPh),
3.98 (m, 1H; CH2b-OPh), 4.61 (m, 1H; CH-N), 4.64 (m, 1H; CH2a-
OMTPA), 4.54 (m, 1H; CH2a-OMTPA), 3.53 (s, 3H; OMe in
MTPA), 3.49 (s, 3H; OMe, in MTPA). 11b: 1H NMR (600 MHz,
CD3OD) 7.58-6.81 (m, 15H; aromatic), 4.00 (m, 2H; CH2-OPh),
4.60 (m, 1H; CH-N), 4.57 (m, 1H; CH2a-OMTPA), 4.47 (m, 1H;
CH2a-OMTPA), 3.52 (s, 3H; OMe in MTPA), 3.48 (s, 3H; OMe,
in MTPA).
Preparation of (R)- and (S)-MTPA Derivatives of (R)-2-
Amino-1-phenylethanol (12a and 12b). To a solution of the
commercially available (R)-2-amino-1-phenylethanol (20 mg) in
pyridine (50 µL) was added (S)-MTPACl (60 µL). The reaction
mixture was stirred at room temperature for 30 min and then
evaporated. The residue was partitioned between EtOAc and H2O
with 0.1 M Na2CO3. The organic layer was subjected to silica gel
flash chromatography with CHCl3 to yield (R)-MTPA derivative
12a (14 mg). The (S)-MTPA derivative 12b (18 mg) was prepared
in the same way. 12a: 1H NMR (600 MHz, CD3OD) 7.46-7.14
(m, 15H; aromatic), 6.12 (m, 1H; CH-O), 3.74 (m, 1H; CH2a-N),
3.57 (m, 1H; CH2b-N), 3.38 (s, 3H; OMe in MTPA), 3.30 (s, 3H;
OMe, in MTPA). 12b: 1H NMR (600 MHz, CD3OD) 7.44-7.32
(m, 15H; aromatic), 6.17 (m, 1H; CH-O), 3.68(m, 2H; CH2-N),
3.40 (s, 3H; OMe in MTPA), 3.20 (s, 3H; OMe, in MTPA).
Preparation of (R)- and (S)-MTPA Derivatives of 1 (13a and
13b). To a solution of 1 (5 mg) in pyridine (50 µL) was added
(S)-MTPACl (60 µL). The reaction mixture was stirred at room
temperature for 30 min and then evaporated. The residue was
partitioned between EtOAc and H2O with 0.1 M Na2CO3.The
organic layer was purified by ODS HPLC (gradient elution of 90-
100% aqueous MeOH) to yield (R)-MTPA derivative 13a (6 mg).
The (S)-MTPA derivative 13b (5.3 mg) was prepared in the same
way. 13a: 1H NMR (600 MHz, CD3OD) 5.07 (m, 1H; H-2), 1.74
(m, 1H; H2-3a), 1.86 (m, 1H; H2-3b), 1.26 (m, 1H; H2-4a), 1.45
(m, 1H; H2-4b), 1.62 (m, 1H; H2-5a), 1.75 (m, 1H; H2-5b), 5.00
(m, 1H; H-6), 2.90 (dd, J ) 9.0, 4.2, 1H; H-7), 2.98 (m, 1H; H-8),
5.07 (m, 1H; H-16), 1.26 (m, 2H; H2-26), 1.46 (m, 2H; H2-27),
3.33 (m, 2H; H2-28), 3.41 (m, 1H; H2-29a), 3.39 (m, 1H; H2-29b),
4.39 (m, 1H; H-30), 4.50 (dd, J ) -14.4, 7.6, 1H; H2-31a), 4.39
(m, 1H; H2-31b), 3.67 (m, 1H; H2-1′a), 3.56 (dd, J ) -14.4, 6,2,
1H; H2-1′b), 6.01 (t, J ) 6.2, 1H; H-2′), 7.14 (d, J ) 8.9, 2H;
H-4′/8′), 6.67 (d, J ) 8.9, 2H; H-5′/7′), 4.11 (dd, J ) -9.6, 8.3,
1H; H2-9′a), 3.97 (dd, J ) -9.6, 5.5, 1H; H2-9′b), 4.76 (m, 1H;
H-10′), 4.59 (dd, J ) -11.0, 8.3, 1H; H2-11′a), 3.86 (m, 1H; H2-
11′b), 2.53 (m, 1H, H2-13′a), 2.35 (m, 1H; H2-13′b), 0.95 (t, J )
7.3, 3H; H3-14′). 13b: 1H NMR (600 MHz, CD3OD) 4.93 (m, 1H;
H-2), 1.51 (m, 1H; H2-3a), 1.65 (m, 1H; H2-3b), 1.15 (m, 1H; H2-
4a), 1.31 (m, 1H; H2-4b), 1.46 (m, 1H; H2-5a), 1.58 (m, 1H; H2-
5b), 4.92 (m, 1H; H-6), 2.96 (dd, J ) 9.3, 4.2, 1H; H-7), 3.02 (m,
1H; H-8), 5.08 (m, 1H; H-16), 1.31 (m, 2H; H2-26), 1.52 (m, 2H;
H2-27), 3.40 (m, 2H; H2-28), 3.47 (dd, J ) -10.3, 5.5, 1H; H2-
29a), 3.43 (dd, J ) -10.3, 6.2, 1H; H2-29b), 4.39 (m, 1H; H-30),
4.50 (dd, J ) -11.0, 4.1, 1H; H2-31a), 4.34 (dd, J ) -11.0, 6.9,
1H; H2-29a), 3.63 (dd, J ) -14.5, 6,2, 1H; H2-1′a), 3.56 (m, 1H;
H2-1′b), 6.04 (t, J ) 6.2, 1H; H-2′), 7.26 (d, J ) 8.9, 2H; H-4′/8′),
6.61 (d, J ) 8.9, 2H; H-5′/7′), 3.93 (dd, J ) -9.3, 6.6, 1H; H2-
9′a), 3.74 (br, 1H; H2-9′b), 4.70 (m, 1H; H-10′), 4.73 (dd, J )
-11.7, 5.5, 1H; H2-11′a), 4.54 (dd, J ) -11.7, 6.2, 1H; H2-11′b),
2.52 (m, 1H, H2-13′a), 2.25 (m, 1H; H2-13′b), 0.97 (t, J ) 7.2, 3H;
H3-14′).
Preparation of (R)- and (S)-MTPA Derivatives of 5 (14a and
14b). To a solution of 5 (2 mg) in pyridine (50 µL) was added
(S)-MTPACl (40 µL). The reaction mixture was stirred at room
temperature for 30 min and then evaporated. The residue was
partitioned between EtOAc and H2O with 0.1 M Na2CO3. The
organic layer was purified by ODS HPLC (gradient elution of 90-
100% aqueous MeOH) to yield (R)-MTPA derivative 14a (3.3 mg).
The (S)-MTPA derivative 14b (3.6 mg) was prepared in the same
way. 14a: 1H NMR (600 MHz, CD3OD) 5.08 (m, 1H; H-2), 1.79
(m, 1H; H2-3a), 1.55 (m, 1H; H2-3b), 1.55 (m, 1H; H2-4a), 1.42
(m, 1H; H2-4b), 1.53 (m, 1H; H2-5a), 1.45 (m, 1H; H2-5b), 3.95
(m, 1H; H-6), 3.62 (m, 1H; H-7), 4.02 (m, 1H; H-8), 5.08 (m, 1H;
H-16), 3.38 (m, 2H; H2-28), 3.41 (m, 2H; H2-29), 4.06 (m, 1H;
H-30), 4.40 (dd, J ) -11.2, 5.0, 1H; H2-31a), 4.34 (dd, J ) -11.2,
7.3, 1H; H2-31b), 3.66 (m, 1H; H2-1′a), 3.59 (m, 1H; H2-1′b), 6.00
(dd, J ) 7.3, 5.5, 1H; H-2′), 7.15 (d, J ) 8.7, 2H; H-4′/8′), 6.68
(d, J ) 8.7, 2H; H-5′/7′), 4.11 (dd, J ) -10.5, 9.0, 1H; H2-9′a),
3.97 (dd, J ) -10.5, 5.5, 1H; H2-9′b), 4.77 (m, 1H; H-10′), 4.60
(dd, J ) -11.0, 8.5, 1H; H2-11′a), 3.84 (br, 1H; H2-11′b), 2.54
(m, 1H, H2-13′a), 2.37 (m, 1H; H2-13′b), 0.96 (t, J ) 7.4, 3H; H3-
14′a), 5.06 (d, J ) -12.4, 1H; CH2 in Cbz), 5.03 (d, J ) -12.4,
1H; CH2 in Cbz), 1.39 (s, 3H; CH3 in acetonide), 1.34 (s, 3H; CH3
in acetonide). 14b: 1H NMR (600 MHz, CD3OD) 5.00 (dd, J )
8.7, 4.1, 1H; H-2), 1.66 (m, 1H; H2-3a), 1.41 (m, 1H; H2-3b), 1.35
(m, 1H; H2-4a), 1.24 (m, 1H; H2-4b), 1.47 (m, 1H; H2-5a), 1.37
(m, 1H; H2-5b), 3.87 (m, 1H; H-6), 3.58 (m, 1H; H-7), 3.98 (m,
1H; H-8), 5.08 (m, 1H; H-16), 3.38 (t, J ) 6.4, 2H; H2-28), 3.40
1224 J. Org. Chem., Vol. 72, No. 4, 2007

Structure Elucidation of Shishididemniols
(m, 2H; H2-29), 4.05 (m, 1H; H-30), 4.42 (dd, J ) -11.0, 5.0, 1H;
H2-31a), 4.34 (dd, J ) -11.0, 6.9, 1H; H2-31b), 3.68 (dd, J )
-14.0, 6.2, 1H; H2-1′a), 3.57 (m, 1H; H2-1′b), 6.05 (dd, J ) 7.3,
6.5, 1H; H-2′), 7.27 (d, J ) 8.7, 2H; H-4′/8′), 6.62 (d, J ) 8.7, 2H;
H-5′/7′), 3.93 (dd, J ) -11.6, 6.4, 1H; H2-9′a), 3.75 (br, 1H; H2-
9′b), 4.70 (m, 1H; H-10′), 4.74 (dd, J ) -11.5, 5.5, 1H; H2-11′a),
3.54 (dd, J ) -11.5, 5.6, 1H; H2-11′b), 2.53 (m, 1H, H2-13′a),
2.26 (m, 1H; H2-13′b), 0.98 (t, J ) 7.3, 3H; H3-14′a), 5.03 (s, 2H;
CH2 in Cbz), 1.38 (s, 3H; CH3 in acetonide), 1.33 (s, 3H; CH3 in
acetonide).
1.17 (m, 1H; H-10b), 1.61-0.78 (m, 32H; H2-11 to H2-15, H2-17-
H2-27), 1.19 (s, 9H, tert-butyl), 1.13 (s, 9H, tert-butyl).
Preparation of N-Cbz Derivative 18. To a solution of 2 (40
mg, 0.046 mmol) in CHCl3/MeOH (3:1, 0.4 mL) were added
triethylamine (66.4 µL) and Cbz-Cl (17 µL). After stirring at room
temperature for 16 h, the reaction mixture was evaporated,
suspended in H2O, and extracted with EtOAc. The organic layer
was purified by ODS HPLC (gradient elution of 70-100% aqueous
MeOH) to afford 18 (21.7 mg, 46.7%). 18: ESIMS m/z 1010.45044
1
and 1012.44065 (M + H)+. H NMR (600 MHz, CD3OD) 7.37-
Preparation of 16. A 10 mg portion of 4 was treated with NaIO4
(20 mg) in MeOH/H2O (4:1, 1 mL) at room temperature for 1 h,
followed by reduction with NaBH4 (23 mg). The reaction mixture
was evaporated, and the residue was purified by ODS HPLC
(gradient elution of 80-100% aqueous MeOH) to obtain 15 (5.5
mg). To a solution of 15 (4 mg) in pyridine (50 µL) was added
tert-butyldiphenylchlorosilane (TBDPSCl) (10 µL). After stirring
at room temperature overnight, the reaction mixture was evaporated,
suspended in H2O, and extracted with EtOAc. The organic layer
was evaporated, and the residue was separated by silica gel flash
chromatography (n-hexane/EtOAc ) 4:1) to yield 16 (4.6 mg).
7.26 (m, 5H), 7.29 (d, J ) 8.7, 2H), 6.94 (d, J ) 8.3, 2H), 5.08 (d,
J ) -12.3, 1H), 5.05 (d, J ) -12.3, 1H), 4.71 (dd, J ) 7.6, 4.8,
1H), 4.23 (m, 1H), 4.05 (dd, J ) 5.5, 2.3, 2H), 4.03-3.99 (m,
2H), 3.78 (m, 1H), 3.70 (d, J ) 5.5, 2H), 3.67 (m 1H), 3.58 (m,
2H), 3.52-3.39 (m, 7H), 3.35 (m, 1H), 2.24 (q, J ) 7.8, 2H), 1.89-
1.25 (m, 42H), 1.12 (t, J ) 7.8, 3H).
Preparation of Acetonide 19 from 18. To the mixture of 18 (5
mg) and PPTS (1.7 mg) in CH2Cl2 (50 µL) was added 2,2-
dimethoxypropane (10 µL). The reaction mixture was stirred at
room temperature for 1 h and was then evaporated. The residue
was partition between EtOAc and H2O. The organic layer was
purified by ODS HPLC (gradient elution of 80-100% aqueous
MeOH) to give 19 (5.2 mg): ESIMS m/z 1050.45589 and
1
15: ESIMS m/z 644.22893 and 646.23454 (M + H)+. H NMR
(600 MHz, CDCl3) 7.36-7.28 (m, 5H), 5.43 (m, 1H), 5.09 (s, 2H),
4.12 (m, 1H), 3.85-3.53 (m, 8H), 3.40 (t, J ) 6.7, 2H), 1.85-
1.20 (m, 36H). 16: ESIMS m/z 1120.39025 and 1122.40749 (M
+ H)+. 1H NMR (600 MHz, benzene-d6) 7.80-7.72 (m, 5H), 7.26-
7.20 (m, 16H), 5.10 (s, 2H), 5.03 (d, J ) 8.7, 1H), 4.20 (m, 1H),
3.97-3.85 (m, 3H), 3.83 (dd, J ) -10.5, 6.0, 1H), 3.74 (dd, J )
-9.4, 6.4, 1H), 3.57 (dd, J ) -8.6, 3.9, 1H), 3.46 (m, 1H), 3.37
(dd, J ) -8.6, 5.5, 1H), 3.20 (t, J ) 6.4, 2H), 1.86-1.17 (m, 36H),
1.20 (s, 9H), 1.14 (s, 9H).
1
1052.50406 (M + H)+. H NMR (600 MHz, CD3OD) 7.37-7.26
(m, 5H), 7.30 (d, J ) 8.7, 2H), 6.93 (d, J ) 8.7, 2H), 5.08 (d, J )
-12.3, 1H), 5.06 (d, J ) -12.3, 1H), 4.70 (dd, J ) 7.5, 4.8, 1H),
4.22 (m, 1H), 4.05 (dd, J ) 5.6, 2.7, 2H), 4.04-3.98 (m, 2H),
3.96 (m, 1H), 3.80-3.74 (m, 2H), 3.69 (d, J ) 5.4, 2H), 3.57 (m,
2H), 3.52-3.39 (m, 6H), 3.35 (m, 1H), 2.24 (q, J ) 7.8, 2H), 1.84-
1.25 (m, 42H), 1.38 (s, 3H), 1.37 (s, 3H), 1.12 (t, J ) 7.8, 3H).
Preparation of 3 from 18. To a solution of 18 (5 mg) in 500
µL of MeOH was added potassium carbonate (18 mg). The reaction
mixture was stirred at room temperature for 1.5 h, quenched with
acetic acid, and then evaporated. The residue was purified by ODS
HPLC (gradient elution of 50-60% aqueous MeCN) to yield 3
Preparation of (R)- and (S)-2NMA Derivatives (17a and 17b).
To a solution of 16 (1.5 mg) in CH2Cl2 (300 µL) were added (R)-
2-naphthylmethoxyacetic acid ((R)-2NMA), 1-ethyl-3-(3-dimethy-
laminopropyl)-carbodiimide hydrochloride (EDC) (5 mg), DMAP
(0.5 mg), and DMAP‚HCl (0.5 mg), and the mixture was stirred at
room temperature overnight. The reaction mixture was evaporated
and partitioned between EtOAc and H2O. The organic layer was
subjected to silica gel flash chromatography (n-hexane/EtOAc )
9:1) to obtain the (R)-2NMA derivative 17a (1.5 mg). The (S)-
2NMA derivative 17b (1.5 mg) was prepared in the same way.
17a: 1H NMR (600 MHz, C6D6) 8.67-7.00 (m, 32H, aromatic),
5.37 (s, 1H, -CH-O in 2NMA), 5.09 (s, 2H; -CH2- in Cbz), 5.08
(m, 1H; H-16), 5.01 (d, J ) 8.7, 1H; NH-30), 4.20 (m, 1H; H-30),
3.95 (m, 1H; H-8), 3.92 (m, 1H; H-7a), 3.89 (m, 1H; H-31a), 3.85
(dd, J ) -10.6, 5.4, 1H; H-7b), 3.73 (dd, J ) -9.6, 6.8, 1H;
H-31b), 3.57 (br, 1H; H-29a), 3.36 (br, 1H; H-29b), 3.28 (s, 3H;
OMe in 2NMA), 3.20 (m, 2H; H2-28), 1.79 (m, 1H; H-9a), 1.72
(m, 1H; H-9b), 1.46 (m, 1H; H-10a), 1.29 (m, 1H; H-10b), 1.55-
0.80 (m, 32H; H2-11 to H2-15, H2-17-H2-27), 1.19 (s, 9H; tert-
butyl), 1.13 (s, 9H; tert-butyl). 17b: 1H NMR (600 MHz, C6D6)
8.03-7.02 (m, 32H, aromatic), 5.12 (m, 1H; H-16), 5.09 (s, 2H;
-CH2- in Cbz), 5.01 (d, J ) 8.7, 1H; NH-30), 4.92 (s, 1H, -CH-O
in 2NMA), 4.20 (m, 1H; H-30), 3.92 (m, 1H; H-8), 3.90 (m, 1H;
H-7a), 3.89 (m, 1H; H-31a), 3.83 (dd, J ) -10.6, 5.4, 1H; H-7b),
3.73 (dd, J ) -9.6, 6.8, 1H; H-31b), 3.57 (br, 1H; H-29a), 3.36
(br, 1H; H-29b), 3.30 (s, 3H; OMe in 2NMA), 3.20 (m, 2H; H2-
28), 1.72 (m, 1H; H-9a), 1.65 (m, 1H; H-9b), 1.34 (m, 1H; H-10a),
1
(3.5 mg) whose H NMR spectrum and [R]D value (3 from 18:
[R]20 -17.2 (c 0.10 MeOH), 3 from 1: [R]20 -17.6 (c 0.10
D
D
MeOH)) were indistinguishable from those of 3 prepared from 1.
Acknowledgment. Thanks are due to the crew of R/V
Toyoshio-maru for assistance in collecting the tunicate. We
thank Prof. Takenori Kusumi, Tokushima University, for
supplying both (R)- and (S)-2NMA. We also thank Prof.
Hidenori Watanabe, University of Tokyo, for valuable discussion
and Kohtaro Okada and Yoshinari Miyata for technical support.
This work was partially supported by Grant-in-aids from MEXT
(Priority Area 16073207) and JSPS (Area B 16380142).
Supporting Information Available: FAB-MS/MS data of 1
1
and 2; H NMR, 13C NMR, COSY, TOCSY, HMBC, and HSQC
1
1
spectra of 1 and 2 in DMSO-d6; H NMR spectra of 3-19; H
NMR, COSY, and TOCSY spectra of 13a and 13b in CD3OD; 1H
NMR and COSY spectra of 14a and 14b in CD3OD; and NOESY
spectra of 5 and 19 in CD3OD. This material is available free of
charge via the Internet at http://pubs.acs.org.
JO062013M
J. Org. Chem, Vol. 72, No. 4, 2007 1225