Journal of Natural Products
NOTE
z 395.1075 [MþH]þ (calcd for C17H2479BrN4O2, 395.1077), 397.1054
(calcd for C17H2481BrN4O2, 397.1057).
7.36 (1H, dd, J = 8.3, 1.6 Hz, H-5), 3.45 (2H, m, H2-11), 3.05 (6H, m,
H2-13, H2-15, H2-18), 1.98 (2H, m, H2-12), 1.70 (2H, m, H2-16), 1.55
(2H, m, H2-17), 1.41 (9H, s, OBut); 13C NMR (CD3OD, 100 MHz) δ
182.1 (C-8), 166.0 (C-9), 158.7 (C-10), 140.1 (C-2), 138.8 (C-7a),
127.0 (C-5), 126.9 (C-3a), 124.4 (C-4), 118.1 (C-6), 116.2 (C-7), 114.0
(C-3), 80.1 (C-20), 48.8 (C-15), 46.5 (C-13), 40.5 (C-18), 36.9 (C-11),
28.7 (OBut), 28.1 (C-17), 27.5 (C-12), 24.6 (C-16); (þ)-ESIMS m/z
495 [M þ H]þ (50%), 497 [M þ H]þ (50%); (þ)-HRESIMS m/z
495.1597 [M þ H]þ (calcd for C22H3279BrN4O4, 495.1601), 497.1579
(calcd for C22H3281BrN4O4, 497.1582).
Didemnidine B (2). To a solution of 6 (9.8 mg, 16.1 μmol) in
CH2Cl2 (20 mL) was added TFA (10 mL). The mixture was stirred at
room temperature for 70 min, solvent was removed under reduced
pressure, and the crude product was purified by Sephadex LH20 column
chromatography (MeOH þ 0.05% TFA) to yield didemnidine B (2) as
the bistrifluoroacetate salt (7.0 mg, 70%, yellow oil): IR (neat) νmax
3254, 2954, 1672, 1605, 1504, 1125 cm-1; UV, 1H and 13C NMR data
were identical to those observed for the isolated natural product; (þ)-
HRESIMS m/z 395.1069 [M þ H]þ (calcd for C17H2479BrN4O2,
395.1077), 397.1049 (calcd for C17H2481BrN4O2, 397.1057).
N1-(Indolyl-3-glyoxamido)-N8-tert-butoxycarbonylspermidine (4).
N8-tert-Butoxycarbonyl spermidine9,10 (18.1 mg, 73.4 μmol), 2-(indol-
3-yl)glyoxylic acid (3)8 (11.6 mg, 61.2 μmol), and PyBOP (35.0 mg,
67.1 μmol) were dissolved in dry DMF (10 mL), triethylamine (34.1
μL) was added, and the reaction was left to stir at room temperature for
24 h. The crude reaction mixture was subjected to combinations of C8,
C18, and Sephadex LH-20 (MeOH, H2O-TFA (0.05%)) column
chromatography to afford N1-(indolyl-3-glyoxamido)-N8-tert-butoxy-
carbonylspermidine trifluoroacetate salt (4) (15.3 mg, 47%) as a pale
yellow oil: IR (neat) νmax 3389, 1677, 1437, 1204, 1136 cm-1; 1H NMR
(CD3OD, 400 MHz) δ 8.79 (1H, s, H-2), 8.29 (1H, m, H-4), 7.47 (1H,
m, H-7), 7.26 (2H, m, H-5 and H-6), 3.46 (2H, m, H2-11), 3.05 (6H, m,
H2-13, H2-15, H2-18), 1.98 (2H, m, H2-12), 1.71 (2H, m, H2-16), 1.56
(2H, m, H2-17), 1.41 (9H, s, OBut); 13C NMR (CD3OD, 100 MHz) δ
181.5 (C-8), 166.5 (C-9), 158.7 (C-10), 139.6 (C-2), 138.0 (C-7a),
128.0 (C-3a), 125.0 (C-6), 124.0 (C-5), 123.0 (C-4), 114.0 (C-3), 113.2
(C-7), 80.2 (C-20), 48.4 (C-15, obscured by solvent), 46.6 (C-13), 40.5
(C-18), 36.9 (C-11), 28.8 (OBut), 28.1 (C-17), 27.6 (C-12), 24.6 (C-
16); (þ)-ESIMS m/z 417 [M þ H]þ; (þ)-HRESIMS m/z 417.2509
[M þ H]þ (calcd for C22H33N4O4, 417.2496).
Biological Assays. Details of the neutrophil anti-inflammatory
assay,16 PLA2 and farnesyltransferase assays,17 and whole organism
parasite assay protocols18 have been reported elsewhere.
Didemnidine A (1). To a solution of 4 (10.1 mg, 19.1 μmol) in
CH2Cl2 (20 mL) was added TFA (10 mL). The mixture was stirred at
room temperature for 70 min, solvent was removed under reduced
pressure, and the crude product was purified by Sephadex LH20 column
chromatography (MeOH þ 0.05% TFA) to yield didemnidine A (1) as
the bistrifluoroacetate salt (7.2 mg, 70%, yellow oil): IR (neat) νmax
3351, 1672, 1633, 1495, 1436, 1200, 1141 cm-1; UV, 1H and 13C NMR
data were identical to those observed for the isolated natural product;
(þ)-ESIMS m/z 317 [M þ H]þ; (þ)-HRESIMS m/z 317.1973 [M þ
H]þ (calcd for C17H25N4O2, 317.1972).
’ ASSOCIATED CONTENT
S
Supporting Information. Taxonomic description and
b
color in situ photo of the Didemnum sp. ascidian, assigned 1H and
13C NMR data (DMSO-d6) for didemnimides A-D as their
trifluoroacetate salts, 1H, 13C, COSY, HSQC, and HMBC NMR
spectra (DMSO-d6) for didemnidines A (1) and B (2), 1H NMR
spectra of synthetic intermediates 4 and 6, and 1H and 13C NMR
spectra for synthetic 1 and 2. This material is available free of
2-(6-Bromoindol-3-yl)glyoxylic acid (5). 6-Bromoindole12 (0.535 g,
2.72 mmol) in a 100 mL two-necked round-bottomed flask was
dissolved in anhydrous ether (30 mL). A solution of oxalyl chloride
(2.72 mmol) in anhydrous ether (10 mL) was added dropwise to the
rapidly stirring indole solution at 0 ꢀC under a nitrogen atmosphere over
a period of 10 min. The mixture was stirred for an additional 10 min, and
an orange precipitate formed. Saturated NaHCO3 (10 mL) was added
dropwise to the solution and the mixture then heated to reflux for 30
min. Upon cooling, the reaction mixture was extracted with CH2Cl2 (3
ꢀ 10 mL) and then acidified with 10% HCl(aq) (10 mL) to yield 5 as a
yellow precipitate, which was then filtered and dried (0.650 g, 89%): mp
233-234 ꢀC (dec); IR (neat) νmax 3203, 1712, 1634, 1512, 1410,
1269 cm-1; 1H NMR (DMSO-d6, 400 MHz) δ 12.63 (1H, s, NH), 8.38
(1H, d, J = 3.1 Hz, H-2), 8.05 (1H, d, J = 8.7 Hz, H-4), 7.76 (1H, d, J = 1.4
Hz, H-7), 7.37 (1H, dd, J = 8.7, 1.4 Hz, H-5); 13C NMR (DMSO-d6, 100
MHz) δ 181.1 (C-8), 165.2 (C-9), 138.9 (C-2), 137.8 (C-7a), 125.9 (C-
5), 124.8 (C-3a), 123.0 (C-4), 116.5 (C-6), 115.8 (C-7), 112.5 (C-3);
(-)-ESIMS m/z 266 [M - H]- (50%), 268 [M - H]- (50%);
(-)-HRESIMS m/z 265.9460 [M - H]- (calcd for C10H579BrNO3,
265.9458), 267.9443 (calcd for C10H581BrNO3, 267.9438).
’ AUTHOR INFORMATION
Corresponding Author
*Tel: þ64 9 373 7599, ext 88284. Fax: þ64 9 373 7422. E-mail:
’ ACKNOWLEDGMENT
We acknowledge funding from the New Zealand Foundation
for Research Science and Technology (contract CO1X0205) and
the University of Auckland. We thank A. Longeon (MNHN, FRE
3206 CNRS, Paris, France) for PLA2 assay results, J. Dubois
(ICSN, CNRS Gif sur Yvette, France) for farnesyltransferase
assay results, the DTP branch of the National Cancer Institute for
cytotoxicity assays, M. Cal, S. Sax, and C. Stalder (Swiss TPH) for
parasite assay results, Dr. M. Schmitz for assistance with NMR
data acquisition, and Ms. R. Imatdieva for MS data. R.F. thanks
the University of Auckland for a doctoral scholarship.
N1-(6-Bromoindolyl-3-glyoxamido)-N8-tert-butoxycarbonylsper-
midine (6). N8-tert-Butoxy carbonylspermidine9,10 (22.0 mg, 89.7
μmol), 2-(6-bromoindol-3-yl)glyoxylic acid (5) (20.1 mg, 75.3 μmol),
and PyBOP (40.0 mg, 76.8 μmol) were dissolved in dry DMF (10 mL),
triethylamine (40 μL) was added, and the reaction was left to stir at room
temperature for 24 h. The crude reaction mixture was subjected to
combinations of C8 flash, C18 flash, and Sephadex LH-20 (MeOH,
H2O-TFA (0.05%)) column chromatography to afford N1-(6-bro-
moindolyl-3-glyoxamido)-N8-tert-butoxycarbonylspermidine trifluoro-
acetate (6) (18.2 mg, 40%) as a pale yellow oil: IR (neat) νmax 3388,
1675, 1460, 1205, 1131 cm-1; 1H NMR (CD3OD, 400 MHz) δ 8.79
(1H, s, H-2), 8.20 (1H, d, J = 8.3 Hz, H-4), 7.66 (1H, d, J = 1.6 Hz, H-7),
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