Thiodiketopiperazines from Epicoccum nigrum
Journal of Natural Products, 2010, Vol. 73, No. 7 1247
measurements yielded 4101 independent reflections after equivalent data
were averaged and Lorentz and polarization corrections were applied.
The structure was solved by direct methods using the SHELXL-97
program, expanded using difference Fourier techniques, and refined
by the program SHELXL-97 and full-matrix least-squares calculation-
s.The final refinement gave R1 ) 0.0358 and wR2 ) 0.0808.
Preparation of (R)- and (S)-MTPA Esters from 4. Following the
procedure summarized above, compound 4 (2.5 mg, 5.2 µmol) was
acylated in an NMR tube with (S)-MTPA chloride (4.0 µL, 21.2 µmol)
in pyridine-d5. The selective acylation was achieved at 0 °C for 30
min, with compound 4 being transformed entirely into the (R)-MTPA
ester 4b: HPLC/MS m/z 913 [M + Na]+; 1H NMR (500 MHz, pyridine-
1
d5, assigned by its H-1H COSY spectrum) δH 2.490 (1H, br d, J )
Ent-epicoccin G (2): colorless needles (MeOH-H2O); [R]20
D
13.0 Hz, H-3/3′a), 2.311 (1H, br d, J ) 13.0 Hz, H-3/3′b), 2.700 (1H,
m, H-4/4′), 5.300 (1H, m, H-5/5′), 2.275 (1H, m, H-6/6′a), 1.544 (1H,
m, H-6/6′b), 2.242 (1H, m, H-7/7′a), 1.544 (1H, m, H-7/7′b), 3.970
(1H, m, H-8/8′), 3.970 (1H, m, H-9/9′). In the same fashion, compound
4 was treated with (R)-MTPA chloride in pyridine-d5 to give the
expected (S)-MTPA ester 4a: 1H NMR (500 MHz, pyridine-d5, assigned
by its 1H-1H COSY spectrum) δH 2.682 (1H, br d, J ) 13.0 Hz, H-3/
3′a), 2.645 (1H, br d, J ) 13.0 Hz, H-3/3′b), 2.853 (1H, m, H-4/4′),
5.226 (1H, m, H-5/5′), 2.212 (1H, m, H-6/6′a), 1.450 (1H, m, H-6/
6′b), 2.212 (1H, m, H-7/7′a), 1.527 (1H, m, H-7/7′b), 3.826 (1H, m,
H-8/8′), 4.049 (1H, m, H-9/9′); 1H NMR (500 MHz, pyridine-d5,
assigned by spin decoupling) of 4, as a reference spectrum for the
selective Mosher’s reaction, δH 3.011 (1H, br d, J ) 13.0 Hz, H-3/
3′a), 2.287 (1H, br d, J ) 13.0 Hz, H-3/3′b), 2.768 (1H, m, H-4/4′),
3.716 (1H, m, H-5/5′), 2.266 (1H, m, H-6/6′a), 1.645 (1H, m, H-6/
6′b), 2.093 (1H, m, H-7/7′a), 1.564 (1H, m, H-7/7′b), 3.807 (1H, m,
H-8/8′), 3.662 (1H, m, H-9/9′).
-141.5 (c 0.1, MeOH); UV (MeOH) λmax 205 nm; IR νmax 3465, 2919,
1
1705, 1640, 1419, 1071 cm-1; H NMR data, see Table 2; 13C NMR
data, see Table 1; ESIMS m/z 455 [M + H]+, 477 [M + Na]+, 453 [M
- H]-, 489 [M + Cl]-; HRESIMS m/z 477.11381 [M + Na]+ (calcd
for C20H26N2O6S2Na, 477.1130).
Preparation of (R)- and (S)-MTPA Esters from 2. After dissolving
in pyridine (0.5 mL) under a gentle argon stream, compound 2 (1.5
mg, 3.5 µmol) was placed in a dried, clean NMR tube, and (S)-MTPA
chloride (2.0 µL, 10.6 µmol) was added and immediately shaken until
uniformly mixed. The NMR tube was kept overnight at room
temperature. The residue was then purified by RP-HPLC using 80%
CH3CN in H2O to afford the corresponding (R)-MTPA ester 2b (1.1
1
1
mg): H NMR (500 MHz, DMSO-d6, assigned by its H-1H COSY
spectrum) δH 2.763 (1H, br d, J ) 13.0 Hz, H-3/3′a), 2.363 (1H, dd,
J ) 13.0, 8.0 Hz, H-3/3′b), 2.829 (1H, br dd, J ) 8.0, 8.0 Hz, H-4/4′),
2.453 (1H, m, H-6/6′a), 2.344 (1H, m, H-6/6′b), 2.321 (1H, m, H-7/
7′a), 2.260 (1H, m, H-7/7′b), 5.852 (1H, m, H-8/8′), 4.326 (1H, br d,
J ) 8.0 Hz, H-9/9′). In an identical fashion, compound 2, (R)-MTPA
chloride, and pyridine were combined in an NMR tube and allowed to
stand overnight. 1H NMR measurements showed the quantitative
conversion of compound 2 to the anticipated (S)-MTPA ester 2a (1.3
mg): 1H NMR (500 MHz, DMSO-d6, assigned by 1H-1H COSY
spectrum) δH 2.789 (1H, br d, J ) 13.5 Hz, H-3/3′a), 2.400 (1H, dd,
J ) 13.0, 8.0 Hz, H-3/3′b), 3.036 (1H, br dd, J ) 8.0, 8.0 Hz, H-4/4′),
2.328 (1H, m, H-6/6′a), 2.062 (1H, m, H-6/6′b), 2.271 (1H, m, H-7/
7′a), 2.168 (1H, m, H-7/7′b), 5.789 (1H, m, H-8/8′), 4.439 (1H, br d,
J ) 8.0 Hz, H-9/9′).
Epicoccin L (5): white powder; [R]20 -88.0 (c 0.1, MeOH); UV
D
(MeOH) λmax 205 nm; IR νmax 3348, 2926, 1700, 1640, 1406, 1110
cm-1; 1H NMR data, see Table 2; 13C NMR data, see Table 1; ESIMS
m/z 457 [M + H]+, 479 [M + Na]+, 455 [M - H]-, 491 [M + Cl]-;
HRESIMS m/z 479.1262 [M + Na]+ (calcd for C20H28N2O6S2Na,
479.1286).
Epicoccin M (6): white powder; [R]20D +339.4 (c 0.1, MeOH); UV
(MeOH) λmax 205 nm; IR νmax 3373, 2923, 1665, 1403, 1000 cm-1; 1H
NMR data, see Table 3; 13C NMR data, see Table 1; ESIMS m/z 471
[M + H]+, 493 [M + Na]+, 469 [M - H]-, 505 [M + Cl]-; HRESIMS
m/z 471.0717 [M + H]+ (calcd for C19H23N2O6S3, 471.0718).
Preparation of (R)- and (S)-MTPA Esters from 6. After dissolving
in pyridine (0.5 mL) under a gentle argon stream, compound 6 (2.5
mg, 5.2 µmol) was placed in a dried, clean NMR tube and (S)-MTPA
chloride (2.0 µL, 10.6 µmol) was added and immediately shaken until
uniformly mixed. The NMR tube was kept overnight at room
temperature. The residue was purified by RP-HPLC using 80% CH3CN
X-ray Crystallographic Analysis of 220. Upon crystallization from
MeOH-H2O (1:1) using the vapor diffusion method, colorless crystals
of 2 were obtained. A crystal (0.57 mm × 0.40 mm × 0.34 mm) was
separated from the sample and mounted on a glass fiber, and data were
collected using a MM007HF + CCD (Saturn724+) area detector with
a graphite monochromator and Cu KR radiation, λ ) 1.54178 Å at
173(2) K. Crystal data: C20H34N2O10S2, M ) 526.61, space group
monoclinic, P21; unit cell dimensions were determined to be a )
10.949(2) Å, b ) 8.7488(17) Å, and c ) 13.036(3) Å; V ) 1232.7(4)
Å3, Z ) 2, Dcalcd ) 1.419 mg/m3, F(000) ) 560. The 12 282
measurements yielded 4399 independent reflections after equivalent data
were averaged and Lorentz and polarization corrections were applied.
The structure was solved by direct methods using the SHELXL-97
program, expanded using difference Fourier techniques, and refined
by the program SHELXL-97 and full-matrix least-squares calculations.
The final refinement gave R1 ) 0.0372 and wR2 ) 0.0955.
1
in H2O to afford the corresponding (R)-MTPA ester 6b (0.5 mg): H
NMR (500 MHz, DMSO-d6, assigned by its 1H-1H COSY spectrum)
δH 2.815 (1H, br d, J ) 13.0 Hz, H-3a), 2.448 (1H, dd, J ) 13.0, 8.0
Hz, H-3b), 2.839 (1H, br dd, J ) 8.0, 8.0 Hz, H-4), 2.355 (1H, m,
H-6a), 2.322 (1H, m, H-6b), 2.297 (1H, m, H-7a), 2.252 (1H, m, H-7b),
5.901 (1H, s, H-8), 4.420 (1H, br d, J ) 8.0 Hz, H-9), 2.810 (1H, br
d, J ) 13.0 Hz, H-3′a), 2.417 (1H, dd, J ) 13.0, 8.5 Hz, H-3′b), 2.926
(1H, br dd, J ) 8.5, 8.5 Hz, H-4′), 2.734 (1H, m, H-6′a), 2.625 (1H,
m, H-6′b), 4.236 (1H, m, H-7′), 5.901 (1H, s, H-8′), 4.564 (1H, br d,
J ) 8.0 Hz, H-9′). Treatment of 6 in the same manner with (R)-MTPA
chloride in pyridine showed a quantitative conversion to the (S)-MTPA
ester 6a (0.7 mg): 1H NMR (500 MHz, DMSO-d6, assigned by its
1H-1H COSY spectrum) δH 2.863 (1H, br d, J ) 13.0 Hz, H-3a), 2.490
(1H, dd, J ) 13.0, 8.0 Hz, H-3b), 3.098 (1H, br dd, J ) 8.0, 8.0 Hz,
H-4), 2.349 (1H, m, H-6a), 2.063 (1H, m, H-6b), 2.267 (1H, m, H-7a),
2.135 (1H, m, H-7b), 5.855 (1H, s, H-8), 4.517 (1H, br d, J ) 8.0 Hz,
H-9), 2.847 (1H, br d, J ) 13.0 Hz, H-3′a), 2.449 (1H, dd, J ) 13.0,
8.5 Hz, H-3′b), 3.211 (1H, br dd, J ) 8.5, 8.5 Hz, H-4′), 2.525 (1H,
m, H-6′a), 2.380 (1H, m, H-6′b), 4.151 (1H, m, H-7′), 5.855 (1H, s,
H-8′), 4.646 (1H, br d, J ) 8.5 Hz, H-9′).
Epicoccin J (3): white powder; [R]20 -75.9 (c 0.1, MeOH); UV
D
(MeOH) λmax 205 nm; IR νmax 3412, 2924, 1700, 1666, 1397, 1073
cm-1; 1H NMR data, see Table 2; 13C NMR data, see Table 1; ESIMS
m/z 453 [M + H]+, 475 [M + Na]+, 487 [M + Cl]-; HRESIMS m/z
453.1149 [M + H]+ (calcd for C20H25N2O6S2, 453.1154).
Epicoccin K (4): white powder; [R]20 -87.9 (c 0.1, MeOH); UV
D
(MeOH) λmax 205 nm; IR νmax 3353, 2929, 1637, 1411, 1102 cm-1; 1H
NMR data, see Table 2; 13C NMR data, see Table 1; ESIMS m/z 459
[M + H]+, 481 [M + Na]+, 493 [M + Cl]-; HRESIMS m/z 459.1671
[M + H]+ (calcd for C20H31N2O6S2, 459.1623).
Epicoccin N (7): white powder; [R]20D +229.2 (c 0.1, MeOH); UV
(MeOH) λmax 205 nm; IR νmax 3408, 2923, 1667, 1408, 1073 cm-1; 1H
NMR data, see Table 3; 13C NMR data, see Table 1; ESIMS m/z 487
[M + H]+, 509 [M + Na]+, 485 [M - H]-, 521 [M + Cl]-; HRESIMS
m/z 487.0671 [M + H]+ (calcd for C19H23N2O7S3, 487.0667).
Epicoccin O (8): white powder; [R]20D +106.3 (c 0.1, MeOH); UV
(MeOH) λmax 205 nm; IR νmax 3415, 2929, 1725, 1663, 1412, 1053
cm-1; 1H NMR data, see Table 3; 13C NMR data, see Table 1; ESIMS
m/z 461 [M + Na]+, 437 [M - H]-, 473 [M + Cl]-; HRESIMS m/z
439.1005 [M + H]+ (calcd for C19H23N2O6S3, 439.0998).
Procedure for the in-NMR-Tube Selective Mosher’s Reaction
to Produce Mono-MTPA Esters.6 The sample is dissolved in
1
pyridine-d5 in an NMR tube dried under a gentle argon stream. A H
NMR spectrum is recorded as a reference. The NMR tube is precooled
to 0 °C, and a calculated amount of MTPA chloride is added. The
NMR tube is rigorously shaken until the liquids are evenly mixed. The
1H NMR spectrum is recorded every 15 min. When evidence of
acylation is observed, the NMR tube is maintained at that temperature
until complete conversion has been observed. The acylation reaction
is quenched by adding one drop of D2O (to hydrolyze excess MTPA
chloride). The 1H NMR and 1H-1H COSY spectra are acquired to
Preparation of (R)- and (S)-MTPA Esters from 8. After dissolving
in pyridine (0.5 mL) under a gentle argon stream, compound 8 (1.0
mg, 2.3 µmol) was placed in a dried, clean NMR tube and (S)-MTPA
chloride (1.0 µL, 5.3 µmol) was added and immediately shaken until
1
derive H NMR data of the MTPA esters, resolving the overlapping
MTPA product signals when necessary.