Semisynthesis and antitumor activities of new styryl-lactone derivatives

Article abstract of DOI:10.1248/cpb.53.1387

Nineteen new derivatives (2-20) of the naturally occurring compound, goniothalamin (1), were prepared by chemical modification and semi-synthetic methods. The antitumor activities of these derivatives and goniothalamin were evaluated in vitro against huma

Full text of DOI:10.1248/cpb.53.1387

November 2005
Chem. Pharm. Bull. 53(11) 1387—1391 (2005)
1387
Semisynthesis and Antitumor Activities of New Styryl-Lactone Derivatives
a
b
Fu Sheng ZHOU, Wei Dong TANG, Qing MU, ,a Guo Xun YANG,a Yang WANG,a Gao Lin LIANG,a and
Li Guang LOU
*
b
a School of Pharmacy, Fudan University; Shanghai 200032, China: and b Shanghai Institute of Materia Medica, Shanghai
Institutes for Biological Science, Chinese Academy of Sciences; Shanghai 200032, China.
Received February 10, 2005; accepted June 12, 2005
Nineteen new derivatives (2—20) of the naturally occurring compound, goniothalamin (1), were prepared by
chemical modification and semi-synthetic methods. The antitumor activities of these derivatives and goniothala-
min were evaluated in vitro against human tumor cell lines, and most of them showed an inhibitory effect against
HL-60 cancer cells. The derivatives 10-nitro-goniothalamin (2) and 10-amino-goniothalamin (4) gave selective in-
hibition concentration (IC50) of 1.10 and 1.14 mg/ml, respectively, against human stomach cancer SGC-7901 cells,
while that of etoposide (vp-16) as the positive control was 6.07 mg/ml. Finally, the partition coefficients, logP (p
values), of these derivative molecules, were evaluated by calculating the additive approximate organic fragment
logP value.
Key words styryl-lactone; semisynthesis; goniothalamin; antitumor activity
The natural styryl-lactones from Goniothalamus species suspension of the amine 4 and succinic anhydride in dry
(Annonaceae) possess potent antitumor activities.1—4) Gonio- CH2Cl2 was stirred at rt for 6 h to give 10-(hydroxysuccinyl)-
thalamin (1), which has been found to have good activity amino-goniothalamin (6) in 60% yield.15) The amino acid de-
against various tumor cell lines,5,6) was first isolated as a rivatives 14—20 were obtained as trifluoroacetate salts in
plant styryl-lactone in 1967 and later obtained from Gonio- good yield by reacting 4 with various N-tert-butoxycarbonyl
thalamus griffithii and other Annonaceae species.7—9) How- (t-Boc)-protected amino acids in the presence of dicyclo-
ever, semisynthesis study of goniothalamin derivatives has hexylcarbodiimide (DCC), and successive deprotection by
been scant so far, and none of them advanced to activity ex- trifluoroacetic acid (TFA) in dichloromethane.16) All com-
periments.3,10) Goniothalamin is a good starting material, be- pounds were unambiguously confirmed by spectroscopic
cause it is abundant in several Goniothalamus species and methods.
possesses lower oxidation activity compared with other
The antitumor activities of these derivatives were mea-
styryl-lactones.7) Some derivatives of 10- and 12-substituted sured in vitro using a SRB (Sulforhodamine B) assay against
goniothalamin were synthesized. Furthermore, in order to human promyelocytic leukemia HL-60 and human hepatoma
improve the water solubility of the compounds, a series of BEL-7402 cells. Compounds 2—5, which possessed signifi-
derivatives with a variety of different a-amino acid groups cant activity against HL-60 cells, were selected for further
were also prepared with moderate yield, and their partition evaluation against human lung carcinoma A549 and human
coefficients, logP or p values were calculated by the approxi- stomach cancer SGC-7901 cells with the reference com-
mate additive organic fragmental method.
The present paper concerns the semi-synthesis and antitu-
mor activities of nineteen new styryl-lactone derivatives in
the goniothalamin series (2—20).
Results and Discussion
The general methods employed for the preparation of
styryl-lactone derivatives 2—20 are outlined in Chart 1. The
nitration of 1 in acetic anhydride with a mixture of concen-
trated nitric acid and glacial acetic acid afforded two nitro-
goniothalamin derivatives, 10-nitro-goniothalamin (2) and
12-nitro-goniothalamin (3) in 52% and 11% yields, respec-
tively.11) Subsequent reduction of the nitro group in 2 was at-
tempted under different sets of conditions.12,13) Firstly, when
Fe/HCl was used as the reductant, the reactant disappeared
but no target product was obtained. Then, by sodium hydro-
sulfite in methanol/water (90%) at room temperature (rt), 2
was reduced to 10-amino-goniothalamin (4) in a low yield of
5%. In the condition using 0.5 M stannous chloride dihydrate
(14 eq) in N,N-dimethylformamide (DMF)/dichloromethane
(1 : 1), 2 was successfully reduced to 4 in moderate yield
(61%). Acetylization of 4 with Ac2O at rt gave 10-acetyl-
Chart 1. The Scheme of the Semisynthesis of Goniothalamin Derivatives
amino-goniothalamin (5) in satisfactory yield (92%).14)
A
∗ To whom correspondence should be addressed. e-mail: qingmu@shmu.edu.cn
© 2005 Pharmaceutical Society of Japan

1388
Vol. 53, No. 11
pound, goniothalamin. For two reasons, VP-16 (etoposide, could not enhance the antitumor activity in vitro.
21) was chosen as a positive control standard. First, the
The approximate calculating logP (p value), or log of the
etoposide is also the derivative of a naturally originated com- partition coefficient, for a molecule was used for evaluating
pound. Secondly, it had consecutively been used as the con- the water solubility of the derivatives. P is logarithmically re-
trol standard in our previous antitumor tests of the natural lated to free energy, generally expressed as logP, and is there-
styryllactones isolated from Goniothalamus plants.9) The re- fore the sum of the hydrophobic and hydrophilic characteris-
sults, expressed as IC50 values, are summarized in Table 2.
tics of the organic functional groups making up the structure
In the antitumor tests, the semi-synthetic styryllactone de- of the molecule.17) According to the p value for organic frag-
rivatives 2, 3 and 4, as well as goniothalamin (1) showed ments,18—20) each derivative compound in this experiment
stronger inhibitory effects (IC50 1.07—1.56 mg/ml) than the was calculated and included in Table 2. The higher logP
positive control drug etoposide (IC50 6.07 mg/ml) against the value corresponds to the stronger hydrophobic or weaker hy-
tumor cell strain SGC-7901 (human stomach tumor), even drophilic nature of the molecule.
though most of the derivatives generally gave lower inhibi-
tion to HL-60, Bel-7402 tumor cell lines (Table 2).
After the chemical modification, most of the amino acid
derivatives of gonithalamin which possessed higher logP val-
The compounds 2, 3 and 4, however, showed comparable ues (ꢀ4.8 to ꢀ7.3), expressed poor antitumor activities
or superior activity against HL-60, A549 or SGC-7901 cells against HL-60 cells (IC50 was 2—3 mg/ml); meanwhile the
in comparison to their parent 1 (Table 2). Compound 2 of the compounds 1, 2 and 3, whose logP values were ꢀ4.4 to
main product in nitration was chosen as the starting material ꢀ4.7, possessed good activities (IC50 was 0.18—0.68 mg/ml).
for the next step reaction, and subsequent reduction easily In addition, different amino moieties in the styryllactone de-
gave the amine 4. Its antitumor activity was similar to 3. It rivatives resulted in little difference in antitumor activity.
was noticeable that the formation of the acylamino group of
Experimental
the 10-position made the IC50 of the derivatives increase dra-
matically, from 0.64 to 3.30 mg/ml for A549 cells, and from
1.14 to 25 mg/ml for SGC-7901 cells (Table 2). It was as-
General 1H- and 13C-NMR spectra (d ppm, J in Hz) were run at 400
and 100 MHz on a Mercury Plus 400 NMR spectrometer. EI/MS were
recorded on an Agilent 5973 spectrometer. ESI/MS and HR- (high resolu-
tion) ESI/MS were performed on a Q-Tof microspectrometer. MALDI/MS
(matrix-assisted laser desorption-ionization) and HR-MALDI/MS were
taken on an IonSpec 4.7 Tesla FT (Fourier transform) spectrometer. Silica
gel TLC plates were observed under UV light (254 nm) and after being
sprayed with 20% (v/v) H2SO4 in ethanol.
sumed that the carbonyl group in the acylamide distributed
the lone electrons in the nitrogen because of the conjugated
effect, and this indicated that the free amino group at the 10-
position was favorable for its antitumor activity. Further
study could explore in detail how the substituents at C-10 af-
fect the antitumor activity after the conversion of an amino
group in the amine 4 to other substituted groups. Addition-
ally, several amino acid trifluoroacetate derivatives (15, 17,
18, 20) showed comparable activity against HL-60 cells to
the amine 4, but were less active than parent 1. The data ob-
tained indicated that the introduction of amino acid groups
Extraction and Isolation Dried and powdered leaves (10 kg) of G. grif-
fithii were extracted with alcohol and concentrated. The extract was dis-
solved in MeOH–H2O (1 : 9) and was extracted with petroleum. After being
concentrated, 120 g of brown resin was subjected to silica gel chromatogra-
phy, eluted with a gradient of petroleum/ethyl acetate, which gave goniothal-
amin (1) 8.0 g (colorless crystal, Rfꢁ0.22 petroleum/ethyl acetate (v/v
1
8 : 2)), structurally confirmed by comparing its H- and 13C-NMR data with
those reported.6)
Bioassays Live cancer cells were established by using Trypan blue stain,
and cell generation was measured by means of the SRB method. All test
compounds were solubilized in DMSO. Each compound concentration was
tested in quadruplicate wells. The cancer cells, cultured in 10% bovine
serum, were digested with trypsin solution. The cancer cells at a concentra-
tion of 1ꢂ105 cells/ml were inoculated into every cell of a 96-well dish and
cultured at 37 °C in 5% CO2 for 48 h. The cellular proteins were precipitated
to the plates with trichloroacetic acid and stained with 0.4% SRB. Protein
bound SRB was solubilized with Tris base and read at 515 nm in an ELISA
reader. The protein content of the compound-treated cells was compared to
that of the DMSO control, and IC50 values were obtained.
Preparation of Compounds 2 and 3 To an acetic anhydride (45 ml) so-
lution of 1 (3050 mg, 15 mmol) was added dropwise at 0 °C concentrated ni-
tric acid and glacial acetic acid (18 ml 1 : 3) for 1.5 h. After the addition was
complete, the mixture was stirred at rt for 2 h, then ice and CH2Cl2 was
added, and the reaction products were extracted into CH2Cl2. The residue
Table 1. Styryl-lactone Derivatives 7—20 and Their Corresponding
Yields
Compd.
R
% Yield Compd.
Rꢄ
% Yield
7
8
9
10
11
12
13
NHBoc-L-Gly
NHBoc-L-Ala
NHBoc-L-Val
NHBoc-L-Leu
NHBoc-L-Ile
NHBoc-L-Gln
NHBoc-L-Phe
77
81
90
68
92
80
78
14
15
16
17
18
19
20
L-Glya)
L-Alaa)
L-Vala)
L-Leua)
L-Ilea)
59
69
85
85
91
65
59
L-Glna)
L-Phea)
a) Obtained as trifluoroacetate salt.
Table 2. Antitumor Activities of Styryl-lactone Derivativesa)
Compd.
HL-60b)
BEL-7402c)
A549d)
SGC-7901e)
LogPg)
Compd.
HL-60b)
BEL-7402c)
31.34
LogPg)
1
2
3
4
5
0.58
0.68
0.18
2.27
2.45
3.11
0.13
4.13
8.17
0.35
0.33
1.90
0.64
3.30
1.07
1.10
1.56
1.14
25.00
ꢀ4.7
ꢀ4.4
ꢀ4.4
ꢀ3.3
ꢀ3.4
ꢀ4.8
8
9
3.92
3.81
2.51
2.54
3.21
3.21
2.28
ꢀ5.3
ꢀ6.0
ꢀ7.3
ꢀ4.0
ꢀ5.5
ꢀ5.5
ꢀ6.3
26.88
24.08
32.62
29.90
15.28
13
15
17
18
20
26.27
31.00
7
21f)
0.19
6.07
13.96
a) Inhibition of HL-60, BEL-7402, A549, and SGC-7901 proliferation as measured by SRB assay, IC50, mg/ml. b) IC50 of compounds 6, 10, 11, 14, 16 and 19 with IC50
ꢃ5 mg/ml. c) IC50 of compounds 6, 9—12, 14—16, and 18—19 with IC50 ꢃ40 mg/ml. d) Compounds 6—20 not tested. e) Compounds 6—20 not tested. f) Positive con-
trol standard (VP-16). g) The approximate fragment p values for the calculations is listed in Experimental.

November 2005
1389
was subjected to column chromatography on silica gel (petroleum : ethyl ac-
etateꢁ37 : 19) to afford 1901 mg of 2 (52%) and 400 mg of 3 (11%).
10-Nitro-goniothalamin (2): Yellow solid, Rfꢁ0.24 petroleum/ethyl ac-
6.80 (1H, d, Jꢁ15.65 Hz, H-8), 6.15 (1H, dd, Jꢁ15.65, 6.24 Hz, H-7), 6.06
(1H, d, Jꢁ10.18 Hz, H-3), 5.64 (1H, br, NHBoc), 5.10—5.05 (1H, m, H-6),
3.95 (2H, d, Jꢁ4.30 Hz, Gly CH2), 2.56—2.54 (2H, m, H-5), 1.43 (9H, s,
Boc CH3ꢂ3). MALDI/MS m/z 395 (MꢀNa)ꢀ HR-MALDI/MS Calcd
C20H24N2O5 (MꢀNa)ꢀ: 395.1577, Found 395.1585.
1
etate (v/v 66 : 34) H-NMR (acetone-d6) d: 7.98 (1H, dd, Jꢁ8.05, 1.22 Hz,
Ar-H), 7.81 (1H, dd, Jꢁ8.04, 1.22 Hz, Ar-H), 7.70 (1H, ddd, Jꢁ7.80, 7.30,
1.22 Hz, Ar-H), 7.55 (1H, ddd, Jꢁ7.80, 7.60, 1.22 Hz, Ar-H), 7.11 (1H, d,
Jꢁ15.85 Hz, H-8), 7.07—7.03 (1H, m, H-4), 6.47 (1H, dd, Jꢁ15.85,
6.09 Hz, H-7), 5.99—5.96 (1H, m, H-3), 5.24—5.19 (1H, m, H-6), 2.73—
2.65 (1H, m, H-5), 2.60—2.51 (1H, m, H-5). EI/MS m/z 246 (MꢀH)ꢀ. HR-
MALDI/MS Calcd C13H12NO4 (MꢀH)ꢀ: 246.0761, Found 246.0770.
12-Nitro-goniothalamin (3): Yellow powder, Rfꢁ0.23 petroleum/ethyl ac-
etate (v/v 65 : 35) 1H-NMR (acetone-d6) d: 8.22 (2H, dt, Jꢁ8.84, 2.46,
1.96 Hz, Ar-H), 7.78 (2H, dt, Jꢁ8.59, 2.46, 1.96 Hz, Ar-H), 7.08—7.04 (1H,
m, H-4), 6.91 (1H, d, Jꢁ15.97 Hz, H-8), 6.71 (1H, dd, Jꢁ15.97, 5.89 Hz, H-
7), 5.99 (1H, m, H-3), 5.24—5.19 (1H, m, H-6), 2.74—2.52 (2H, m, H-5).
EI/MS m/z 245 Mꢀ HR-MALDI/MS Calcd C13H12NO4 (MꢀH)ꢀ: 246.0761,
Found 246.0750.
10-(N-a-tert-Butoxycarbonyl-L-alanyl)amino-goniothalamin (8)
A
mixture of DCC (35 mg, 0.17 mmol), compound 4 (24 mg, 0.11 mmol), and
N-tert-butoxycarbonyl-L-alanine (32 mg, 0.17 mmol) in CH2Cl2 (2.0 ml) was
stirred for 1 h at rt. The solids were filtered and the filtrate was concentrated
and chromatographed (petroleum : ethyl acetateꢁ3 : 2) to give 8 as a yellow
amorphous solid (35 mg, 81%). Compound 8: Rfꢁ0.23 petroleum/ethyl ac-
etate (v/v 6 : 4) 1H-NMR (CDCl3) d: 8.42 (1H, br s, NHCO), 7.70 (1H, t, Jꢁ
7.82 Hz, Ar-H), 7.40 (1H, d, Jꢁ7.82 Hz, Ar-H), 7.25 (1H, t, Jꢁ7.43 Hz, Ar-
H), 7.13 (1H, t, Jꢁ7.83, 7.43 Hz, Ar-H), 6.94—6.87 (1H, m, H-4), 6.82 (1H,
d, Jꢁ16.04 Hz, H-8), 6.17 (1H, dd, Jꢁ15.65, 6.65 Hz, H-7), 6.07—6.04 (1H,
dm, H-3), 5.31—5.28 (1H, br, NHBoc), 5.11—5.04 (1H, m, H-6), 4.34 (1H,
d, Ala a-CH), 2.56—2.54 (2H, m, H-5), 1.45—1.41 (12H, m, Ala b-CH3
and Boc CH3ꢂ3). Compound 8: ESI/MS m/z 387 (MꢀH)ꢀ HR-ESI/MS
Calcd C21H26N2O5Na (MꢀNa)ꢀ: 409.1742, Found 409.1739.
10-Amino-goniothalamin (4) 2 (800 mg, 3.3 mmol) was treated with
tin(II) chloride dihydrate (10310 mg, 45.7 mmol) in DMF/CH2Cl2 (91 ml,
1 : 1) under N2 for 3 h at rt. The CH2Cl2 was removed in vacuo at rt. The re-
maining solution was adjusted to pH 8 with 10% Na2CO3 (aq.), and the mix-
ture was extracted with ethyl acetate. The organic layer was washed with
water and brine, then dried over anhydrous Na2SO4. The solvent was evapo-
rated under reduced pressure and the residue was subjected to column chro-
matography on silica gel (petroleum : ethyl acetateꢁ3 : 2) to afford 4 as a
yellow solid (425 mg, 61%). Compound 4: Rfꢁ0.25 petroleum/ethyl acetate
(v/v 58 : 42) 1H-NMR (CDCl3) d: 7.25 (1H, dd, Jꢁ7.42, 1.17 Hz, Ar-H),
7.10 (1H, ddd, Jꢁ7.80, 7.42, 1.17 Hz), 6.95—6.90 (1H, m, H-4), 6.80 (1H,
d, Jꢁ15.61 Hz, H-8), 6.76 (1H, t, Jꢁ7.42 Hz, Ar-H), 6.69 (1H, dd, Jꢁ7.80,
1.17 Hz, Ar-H), 6.16 (1H, dd, Jꢁ15.61, 6.25 Hz, H-7), 6.09 (1H, dt, Jꢁ9.75,
1.95, 1.57 Hz, H-3), 5.12—5.07 (1H, m, H-6), 3.81 (2H, br s, NH2), 2.56—
2.52 (2H, m, H-5). EI/MS m/z 215 Mꢀ HR-MALDI/MS Calcd C13H14NO2
(MꢀH)ꢀ: 216.1038, Found 216.1029.
10-(N-a-tert-Butoxycarbonyl-L-valyl)amino-goniothalamin (9) To a
solution of 4 (25 mg, 0.12 mmol), N-tert-butoxycarbonyl-L-valine (78 mg,
0.36 mmol) and CH2Cl2 (3.0 ml) was added DCC (74 mg, 0.36 mmol). The
solution was stirred for 12 h at rt, filtered, and concentrated. The residue was
chromatographed (petroleum : ethyl acetateꢁ32 : 18) to give 9 as a yellow
amorphous solid (43 mg, 90%). Compound 9: Rfꢁ0.26 petroleum/ethyl ac-
1
etate (v/v 6 : 4) H-NMR (CDCl3) d: 8.39 (1H, br, NHCO), 7.60 (1H, t, Jꢁ
9.39, 8.21 Hz, Ar-H), 7.41 (1H, d, Jꢁ7.83 Hz, Ar-H), 7.24 (1H, t, Jꢁ7.82,
7.43 Hz, Ar-H), 7.13 (1H, t, Jꢁ7.82, 7.43 Hz, Ar-H), 6.90—6.80 (1H, m, H-
4), 6.78 (1H, d, Jꢁ16.00 Hz, H-8), 6.15 (1H, dd, Jꢁ16.00, 6.66 Hz, H-7),
6.01 (1H, t, Jꢁ9.39, 8.61 Hz, H-3), 5.37 (1H, br, NHBoc), 5.03—4.93 (1H,
m, H-6), 4.16—4.10 (1H, m, Val a-CH), 2.49—2.39 (2H, m, H-5), 2.25—
2.19 (1H, m, Val b-CH), 1.39 (9H, d, Jꢁ5.86 Hz, Boc CH3ꢂ3), 1.04—0.98
(6H, m, Val b-CH3 and g-CH3). MALDI/MS m/z 437 (MꢀNa)ꢀ HR-
MALDI/MS Calcd C23H30N2O5Na (MꢀNa)ꢀ: 437.2047, Found 437.2053.
10-Acetylamino-goniothalamin (5) The mixture of 4 (20 mg, 0.09
mmol) and Ac2O (1.0 ml). was stirred for 40 min at rt. Water and CHCl3 was
added to the reaction mixture and the organic layer was washed with water.
The organic solvent was evaporated under reduced pressure. The residue
was purified by silical gel column chromatography (CHCl3 : MeOHꢁ
96 : 1.5) to give 5 (22 mg, 92%) as a yellow amorphous powder. Compound
10-(N-a-tert-Butoxycarbonyl-L-leucyl)amino-goniothalamin (10)
A
mixture of DCC (49 mg, 0.24 mmol), compound 4 (25 mg, 0.12 mmol), N-
tert-butoxycarbonyl-L-leucine (55 mg, 0.24 mmol), and CH2Cl2 (2.0 ml) was
stirred at rt for 12 h. The solids were filtered and the filtrate was concen-
trated and chromatographed (petroleum : ethyl acetateꢁ35 : 15) to give 10 as
a white powder (34 mg, 68%). Compound 10: Rfꢁ0.32 petroleum/ethyl ac-
1
5: Rfꢁ0.22 petroleum/ethyl acetate (v/v 17 : 83) H-NMR (CDCl3) d: 7.67
1
(1H, d, Jꢁ7.65 Hz, Ar-H), 7.42 (1H, br, NH), 7.39 (1H, d, Jꢁ7.75 Hz, Ar-
H), 7.27 (1H, t, Jꢁ7.75 Hz, Ar-H), 7.14 (1H, t, Jꢁ7.75 Hz, Ar-H), 6.94—
6.90 (1H, m, H-4), 6.83 (1H, d, Jꢁ15.92 Hz, H-8), 6.15 (1H, dd, Jꢁ15.91,
6.13 Hz, H-7), 6.07 (1H, d, Jꢁ9.79 Hz, H-3), 5.12—5.07 (1H, m, H-6),
2.55—2.52 (2H, m, H-5), 2.20 (3H, s, CH3). 13C-NMR (CDCl3) d: 168.3
(NHCO), 163.3 (C-2), 144.4 (C-4), 134.2 (C-9), 128.8 (C-10), 128.6 (s),
128.3 (sꢂ2), 126.4 (s), 125.4 (s), 124.5 (s), 121.2 (C-3), 77.9 (C-6), 30.1 (C-
5), 24.5 (CH3). EI/MS m/z 257 Mꢀ HR-MALDI/MS Calcd C15H15NO3Na
(MꢀNa)ꢀ: 280.0944, Found 280.0955.
etate (v/v 6 : 4) H-NMR (CDCl3) d: 8.40 (1H, br, NHCO), 7.69 (1H, d, Jꢁ
8.22 Hz, Ar-H), 7.40 (1H, d, Jꢁ7.44 Hz, Ar-H), 7.25 (1H, t, Jꢁ7.83 Hz, Ar-
H), 7.13 (1H, t, Jꢁ7.83, 7.43 Hz, Ar-H), 6.93—6.84 (1H, m, H-4), 6.82 (1H,
d, Jꢁ16.04 Hz, H-8), 6.27 (1H, dd, Jꢁ16.04, 6.26 Hz, H-7), 6.07—6.03 (1H,
m, H-3), 5.21 (1H, br, NHBoc), 5.09—5.00 (1H, m, H-6), 4.30 (1H, s, Leu
a-CH), 2.56—2.48 (2H, m, H-5), 1.78—1.74 (2H, m, Leu b-CH2), 1.61—
1.57 (1H, m, Leu g-CH), 1.41 (9H, d, Jꢁ5.09 Hz, Boc CH3ꢂ3), 0.98—0.94
(6H, m, Leu CH3ꢂ2). MALDI/MS m/z 451 (MꢀNa)ꢀ HR-MALDI/MS
Calcd C24H32N2O5Na (MꢀNa)ꢀ: 451.2203, Found 451.2208.
10-(Hydroxysuccinyl)amino-goniothalamin (6) A suspension of the
amine 4 (15 mg, 0.07 mmol) and succinic anhydride (120 mg, 1.20 mmol) in
dry CH2Cl2 (3 ml) was stirred for 6 h at rt. The solids were filtered and the
filtrate was concentrated and chromatographed (CHCl3 : MeOHꢁ46 : 4) to
give 6 as a yellow amorphous solid (13 mg, 60%). Compound 6: Rfꢁ0.25
CHCl3/MeOH (v/v 9 : 1) 1H-NMR (CDCl3) d: 8.21 (1H, br s, NH), 7.65 (1H,
d, Jꢁ7.80 Hz, Ar-H), 7.45 (1H, d, Jꢁ6.64 Hz, Ar-H), 7.28 (1H, t, Jꢁ
7.60 Hz, Ar-H), 7.14 (1H, t, Jꢁ7.81 Hz, Ar-H), 7.04—7.00 (1H, m, H-4),
6.95 (1H, d, Jꢁ16.00 Hz, H-8), 6.16—6.11 (2H, m, H-4, H-7), 5.21—5.16
(1H, m, H-6), 2.84—2.45 (6H, m, H-5, COCH2CH2COOH). 13C-NMR
(CDCl3) d: 174.9 (COOH), 170.5 (NHCO), 165.5 (C-2), 145.9 (C-4), 134.1
(C-9), 128.6 (C-10), 128.3 (s), 126.3 (s), 126.0 (s), 125.3 (s), 125.0
(s), 124.9 (s), 120.8 (C-3), 77.2 (C-6), 32.5 (s), 30.6 (s), 30.4 (s). EI/MS m/z
315 Mꢀ HR-MALDI/MS Calcd C17H18NO5 (MꢀH)ꢀ: 316.1180, Found
316.1190.
10-(N-a-tert-Butoxycarbonyl-L-glycyl)amino-goniothalamin (7) To a
stirred solution of 4 (30 mg, 0.14 mmol) and N-tert-butoxycarbonyl-L-
glycine (147 mg, 0.84 mmol) in CH2Cl2 (3.5 ml), DCC (173 mg, 0.84 mmol)
was added. The reaction mixture was stirred at rt for 1.5 h and the solids
were filtered. The filtrate was concentrated in vacuo and the residue was
chromatographed (petroleum : ethyl acetateꢁ8 : 5). Initially obtained was 7
as a white amorphous solid (40 mg, 77%). Compound 7: Rfꢁ0.24 petro-
leum/ethyl acetate (v/v 53 : 47) 1H-NMR (CDCl3) d: 8.37 (1H, br s, NHCO),
7.64 (1H, d, Jꢁ8.22 Hz, Ar-H), 7.39 (1H, d, Jꢁ7.44 Hz, Ar-H), 7.25 (1H, t,
Jꢁ7.43 Hz, Ar-H), 7.13 (1H, t, Jꢁ7.43 Hz, Ar-H), 6.94—6.89 (1H, m, H-4),
10-(N-a-tert-Butoxycarbonyl-L-isoleucyl)amino-goniothalamin
(11)
A mixture of DCC (62 mg, 0.30 mmol), compound 4 (17 mg, 0.08 mmol), N-
tert-butoxycarbonyl-L-isoleucine (70 mg, 0.30 mmol), and CH2Cl2 (2.0 ml)
was stirred at rt for 24 h. The solids were filtered and the filtrate was concen-
trated and chromatographed (petroleum : ethyl acetateꢁ35 : 15) to give 11 as
a white amorphous powder (31 mg, 92%). Compound 11: Rfꢁ0.29 petro-
leum/ethyl acetate (v/v 6 : 4) 1H-NMR (CDCl3) d: 8.26 (1H, br, NHCO),
7.64 (1H, t, Jꢁ8.21 Hz, Ar-H), 7.42 (1H, d, Jꢁ7.83 Hz, Ar-H), 7.26 (1H, t,
Jꢁ8.21 Hz, Ar-H), 7.15 (1H, t, Jꢁ7.83, 7.43 Hz, Ar-H), 6.92—6.82 (1H, m,
H-4), 6.80 (1H, d, Jꢁ15.65 Hz, H-8), 6.17 (1H, dd, Jꢁ16.04, 6.55 Hz, H-7),
5.28 (1H, br, NHBoc), 5.06—4.97 (1H, m, H-6), 4.18—4.08 (1H, m, Ile a-
CH), 2.53—2.43 (2H, m, H-5), 2.03—1.97 (1H, m, Ile b-CH), 1.59 ( m, 1H,
Ile g-CH2a), 1.41 (9H, d, Jꢁ5.87 Hz, Boc CH3ꢂ3), 1.26—1.16 (2H, m, Ile
g-CH2b), 1.01 (3H, dd, Jꢁ7.65, 2.34 Hz, Ile b-CH3), 0.92 (3H, t, Jꢁ7.44 Hz,
Ile g-CH3). MALDI/MS m/z 451 (MꢀNa)ꢀ HR-MALDI/MS Calcd
C24H32N2O5Na (MꢀNa)ꢀ: 451.2203, Found 451.2207.
10-(N-a-tert-Butoxycarbonyl-L-glutaminyl)amino-goniothalamin (12)
A mixture of DCC (62 mg, 0.30 mmol), compound 4 (21 mg, 0.10 mmol), N-
tert-butoxycarbonyl-L-glutamine (74 mg, 0.30 mmol), and CH2Cl2 (2.0 ml)
was stirred at rt for 2 h. The solids were filtered and the filtrate was concen-
trated and chromatographed (CHCl3 : CH3OHꢁ50 : 1) to give 12 as a foam
1
(34 mg, 80%). Compound 12: Rfꢁ0.20 CHCl3/MeOH (v/v 96 : 4) H-NMR
(CDCl3) d: 9.01 (1H, br, NHCO), 7.51 (1H, s, Ar-H), 7.41 (1H, dd, Jꢁ7.83,
7.43 Hz, Ar-H), 7.22 (1H, t, Jꢁ7.43 Hz, Ar-H), 7.13 (1H, t, Jꢁ7.43 Hz, Ar-
H), 6.93—6.82 (2H, m, H-8, H-4), 6.75 (1H, br, NHBoc), 6.34 (1H, br, NH),

1390
Vol. 53, No. 11
6.17—6.00 (3H, m, NH, H-7, H-3), 5.11—5.06 (1H, m, H-6), 4.41—4.37
(1H, m, Gln a-CH), 2.54—2.42 (4H, m, H-5, Gln g-CH2), 2.18—2.01 (2H,
m, Gln b-CH2), 1.42 (9H, s, Boc CH3ꢂ3). ESI/MS m/z 444 (MꢀH)ꢀ HR-
ESI/MS Calcd C23H29N3O6Na (MꢀNa)ꢀ: 466.1954, Found 466.1956.
10-(N-a-tert-Butoxycarbonyl-L-phenylalanyl)amino-goniothalamin
m, H-7, H-3), 5.02 (1H, m, H-6), 4.28 (1H, t, Jꢁ7.43 Hz, Ile a-CH), 2.47—
2.41 (2H, m, H-5), 1.91 (1H, m, Ile b-CH), 1.48 (1H, m, Ile g-CH2a), 0.95
(4H, m, Ile g-CH2b, b-CH3), 0.79—0.72 (3H, m, Ile g-CH3). ESI/MS m/z
427 (MꢅH)ꢅ HR-ESI/MS Calcd C21H24F3N2O5 (MꢅH)ꢅ: 441.1637, Found
441.1641.
(13)
A mixture of DCC (56 mg, 0.27 mmol), compound 4 (19 mg,
10-(a-L-Glutaminyl)amino-goniothalamin Trifluoroacetate (19)
A
0.09 mmol), N-tert-butoxycarbonyl-L-phenylalanine (72 mg, 0.27 mmol) and
CH2Cl2 (2.0 ml) was stirred at rt for 2 h. The solids were filtered, the filtrate
concentrated and then chromatographed (petroleum : ethyl acetateꢁ33 : 17)
to give 13 as a white powder (32 mg, 78%). Compound 13: Rfꢁ0.22 petro-
mixture of 12 (18 mg, 0.04 mmol), TFA (0.2 ml) and CH2Cl2 (1.5 ml) was
stirred 2 h at rt and concentrated to dryness. The residue was chro-
matographed (CHCl3 : MeOHꢁ46 : 6) to give 19 as a foam (12 mg, 65%).
1
Compound 19: Rfꢁ0.20 CHCl3/MeOH (v/v 86 : 14) H-NMR (CD3OD) d:
1
leum/ethyl acetate (v/v 64 : 36) H-NMR (CDCl3) d: 7.93 (1H, br, NHCO),
7.65 (1H, d, Jꢁ7.43 Hz, Ar-H), 7.45 (1H, d, Jꢁ7.82 Hz, Ar-H), 7.36—7.27
(2H, m, Ar-H), 7.11—7.06 (1H, m, H-4), 6.92 (1H, d, Jꢁ16.04 Hz, H-8),
6.36 (1H, m, H-7), 6.05 (1H, dd, Jꢁ9.78, 1.56 Hz, H-3), 5.22—5.17 (1H, m,
H-6), 4.19 (1H, t, Jꢁ6.26 Hz, Gln a-CH), 2.71—2.50 (4H, m, H-5 and Gln
g-CH2), 2.31—2.21 (2H, m, Gln b-CH2). ESI/MS m/z 456 (MꢅH)ꢅ HR-
ESI/MS Calcd C20H21F3N3O6 (MꢅH)ꢅ: 456.1382, Found 456.1386.
7.66—7.59 (1H, m, Ar-H), 7.38 (1H, d, Jꢁ7.81 Hz, Ar-H), 7.31—7.22 (6H,
m, Ar-H, Phe-H), 7.15 (1H, t, Jꢁ7.42 Hz, Ar-H), 6.94—6.88 (1H, m, H-4),
6.53 (1H, d, Jꢁ16.00 Hz, H-8), 6.11 (1H, dd, Jꢁ16.00, 6.63 Hz, H-7),
6.10—6.06 (1H, m, H-3), 5.25 (1H, br, NHBoc), 5.04—4.96 (1H, m, H-6),
4.50 (1H, d, Jꢁ6.25 Hz, Phe a-CH), 3.23—3.10 (2H, m, Phe b-CH2),
2.53—2.50 (2H, m, H-5), 1.41 (9H, s, Boc CH3ꢂ3). MALDI/MS m/z 485
(MꢀNa)ꢀ HR-MALDI/MS Calcd C27H30N2O5Na (MꢀNa)ꢀ: 485.2047,
Found. 485.2047.
10-(a-L-Phenylalanyl)amino-goniothalamin Trifluoroacetate (20)
A
mixture of 13 (28 mg, 0.06 mmol), TFA (0.5 ml) and CH2Cl2 (1.5 ml) was
stirred 1 h at rt and concentrated to dryness. The residue was chro-
matographed (CHCl3 : MeOHꢁ49 : 2) to give 20 as an amorphous powder
10-(a-L-Glycyl)amino-goniothalamin Trifluoroacetate (14) To a solu-
tion of trifluoroacetic acid (0.5 ml) and methylene chloride (1.5 ml) was
added 7 (18 mg, 0.05 mmol), and the mixture was stirred 1 h at rt. The sol-
vent was removed in vacuo and the residue was chromatographed
(CHCl3 : MeOHꢁ50 : 2) to give 14 as a yellow solid (11 mg, 59%). Com-
pound 14: Rfꢁ0.21 CHCl3/MeOH (v/v 92 : 8) 1H-NMR (CD3OD) d: 7.63
(1H, dd, Jꢁ7.81, 1.58 Hz, Ar-H), 7.44 (1H, d, Jꢁ7.81 Hz, Ar-H), 7.32 (1H,
t, Jꢁ7.81, 7.41 Hz, Ar-H), 7.26 (1H, t, Jꢁ7.41 Hz, Ar-H), 7.09 (1H, m, H-
4), 6.90 (1H, d, Jꢁ16.00 Hz, H-8), 6.36 (1H, dd, Jꢁ16.00, 6.24 Hz, H-7),
6.06—6.03 (1H, m, H-3), 5.21—5.16 (1H, m, H-6), 3.85 (2H, s, Gly CH2),
2.70—2.52 (2H, m, H-5). ESI/MS m/z 385 (MꢅH)ꢅ HR-ESI/MS Calcd
C17H16F3N2O5 (MꢅH)ꢅ: 385.1011, Found 385.1015.
10-(a-L-Alanyl)amino-goniothalamin Trifluoroacetate (15) 8 (38 mg,
0.10 mmol) was dissolved in CH2Cl2 (1.0 ml) and added to a stirring solution
of TFA (0.5 ml) and CH2Cl2 (0.5 ml). The mixture was stirred 1 h at rt, then
the solvent was removed in vacuo and chromatographed (CHCl3 : MeOHꢁ
45 : 5) to give 15 (27 mg, 69%) as a foam. Compound 15: Rfꢁ0.18
CHCl3/MeOH (v/v 94 : 6) 1H-NMR (CD3OD) d: 7.64 (1H, d, Jꢁ7.82 Hz,
Ar-H), 7.38 (1H, d, Jꢁ7.83 Hz, Ar-H), 7.34—7.26 (2H, m, Ar-H), 7.10—
7.06 (1H, m, H-4), 6.87 (1H, d, Jꢁ15.65 Hz, H-8), 6.36 (1H, dd, Jꢁ15.65,
6.26 Hz, H-7), 6.04 (1H, dd, Jꢁ9.78, 1.17 Hz, H-3), 5.20—5.15 (1H, m, H-
6), 4.04 (1H, q, Jꢁ7.42 Hz, Ala a-CH), 2.60 (2H, m, H-5), 1.60 (3H, d, Jꢁ
7.04 Hz, Ala b-CH3). ESI/MS m/z 399 (MꢅH)ꢅ HR-ESI/MS Calcd
C18H18F3N2O5 (MꢅH)ꢅ: 399.1168, Found 399.1171.
1
(17 mg, 59%). Compound 20: Rfꢁ0.20 CHCl3/MeOH (v/v 97 : 3) H-NMR
(CDCl3) d: 9.57 (1H, br, NHCO), 7.55 (1H, d, Jꢁ7.83 Hz, Ar-H), 7.33 (1H,
d, Jꢁ7.82 Hz, Ar-H), 7.29—7.14 (6H, m, Ar-H, Ph-H), 7.07 (1H, t, Jꢁ
7.83 Hz, Ar-H), 6.90—6.85 (1H, m, H-4), 6.64 (1H, dd, Jꢁ15.65, 5.87 Hz,
H-8), 6.06 (1H, dd, Jꢁ15.65, 6.62 Hz, H-7), 6.02 (1H, m, H-3), 5.04—4.99
(1H, m, H-6), 4.20 (2H, br s, NH2), 4.20 (1H, m, Phe a-CH), 3.26 (1H, dd,
Jꢁ13.69, 5.87 Hz, Phe b-CH2a), 3.06—2.99 (1H, m, Phe b-CH2b), 2.51—
2.43 (2H, m, H-5). ESI/MS m/z 475 (MꢅH)ꢅ HR-ESI/MS Calcd
C24H22F3N2O5 (MꢅH)ꢅ: 475.1481, Found 475.1484.
Approximate organic fragments p value for the Calculating the p Value
of the Derivative Molecules17—20): C (aliphatic) ꢀ0.5, phenyl ꢀ0.2, Cl ꢀ0.5,
OꢁC–O (carboxyl) ꢅ0.7, OꢁC–N (amide, imide) ꢅ0.7, O (hydroxyl, phe-
nol, ether) ꢅ0.1, N (amine) ꢅ1.0, NO2 (aromatic) ꢅ0.28.
Acknowledgements This work was supported by the Shanghai Natural
Science Foundation of China (2002ZB14020). The anticancer activity tests
were carried out by the Tumor Research Laboratory, Shanghai Institute of
Medicine Material, Chinese Academy of Sciences.
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10-(a-L-Valyl)amino-goniothalamin Trifluoroacetate (16) To a solu-
tion of TFA (0.5 ml) and CH2Cl2 (1.5 ml) was added 9 (32 mg, 0.08 mmol),
and the mixture was stirred 1 h at rt. The solvent was removed under reduced
pressure and the residue was chromatographed (CHCl3 : MeOHꢁ48 : 2) to
give 16 as a yellow solid (28 mg, 85%). Compound 16: Rfꢁ0.19 CHCl3/
MeOH (v/v 97 : 3) 1H-NMR (CD3OD) d: 7.61 (1H, dd, Jꢁ7.80, 1.17 Hz, Ar-
H), 7.45 (1H, dd, Jꢁ7.81, 1.17 Hz, Ar-H), 7.31 (1H, t, Jꢁ7.81, 7.41 Hz, Ar-
H), 7.24 (1H, t, Jꢁ7.41 Hz, Ar-H), 7.10—7.05 (1H, m, H-4), 6.92 (1H, d,
Jꢁ15.61 Hz, H-8), 6.35 (1H, dd, Jꢁ15.61, 6.22 Hz, H-7), 6.04 (1H, m, H-3),
5.20—5.15 (1H, m, H-6), 3.37 (1H, d, Jꢁ5.07 Hz, Val a-CH), 2.69—2.51
(2H, m, H-5), 2.17—2.12 (1H, m, Val b-CH), 1.09 (3H, d, Jꢁ7.02 Hz, Val
b-CH3), 1.02 (3H, d, Jꢁ6.63 Hz, Val g-CH3). ESI/MS m/z 427 (MꢅH)ꢅ
HR-ESI/MS Calcd C20H22F3N2O5 (MꢅH)ꢅ: 427.1484, Found 427.1476.
10-(a-L-Leucyl)amino-goniothalamin Trifluoroacetate (17) A mix-
ture of 10 (17 mg, 0.04 mmol) and TFA/CH2Cl2 (2 ml, 1 : 3) was stirred 1 h
at rt and concentrated to dryness. The residue was chromatographed
(CHCl3 : MeOHꢁ46 : 4) to give 17 as a foam (15 mg, 85%). Compound 17:
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1
Rfꢁ0.21 CHCl3/MeOH (v/v 97 : 3) H-NMR (CD3OD) d: 7.65 (1H, d, Jꢁ
7.42 Hz, Ar-H), 7.37—7.27 (3H, m, Ar-H), 7.10—7.06 (1H, m, H-4), 6.90
(1H, d, Jꢁ15.61 Hz, H-8), 6.37 (1H, dd, Jꢁ15.61, 5.47 Hz, H-7), 6.06—6.03
(1H, m, H-3), 5.20—5.14 (1H, m, H-6), 4.15—4.10 (1H, m, Leu a-CH),
2.71—2.63 (1H, m, H-5), 2.58—2.49 (1H, m, H-5), 1.90—1.81 (3H, m, Leu
b-CH2, g-CH), 1.09 (6H, m, Leu CH3ꢂ2). ESI/MS m/z 441 (MꢅH)ꢅ HR-
ESI/MS Calcd C21H24F3N2O5 (MꢅH)ꢅ: 441.1637, Found 441.1638.
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10-(a-L-Isoleucyl)amino-goniothalamin Trifluoroacetate (18) A mix-
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(CHCl3 : MeOHꢁ48 : 4) to give 18 as an amorphous solid (16 mg, 91%).
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9.72 (1H, br, NHCO), 8.33 (2H, br s, NH2), 7.38—7.32 (2H, m, Ar-H),
7.14—7.05 (2H, m, Ar-H), 6.90—6.78 (2H, m, H-8, H-4), 6.07—6.00 (2H,
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