1748 Bull. Chem. Soc. Jpn., 74, No. 9 (2001)
Synthesis of the Novel Spider Toxin NPTX-594
was concentrated in vacuo to give 4,8-bis(benzyloxycarbonyl)-
N1-[Nα,Nε-bis(benzyloxycarbonyl)lysyl]-N12-asparaginyl-4,8-
diaza-1,12-dodecanediamine (16) as a colorless oil (1.11 g,
92.5%).
This work was partly supported by a glant from the Re-
search for the Future Program from the Japan Society for the
Promotion of Science (JSPS), and a SUNBOR GRANT from
Suntory Institute for Bioorganic Research.
To a solution of a part of compound 16 (0.214 g, 0.219
mmol) in DMF (10 mL) were added 2,4-bis(benzyloxy)phen-
ylacetic acid succinimidyl ester11c (17) (0.107 g, 0.241 mmol)
and TEA (24.4 mg, 0.241 mmol), and the reaction mixture was
stirred for 3 h at r. t. The solvent was removed in vacuo, and
the residue was dissolved in AcOEt (50 mL). The solution was
washed with 10% citric acid (10 mL × 3), saturated aqueous
NaHCO3 (10 mL × 3), and brine (10 mL × 3). The organic
layer was dried over anhydrous MgSO4, and then the solvent
was removed in vacuo. The crude product was purified by sili-
ca-gel column chromatography (10 g, 1.0 × 20 cm, CHCl3:
MeOH = 100:1). The fractions containing desired product
were combined and concentrated in vacuo. The residue was
triturated with hexane to obtain 18 as colorless crystals, and
the thus-obtained crude product was recrystallized from dieth-
References
1
A part of this work was presented at the 25th European
Peptide Symposium: T. Wakamiya, A. Yamamoto, K. Kawaguchi,
Y. Yamaguchi, T. Fujita, M. Hisada, Y. Kan, Y. Itagaki, T.
Nakajima, and M. Andriantsiferana, “Peptides 1998,” ed by S.
Bajusz and F. Hudecz, Akadémiai Kiadó, Budapest (1999), p. 462.
2
(1982).
3
N. Kawai, A. Niwa, and T. Abe, Brain Res., 247, 169
NPTX-594 indicates a nephilatoxin whose molecular
weight is 594. Details about the isolation and structure determina-
tion will be reported soon in Natural Toxins.
4
Abbreviations according to IUPAC-IUB commission, Eur.
J. Biochem., 138, 9 (1984), are used. Asn: L-asparagine; Boc: t-
butoxycarbonyl; Boc2O: di-t-butyl dicarbonate; Bzl: benzyl;
DCHA: dicyclohexylamine; DMF: N,N-dimethylformamide; DM-
SO: dimethyl sulfoxide; EDC: 1-(3-dimethylaminopropyl)-3-eth-
ylcarbodiimide; HRFAB-MS: high resolution fast atom bombard-
ment mass spectrometry; HOBt: 1-hydroxybenzotriazole; HOSu:
N-hydroxysuccinimide; Lys: L-lysine; Ms: methanesulfonyl; Np:
p-nitrophenyl (or 4-nitrophenyl); RPHPLC: reversed-phase high-
performance liquid chromatography; TEA: triethylamine; TFA:
trifluoroacetic acid; THF: tetrahydrofuran; Troc: 2,2,2-trichloro-
ethoxycarbonyl; TrocCl: 2,2,2-trichloroethoxycarbonyl chloride;
Z: benzyloxycarbonyl; ZCl: benzyloxycarbonyl chloride.
1
yl ether and hexane (0.181 g, 63.3%). Mp 130–134 °C; H
NMR (500 MHz, DMSO-d6) δ 1.20–1.72 (6H, m, β-, γ-, and δ-
CH2/Lys; 8H, m, CH2/Dada × 4), 2.39 and 2.44 (each 1H, dd,
β-CH2/Asn), 2.90–3.21 (14H, m, CO–N–CH2/Dada × 6 and ε-
CH2/Lys), 3.42 (2H, s, PhCH2/Dhpa), 3.89 (1H, m, α-CH/Lys),
4.51 (1H, m, α-CH/Asn), 4.96–5.06 (8H, PhCH2/Z × 4), 5.04
and 5.08 (each 2H, s, PhCH2/Bzl), 6.53 (1H, dd, C3-Ph /Dhpa),
6.68 (1H, d, C5-Ph /Dhpa), 7.08 (1H, d, C6-Ph /Dhpa), and
7.24–7.38 (30H, Ph/Z and Bzl × 6); HRFAB-MS: found m/z
1311.6325 (M+H)+ (calcd for C74H86N8O14 + H: 1311.6342);
[α]D19 −3.5° (c 1.0, MeOH).
5
a) Y. Aramaki, T. Yasuhara, K. Shimazaki, A. Kawai, and
N12-[Nα-(2,4-Dihydroxyphenylacetyl)asparaginyl]-N1-ly-
syl-4,8-diaza-1,12-dodecanediamine (NPTX-594) (1). To a
solution of compound 18 (50.0 mg, 0.0382 mmol) in MeOH
(10 mL) and AcOH (20 mL) was added Pd black (50 mg). The
reaction mixture was stirred under an atmosphere of hydrogen
for 2 h at r. t. The catalyst was filtered off, and the filtrate was
concentrated in vacuo. The crude product was dissolved in
water and filtered through Millipore® filter (Millipore Corp.,
Type: HA, pore size: 0.45 µm); the filtrate was then lyo-
philized. The crude product was finally purified by preparative
RPHPLC (10–40% CH3CN–0.1% TFA/8.0 mL min−1). The
fractions containing the desired compound were combined and
lyophilized. The thus-obtained powdery substance was dis-
solved in 0.1 M HCl (10 mL), and the solution was lyophilized
again to obtain 1 as hydrochloride19 (16.7 mg, 59.0%). 1H
NMR (500 MHz, D2O; auto-reference condition: HDO = 4.75
ppm) δ 1.37 (2H, m, γ-CH2/Lys), 1.40–1 52 (4H, m, C10H2 and
C11H2/Dada), 1.62 (2H, m, δ-CH2 /Lys), 1.83 (2H, m, β-CH2/
Lys), 1.85 (2H, m, C2H2/Dada), 1.97 (2H, m, C6H2/Dada), 2.64
and 2.70 (each 1H, dd, β-CH2 /Asn), 2.88 (2H, m, C9H2/Dada),
2.92 (2H, m, ε-CH2/Lys), 2.96 and 3.03 (each 2H, m, C5H2,
C7H2/Dada), 3.01 (2H, m, C3H2/Dada), 3.09 and 3.21 (each
1H, m, C12H2/Dada), 3.21 and 3.31 (each 1H, m, C1H2/Dada),
3.45 (2H, m, CH2/Dhpa), 3.89 (1H, m, α-CH/Lys), 4.49 (1H,
m, α-CH/Asn), 6.37 (1H, s, C3-Ph/Dhpa), 6.38 (1H, d, C5-Ph/
Dhpa), and 7.02 (1H, d, C6-Ph/Dhpa); FAB-MS: found m/z
595.3946 (M+H)+ (calcd for C28H50N8O6 + H: 595.3935);
[α]D19 −7.9° (c 1.0, H2O).
T. Nakajima, Biomed. Res., 8, 241 (1987). b) T. Toki, T. Yasuhara,
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