796
M. Abe et al. / Bioorg. Med. Chem. 13 (2005) 785–797
1.90–2.09 (3H, m, H-4a, H-5a, and H-6a), 1.70–1.90
(2H, m, H-4b and H-5b), 1.45 (1H, m, H-6b); 13C
NMR (100MHz, D2O) d 177.9 (C-2), 175.9 (CHCOO),
170.7 (PhCOO), 136.9 (Ph), 132.7 (Ph · 2), 131.8 (Ph),
131.6 (Ph · 2), 78.1 (OCHCOO), 57.0 (C-3), 47.5 (C-
7), 31.2 (C-4), 30.4 (C-6), 29.1 (C-5); 31P NMR
(243MHz, D2O) d 18.30; HR-ESIMS m/z calcd for
C6H15N4NaO4P (M+Na)+ 229.0830, found 229.0824.
Acknowledgements
We thank Dr. H. Naganawa, Dr. R. Sawa and Ms. Y.
Kubota (Microbial Chemistry Research Center) for
helpful advice and measurement of various spectral
data.
References and notes
7.27. (3R)-3-Amino-1-diaminophosphinyl-2-piperidinone
1/2 (ꢀ)-dibenzoyl-L-tartarate (27c)
1. Heins, J.; Welker, P.; Scho¨nlein, C.; Born, I.; Hartrodt, B.;
Neubert, K.; Tsuru, D.; Barth, A. Biochim. Biophys. Acta
1988, 954, 161.
2. For reviews on DPP-IV, see: (a) Augustyns, K.; Bal, G.;
Thonus, G.; Belyaev, A.; Zhang, X. M.; Bollaert, W.;
Lambeir, A. M.; Durinx, C.; Goossens, F.; Haemers, A.
Curr. Med. Chem. 1999, 6, 311; (b) De Meester, I.; Korom,
The synthetic method similar to that noted for com-
pound 6 was used. (3R)-3-Benzyloxycarbonylamino-1-
diaminophosphinyl-2-piperidinone
(26)
(700.0mg,
2.145mmol) was treated with Pd-black (35.0mg) under
a hydrogen atmosphere. The reaction mixture was fil-
trated, and addition of (ꢀ)-dibenzoyl-L-tartaric acid
(403.7mg, 1.073mmol) afforded compound 27b
(659.8mg) in 83% yield as a colorless powder: 1H
NMR (400MHz, D2O) d 8.13 (2H, d, J = 7.8Hz,
COPh), 7.72 (1H, m, COPh), 7.57 (2H, t, J = 7.8Hz,
COPh), 5.72 (1H, s, OCHCOO), 4.10 (1H, dd,
J = 11.7, 6.8Hz, H-3), 3.65 (2H, td, J = 5.9, 4.9Hz, H-
6), 2.37 (1H, m, H-4a), 1.79–2.07 (3H, m, H-4b and
H-5); 13C NMR (100MHz, D2O) d 175.9 (CHCOO),
174.6 (C-2), 170.7 (PhCOO), 136.9 (Ph), 132.7
(Ph · 2), 131.8 (Ph), 131.6 (Ph · 2), 78.1 (OCHCOO),
53.3 (C-3), 47.3 (C-6), 26.6 (C-4), 23.1 (C-5); 31P
NMR (243MHz, D2O) d 17.33; HR-ESIMS m/z calcd
for C5H13N4NaO2P (M+Na)+ 215.0674, found
215.0683.
´
S.; Van Damme, J.; Scharpe, S. Immunol. Today 1999, 20,
367.
3. Mentlein, R.; Gallwitz, B.; Schmidt, W. E. Eur. J.
Biochem. 1993, 214, 829.
4. (a) Balkan, B.; Kwasnik, L.; Miserendino, R.; Holst, J. J.;
Li, X. Diabetologia 1999, 42, 1324; (b) Pederson, R. A.;
White, H. A.; Schlenzig, D.; Pauly, R. P.; McIntosh, C.
H.; Demuth, H.-U. Diabetes 1998, 47, 1253.
´
5. Ahren, B.; Simonsson, E.; Larsson, H.; Landin-Olsson,
M.; Torgeirsson, H.; Jansson, P. A.; Sandqvist, M.;
˚
Bavenholm, P.; Efendic, S.; Eriksson, J. W.; Dickinson,
S.; Holmes, D. Diabetes Care 2002, 25, 869.
6. Sorbera, L. A.; Revel, L.; Castan˜er, J. Drugs Future 2001,
26, 859.
7. (a) Bristol, L. A.; Sakaguchi, K.; Appella, E.; Doyle, D.;
´
Takacs, L. J. Immunol. 1992, 149, 367; (b) Vivier, I.;
Marguet, D.; Naquet, P.; Bonicel, J.; Black, D.; Li, C. X.;
Bernard, A. M.; Gorvel, J. P.; Pierres, M. J. Immunol.
1991, 147, 447.
7.28. In vitro assay for DPP-IV inhibitory activity
8. Korom, S.; De Weester, I.; Stadlbauer, T. H.; Chandraker,
A.; Schaub, M.; Sayegh, M. H.; Belyaev, A.; Haemers, A.;
0.1M Tris(hydroxymethyl)-aminomethane/maleic acid
buffer solution (pH7.2, 100lL), 3.2mM Gly-Pro-b-
naphthylamide (25lL, Bachem, Switzerland), and an
aqueous solution (50lL) of the synthetic compound
were added into 96 well micro-plates. The resultant solu-
tion was warmed for 10min at 37ꢂC, and then the DPP-
IV solution (25lL) was added to the solution. The com-
bined solution was allowed to react for 1h at 37ꢂC. The
reaction was terminated by adding a solution (100lL) of
0.2% Fast Garnet GBC salt (Sigma, USA) in 0.5M so-
dium citrate buffer solution (pH3.78) including 10%
polyoxyethylene (20) sorbitan monolaulate (Wako, Ja-
pan). The absorbance at 525nm was measured (value
a). Simultaneously, the absorbance of the reaction mix-
ture without the synthetic compound solution was meas-
ured (value b). Moreover, the absorbance of the reaction
mixture without the DPP-IV solution was measured,
respectively (value a0 and value b0). The DPP-IV inhibi-
tory rate (%) was calculated by [(b ꢀ b0) ꢀ (a ꢀ a0)/
(b ꢀ b0)] · 100. From the inhibitory curve, taking inhib-
itory rate (%) at various concentrations of the synthetic
compound on the ordinate and the logarithm of concen-
trations of the synthetic compound on the abscissa, the
concentration for 50% inhibition was obtained. In all
cases, a linear relation was observed between 20% and
80% inhibition. This procedure was repeated three times
or more, and each average was shown as the IC50 value
of the synthetic compound.
´
Scharpe, S.; Kupiec-Weglinski, J. W. Transplantation
1997, 63, 1495.
9. Tanaka, S.; Murakami, T.; Horikawa, H.; Sugiura, M.;
Kawashima, K.; Sugita, T. Int. J. Immunopharmacol.
1997, 19, 15.
10. Umezawa, H.; Aoyagi, T.; Ogawa, K.; Naganawa, H.;
Hamada, M.; Takeuchi, T. J. Antibiot. 1984, 37,
422.
11. Akiyama, T.; Abe, M.; Harada, S.; Kojima, F.; Sawa, R.;
Takahashi, Y.; Naganawa, H.; Homma, Y.; Hamada, M.;
Yamaguchi, A.; Aoyagi, T.; Muraoka, Y.; Takeuchi, T. J.
Antibiot. 2001, 54, 744.
12. Abe, M.; Akiyama, T.; Nakamura, H.; Kojima, F.;
Harada, S.; Muraoka, Y. J. Nat. Prod. 2004, 67,
999.
13. Kokotos, G.; Markidis, T.; Violetta, C.-K. Synthesis 1996,
1223.
14. Ranganathan, S.; Ranganathan, D.; Singh, W. P. Tetra-
hedron Lett. 1988, 29, 3111.
15. Oya, H.; Nagatsu, I.; Nagatsu, T. Biochim. Biophys. Acta
1972, 258, 591.
16. Hughes, T. E.; Mone, M. D.; Russell, M. E.; Weldon, S.
C.; Villhauer, E. B. Biochemistry 1999, 38, 11597.
17. Coutts, S. J.; Kelly, T. A.; Snow, R. J.; Kennedy, C. A.;
Barton, R. W.; Adams, J.; Krolikowski, D. A.; Freeman,
D. M.; Campbell, S. J.; Ksiazek, J. F.; Bachovchin, W. W.
J. Med. Chem. 1996, 39, 2087.
18. Belyaev, A.; Zhang, X.; Augustyns, K.; Lambeir, A. M.;
´
De Meester, I.; Vedernikova, I.; Scharpe, S.; Haemers, A.
J. Med. Chem. 1999, 42, 1041.