Mucronine J, a 14-Membered Cyclopeptide Alkaloid from Zizyphus mucronata Catherine Auvin, Franc ¸ oise Lezenven, Alain Blond, Isabelle Augeven-Bour, Jean-Louis Pousset, and Bernard Bodo* Laboratoire de Chimie des Substances Naturelles, URA 401 CNRS, GDR 964 CNRS, Muse ´ um National d’Histoire Naturelle, 63 rue Buffon, 75231 Paris Cedex 05, France Jose ´ Camara Institut Henri Beaufour, 1 avenue des Tropiques, ZA Courtaboeuf, 91940 Les Ulis, France Received October 25, 1995 X From the CH 2 Cl 2 extract of the root bark of Z. mucronata (Rhamnaceae), a new cyclopeptide alkaloid, named mucronine J, was isolated together with previously known alkaloids abyssenine A and mucronine D. The structure of mucronine J was elucided by mass spectrometry and 1D and 2D NMR. A solution conformation is proposed on the basis of NOE experiments in combination with MM2 calculations. Zizyphus mucronata Willd., a Rhamnaceae of central and southern Africa, is employed by the natives in the treatment of diarrhea, dysentery, and skin infections. 1 Previous chemical studies on the bark revealed cyclo- peptide alkaloids of the mucronine family. 2-7 All were characterized by the presence of a styrylamine group in a 15-membered cyclopeptide alkaloid, except mucro- nine D, which is a 13-membered ring system. From the root bark, a new 14-membered cyclopeptide alkaloid (1) together with the known abyssenine A 6 and mucronine D 7 were isolated. In this paper, we report the isolation, structure determination, and molecular modeling of mucronine J (1). The peptide alkaloid 1 was isolated from the alkaloi- dal fraction of the CH 2 Cl 2 extract of Z. mucronata root bark by successive column chromatography and gave a weak positive reaction with Dragendorff’s reagent. The molecular formula C 27 H 40 N 4 O 4 was derived from the HR positive ion FABMS where the MH + ion was observed at m/z 485.3088. The EIMS exhibited the characteristic fragmentations of a 14-membered cyclopeptide alkaloid such as amphibine. 8,9 The presence of an ion at m/z 135 was typical of p-hydroxystyrylamine. The ions at m/z 86 and 72 suggested leucine or isoleucine as a ring- bonded amino acid. Moreover, the presence of an N,N- dimethylleucine (or N,N-dimethylisoleucine) unit as the terminal residue was deduced from the occurrence of the base peak at m/z 114. The 1 H NMR spectrum of 1 displayed signals for two ethylenic protons, four aromatic protons, and three amino acids, -hydroxyproline, isoleucine, and N,N- dimethylleucine, which were identified and assigned with the aid of 1 H- 1 H COSY experiment (Table 1). 1 H and 13 C assignments arose from the analysis of J-modulated 13 C, HMQC, and HMBC spectra. The carbonyl resonances at C-4, C-7, and C-22 were assigned from their 2 J C-H intraresidue correlations with R-pro- tons at H-5, H-8, and H-23, respectively. HMBC cor- relations from the oxygenated sp 2 quaternary carbon (C- 11) (δ 157.4) to H-12, H-12′, H-13, and H-13′ confirmed a para-substituted aromatic ring. As previously ob- served for 14-membered cyclopeptide alkaloids, 10 con- formational constraints in the macrocyclic ring system induced the nonequivalence of the protons and carbon atoms of each aromatic methine pair (C-12 and -12′, C-13 and -13′). The observed HMBC correlations from C-13 (δ 132.6) to H-12 (δ 7.28) and from C-13′ (δ 130.2) to H-12′ (δ 7.10) allowed their unambiguous identifica- tion. The combined analysis of HMBC and HMQC data ascertained the linkages between constitutive parts of 1 and allowed full resonance assignment (Table 1). The two ethylenic protons at δ 6.27 (H-1) and 6.74 (H-2) had a mutual coupling constant value of 7.7 Hz, indicating a cis stereochemistry, which was confirmed by their strong mutual NOE effect. NH-3 did not give NOE with these ethylenic protons, but a strong effect with aromatic protons H-13 and H-13′. Moreover, the coupling constant value between H-2 and NH-3 ( 3 J ) 10.8 Hz) indicated they were in a trans coplanar position. Strong NOEs were observed between H-9 and both H-19R and H-20R but not between H-19 and H-20, indicating that H-9, H-19R, and H-20R were on X Abstract published in Advance ACS Abstracts, June 1, 1996. 676 J. Nat. Prod. 1996, 59, 676-678 S0163-3864(96)00197-8 CCC: $12.00 © 1996 American Chemical Society and American Society of Pharmacognosy