MAGNETIC RESONANCE IN CHEMISTRY Magn. Reson. Chem. 2005; 43: 595–598 Published online in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/mrc.1600 Spectral Assignments and Reference Data Structural and spectral assignment of three forskolin-like diterpenoids isolated from Plectranthus ornatus Patr´ ıcia Rijo, 1 M. F ´ atima Sim ˜ oes 1 and Benjam´ ın Rodr´ ıguez 2∗ 1 Faculdade de Farm ´ acia da Universidade de Lisboa, CECF, Av. Prof. Gama Pinto, 1649-003 Lisbon, Portugal 2 Instituto de Qu´ ımica Org ´ anica, CSIC, Juan de la Cierva 3, E-28006 Madrid, Spain Received 22 February 2005; revised 5 April 2005; accepted 6 April 2005 Three labdane diterpenoids were isolated from an acetone extract of Plectranthus ornatus. Their structures, closely related to that of forskolin, were determined by NMR studies. Unambiguous and complete assignments of the 1 H and 13 C NMR chemical shifts for these substances are presented. The assignments are based on 2D shift-correlated [ 1 H, 1 H-COSY, 1 H, 13 C-gHSQC- 1 J (C,H), 1 H, 13 C-gHMBC- n J (C,H) (n = 2 and 3)] and NOE experiments. Copyright  2005 John Wiley & Sons, Ltd. KEYWORDS: NMR; 1 H NMR; 13 C NMR; 1D NMR; 2D NMR; Plectranthus ornatus; diterpenes; labdane derivatives INTRODUCTION Many Plectranthus species (family Lamiaceae) are plants of orna- mental, economic and medicinal interest. Several species are used in tropical Africa and Polynesia as vermicides, antiseptics and purga- tives, for the treatment of ear infections, toothache and stomach ache and as a remedy for vomiting and nausea. 1 Several Plectranthus plants have been studied chemically and a large number of abietane, phyl- locladane, kaurane, clerodane and labdane diterpenoids have been isolated from them, 1–4 together with long-chain alkylphenols, 1 aris- tolane sesquiterpenes, 3,5 ursane, oleanane and lupane triterpenoids 1 and flavonoid compounds. 1 Recently, we reported 2 the isolation of three new diterpenoids, one clerodane and two labdane derivatives, from an acetone extract of Plectranthus ornatus Codd. (synonym P. comosus Hochst. ex G ¨ uerke) whole plant. In this paper, we report on the isolation and structure elucidation of three other labdane diterpenoids (1 – 3) that were found, together with the triterpenoids ˛- and ˇ-amyrin and ursolic and oleanolic acids, in the same acetone extract of the plant. Compounds 1 – 3 possess structures closely related to that of forskolin (4), a very interesting substance isolated from Coleus forskohlii Briq. (Lamiaceae), displaying unique cardiotonic, platelet aggregation inhibitory, bronchodilatory, antihypertensive, intraoc- ular pressure-lowering, anti-inflammatory and adenylate cyclase- activating properties, and whose synthesis and chemical reactivity have been extensively studied. 6 This is the first report on 1 – 3 as naturally occurring substances, although they are already known as semisynthetic derivatives. 6-O-Acetylforskolin (1) and 1,6-di-O- acetylforskolin (2) have previously been obtained by acetylation of forskolin (4), 7 whereas 3 (1,6-di-O-acetyl-9-deoxyforskolin) has been reported as an intermediate in a semisynthesis of 4 starting from 9-deoxyforskolin (5). 8,9 Structures 1 – 3 were established by NMR spectroscopic studies and, since only partial 1 H NMR data for 1 and 2 had been reported, 7 Ł Correspondence to: Benjam´ ın Rodr´ ıguez, Instituto de Qu´ ımica Org´ anica, CSIC, Juan de la Cierva 3, E-28006 Madrid, Spain. E-mail: iqor107@iqog.csic.es Contract/grant sponsor: FCT; Contract/grant number: I&D No. 8/94. Contract/grant sponsor: POCTI; Contract/grant number: QCA III. Contract/grant sponsor: FEDER. Contract/grant sponsor: Comisi ´ on Interministerial de Ciencia y Tecnolog´ ıa (CICyT); Contract/grant number: 5653. we decided to perform the complete assignment of the 1 H and 13 C NMR spectra of these substances in order to provide several sets of data that might serve as models for future assignments of other structurally related compounds. RESULTS AND DISCUSSION Repeated chromatographic processes on the fractions from the initial chromatography eluted with 1 : 1 EtOAc–light petroleum of the acetone extract of P. ornatus (see Experimental) allowed the isolation of 1 – 3 together with a mixture of ˛- and ˇ-amyrin (1 : 2) and a mixture of ursolic and oleanolic acids (1 : 1). For the assignment of the 1 H and 13 C NMR spectra of 1 – 3 (Tables 1 and 2, respectively), a combination of two-dimensional COSY, gHSQC and gHMBC experiments was carried out, together with 1D NOESY spectra for establishing the relative stereochemistry and conformations of these substances. Low-resolution mass spectrometry and combustion analysis indicated the molecular formula C 24 H 36 O 8 for 1 and its 1 H and 13 C NMR spectra (Tables 1 and 2, respectively) were almost identical with those reported 10–13 for forskolin (4,C 22 H 34 O 7 ). In fact, the observed differences between the 1 H and 13 C NMR spectra of 1 and 4 were compatible with the presence in the former of a 6ˇ-acetoxyl group instead of the 6ˇ-hydroxyl function in the latter. This conclusion is supported by the following facts. Compound 1 possesses two acetoxyl groups, as was evidenced by its molecular formula and confirmed by the presence of two three-proton singlets (υ 2.13 and 2.00) and two pairs of acetoxyl carbons (υ 170.49 s, 170.31 s, 21.28 q and 20.95 q) in the 1 H and 13 C NMR spectra, respectively. The H-6˛ proton of 1 appeared downfield shifted (υ 6.20) with respect to that of 4 (υ 4.44). 13 The C-1–C-4 and C-8–C-20 carbon atom resonances of 1 (Table 2) were almost identical with those of 4, 13 whereas the observed differences in the chemical shifts of the C-5, C-6 and C-7 carbons [υ D υ1  υ4: 0.2, C1.2 and 1.6 ppm, respectively] are compatible only with the presence in 1 of an acetoxyl substituent at the 6ˇ-position. Finally, the gHMBC spectrum of 1 showed connectivity between the carbonyl carbon of the 6-O-acetyl group (υ 170.49 s) and the H-6˛ proton (υ 6.20 dd), which, in turn, was connected with the C-5, C-7, C-8 and C-10 carbons, whereas the H-7˛ proton showed gHMBC cross peaks with the carbonyl carbon of the other acetate (υ 170.31 s) and the C-6, C-8, C-9 and C-17 carbons. Hence 1 is 6-O-acetylforskolin. Compound 2 is the 1-O-acetyl derivative of 1, as was evidenced by its molecular formula (C 26 H 38 O 9 ) and the presence of three acetoxyl groups in its 1 H and 13 C NMR spectra [υ H 2.09 s, 2.02 s and 2.01 s, 3H each; υ C 169.87 s (2C), 168.40 s, 21.74 q, 21.44 q and 20.82 q]. Comparison of the chemical shift of the H-1ˇ proton of 2 (υ 5.56 t) with respect to those of 1 (υ 5.01 ddd, see Table 1) and 4 (υ 4.54 ddd), 13 together with the diamagnetic shift of the C-2 carbon of 2 (υ 23.11) with respect to those of 1 (υ 27.06) and 4 (υ 26.46), 13 strongly supported that 2 is 1,6-di-O-acetylforskolin. Diterpenoid 3 is the 9-deoxy derivative of 2. The molecular for- mula established for 3 was C 26 H 38 O 8 instead of C 26 H 38 O 9 for 2. The IR spectrum of 3 was devoid of hydroxyl absorptions, whereas 2 Copyright  2005 John Wiley & Sons, Ltd.