Novel steroidal penta and hexacyclic com

Steroids 75 (2010) 1127–1136 Contents lists available at ScienceDirect Steroids journal homepage: www.elsevier.com/locate/steroids Novel steroidal penta- and hexacyclic compounds derived from 12-oxospirostan sapogenins José Oscar H Pérez-Díaz a , José Luis Vega-Baez b , Jesús Sandoval-Ramírez b , Socorro Meza-Reyes b,∗ , Sara Montiel-Smith b , Norberto Farfán c , Rosa Santillan a,∗∗ a b c Departamento de Química, Centro de Investigación y de Estudios Avanzados del IPN, Apdo Postal 14-740, 07000 México D.F., Mexico Facultad de Ciencias Químicas, Benemérita Universidad Autónoma de Puebla, Ciudad Universitaria, Col San Manuel, C.P 72570, Puebla, Pue., Mexico Facultad de Química, Departamento de Química Orgánica, Universidad Nacional Autónoma de México, Ciudad Universitaria 04510, Mexico D.F., Mexico a r t i c l e i n f o Article history: Received 12 May 2010 Received in revised form 12 July 2010 Accepted 14 July 2010 Available online 22 July 2010 Keywords: 12-Oxosapogenins Polycyclic steroids Intramolecular cyclocondensation a b s t r a c t The E ring regioselective acid-catalyzed opening of spirostanic sapogenins possessing a carbonyl group at C-12, such as botogenin and hecogenin, provided the new 12,23-cyclo-22,26-epoxycholesta-11,22-diene skeleton, in addition to new compounds of the already known 12,23-cyclocholest-12(23)-en-22-one frameworks This transformation proceeds in a single step, under slightly acidic conditions Both, pentaand hexacyclic steroids were obtained with retention of configuration of all asymmetric centers © 2010 Elsevier Inc All rights reserved Introduction Steroidal derivatives have been a rich source of agents with potential pharmaceutical applications that have inspired the synthesis of new analogs with increased pharmacological activity By 1990 about 80% of the drugs used were either natural products or synthetic analogs; nowadays almost 50% of the approved drugs are still based on natural products [1] The spirostan sapogenins are a particular type of steroidal derivatives, widely spread in plants and some marine organisms, which exhibit a broad range of biological activities [2–5] These natural products are found as steroidal glycosides (saponins) The sugar moiety is greatly diversified and the carbohydrate units can be obtained through acidic or enzymatic hydrolysis The steroidal sapogenins have served for many years as cheap raw material for the pharmaceutical industry in the synthesis of sex and adrenocortical hormones, analogs of vitamin D, anabolics and antiinflammatory drugs [6–8] Steroidal sapogenins can be divided into cholestanic, furostanic and spirostanic derivatives; the latter ones contain a particular spiroketal system (E/F rings) at the side chain, which is stabilized by anomeric effects [9] ∗ Corresponding author Tel.: +52 22 2229 5500x7382; fax: +52 22 2229 5584 ∗∗ Corresponding author Tel.: +52 55 5747 3725; fax: +52 55 5747 3389 E-mail addresses: msmeza@siu.buap.mx (S Meza-Reyes), rsantill@cinvestav.mx (R Santillan) 0039-128X/$ – see front matter © 2010 Elsevier Inc All rights reserved doi:10.1016/j.steroids.2010.07.008 The variants of 12-oxosapogenins, botogenin (1a) and hecogenin (2a) (Fig 1) are suitable starting materials for the synthesis of 9,11 and 11,12 steroidal epoxides [10], cephalostatines [2,11] and ritterazines [4,12], compounds with potential application for cancer chemoprevention; also they have been used in the synthesis of drugs such as cortisone, betamethasone [13,14], as well as the preparation of compounds which are potent cholesterol absorption inhibitors [15] The C-12 ketone group shows remarkable low reactivity mainly attributed to the steric hindrance caused by the angular C-18 and C19 methyl groups in the ␤ face This hypothesis was corroborated by the reduction of the C-12 carbonyl group with NaBH4 and by hydrogenation [16–18] The cleavage of the spiroketal ring has been widely investigated under a great variety of reaction conditions providing different kinds of skeletons, sometimes under similar reaction conditions [19–22] Suárez and coworkers described for the first time the cleavage of the E ring of spirostanic sapogenins catalyzed by BF3 obtaining the 22,26-epoxy-5␣-cholest-22-ene with an excellent yield, from a non-functionalized (25R)-spirostan compound [21] Singh and Dhar however, obtained the 22,26-epoxycholesta-3,5dien-16-one skeleton 4, treating diosgenin with BF3 ·OEt2 at high temperature [23] Similar results to those of Suárez were obtained by our work group, thus the 22,26-epoxy-5␣-cholest-22-en-12one was prepared by regioselective cleavage of the E ring of (25R)-sapogenins such as hecogenin (Fig 2) [24,25] Further studies showed that when the same reaction conditions are applied to sarsasapogenin, a (25S)-sapogenin with a pronounced steric 1128 J.O.H Pérez-Díaz et al / Steroids 75 (2010) 1127–1136 Fig Spirostan sapogenins with a carbonyl group at C-12 hindrance due to the axial C-27 methyl group, the 22,26-epoxy5␤-cholest-22-ene derivative is obtained in a lower yield together with the acetyl pseudosapogenins and [26] Moreover, modifying reaction conditions, the treatment with BF3 ·OEt2 on sapogenins could direct to the 22-oxocholestanic frameworks and [27,28] More recently, LaCour et al [29] reported that cleavage of hecogenin (2a) with Ph3 P·I2 at high temperatures gives an aromatic hexacyclic product derived from an intramolecular aldol condensation In this transformation, aromaticity is achieved by the elimination of four asymmetric centers In continuation with our studies on the synthesis of new steroidal derivatives, we report hereupon the synthesis of new polycyclic frameworks from botogenin (1b) and hecogenin (2b) acetates Experimental 2.1 General methods IR spectra were acquired on a FT-IR Perkin-Elmer Spectrum GX spectrophotometer using KBr pellets, or a FT-IR Varian 640 spectrophotometer using ATR ( cm−1 ) Optical rotations [˛]25 D were obtained using dichloromethane or chloroform solutions on a Perkin-Elmer 241 polarimeter NMR spectra (1 H, 13 C, DEPT, HSQC, HETCOR and COSY) were determined with a JEOL eclipse +400, and a VARIAN Mercury 400 spectrometer, chemical shifts are stated in ppm (ı), and are referred to the residual H signal (ı = 7.27) or to the central 13 C triplet signal (ı = 77.0) for CDCl3 , compound 12 was referred to the residual H signals (ı = 8.74) or to the 13 C pyridine-d5 (ı = 150.35) HMBC and ROESY experiments were performed on a JEOL ECA 500 Mass spectra were obtained at 70 eV with a Hewlett Packard 5989A and a Hewlett Packard 6890A mass spectrometer High resolution mass spectra (HRMS) were determined on an Agilent Technologies, model 1100 coupled MSD-TOF spectrophotometer with APCI as ionization source Thin-layer chromatograms were developed on aluminum TLC pre-coated sheets with silica gel 60 with fluorescent indicator F254 , visualized by UV and by calcinations with 50% sulfuric acid 2.2 Crystal structure determination Crystals of 16 suitable for X-ray were obtained from hexane: ethyl acetate, by slow evaporation of the solvent at room temperature Data collection was performed at 293 K on a Kappa CCD ˚ The structure diffractometer with Mo K␣-radiation, ␭ = 0.7107 A was solved by direct methods SHELXS-86 [30] and refined using CRYSTALS [31] All non-hydrogen atoms were refined anisotropically The H atoms attached to O atoms were refined freely and the remaining H atoms were refined using a riding model Crystallographic data have been deposited at the Cambridge Crystallographic Center No 757895 Copy of the data can be obtained free of charge from CCDC, 12 Union Road, Cambridge CB2 1EZ, UK E-mail: deposit@ccdc.cam.ac.uk 2.2.1 Crystal data for compound 16 C33 H46 O7 , colorless prisms, formula weight M = 554.72, ˚ ˚ orthorhombic, P 21 21 21 , a = 11.7915(2) A, b = 12.2100(2) A, ˚ V = 3684.13(12) A˚ Z = 4, c = 25.5888(6) A, ˛ = ˇ = = 90◦ , Dx = 1.00 Mgm−3 , = 0.07 mm−1 , F(0 0) = 1200 Collected reflections: 8564 within a theta range of 2.9–27.9◦ (−14 ≤ h ≥ 14, −16 ≤ k ≥ 16, −33 ≤ l ≥ 33) Refinement: final R = 0.075, goodnessof-fit = 1.083; 8564 reflections, 362 parameters, maximum and Fig Variety of structures obtained from the treatment of spirostanic sapogenins with BF3 ·OEt2 J.O.H Pérez-Díaz et al / Steroids 75 (2010) 1127–1136 minimum difference electron densities were −0.24 eA˚ −3 and 0.14 eA˚ −3 respectively 2.3 Synthesis of polycyclic steroidal derivatives from 12-oxosapogenin acetate with BF3 ·OEt2 In a 25 mL round bottom flask were dissolved 0.50 g (1.06 mmol) of sapogenin acetate (1b or 2b) in 5.0 mL of dichloromethane; 0.70 mL (6.36 mmol) of acetic anhydride and 0.72 mL (4.24 mmol) of BF3 ·OEt2 were added slowly The mixture was stirred at 25 ◦ C and monitored by TLC until complete disappearance of starting material (45 min) The reaction mixture was poured over ground ice (20 g) and vigorously shaken The organic phase was extracted with AcOEt, washed with brine (2 × 20 mL), followed by neutralization with saturated solution of NaHCO3 , dried over anhydrous Na2 SO4 and concentrated to dryness under vacuum The crude product of the reaction with botogenin acetate (1b) (0.60 g) was chromatographed over silica gel (230–400 mesh) using hexane/AcOEt (90:10) to give 0.16 g of 10 (27%), 0.13 g of 12 (22%), 0.03 g of 14 (5%), 0.04 g of 16 (7%), and 0.17 g of 17 (29%) The purification of the crude reaction product using hecogenin acetate (2b) (0.54 g) afforded 0.06 g of 11 (12%), 0.31 g of 13 (57%), 0.02 g of 15 (4%), mg of (1%), and 16 mg of 18 (3%) 2.4 Acetolysis of 12-oxosapogenin acetate catalyzed by ZnCl2 To a solution of 0.50 g (1.06 mmol) of the 12-oxosapogenin acetate 1b or 2b in 11.60 mL (105 mmol) of acetic anhydride, were added 0.29 g (2.12 mmol) of anhydrous ZnCl2 , then the mixture was stirred at room temperature for 40 h Once finished, the reaction was poured over ground ice (20 g) and vigorously shaken The organic phase was extracted with AcOEt, washed with brine (2 × 20 mL), followed by neutralization with saturated solution of NaHCO3 , dried over anhydrous Na2 SO4 then concentrated until dryness under vacuum The products from the reaction with botogenin acetate (1b) (0.68 g) were isolated by column chromatography over silica gel (230–400 mesh) using hexane/AcOEt (85:15) to give 0.55 g of 16 (81%), 0.10 g of 17 (10%) and 0.03 g of 19 (5%) The crude reaction product of hecogenin acetate (2b) (0.54 g), was separated by column chromatography over silica gel (230–400 mesh) affording 0.36 g of (65%), 0.14 g of 18 (25%), and 0.05 g of 20 (10%) 2.5 Acid-catalyzed cyclization of 26-hydroxypentacyclic derivatives (12–13) into hexacyclic frameworks (10–11) In a mL round bottom flask were dissolved 0.10 g (0.19 mmol) of the corresponding 26-hydroxypentacyclic derivative (12 or 13) in mL of dichloromethane and 5.0 ␮L of glacial AcOH were added This mixture was magnetically stirred at 25 ◦ C and monitored by TLC for h The reaction mixture was poured over ground ice (3 g) and vigorously shaken The organic phase was extracted with AcOEt and washed with brine (2 × mL), followed by neutralization with saturated NaHCO3 solution, dried over Na2 SO4 and concentrated to dryness under vacuum The reaction products were isolated by column chromatography over silica gel (230–400 mesh) using hexane/AcOEt (9:1) to give 0.093 g of 10 (97%) and 0.082 g of 11 (87%), respectively 2.6 Base-catalyzed cyclization of 26-hydroxypentacyclic derivative (13) into hexacyclic framework (11) In a 10 mL round bottom flask were dissolved 0.10 g (0.19 mmol) of 13 in 5.0 mL of dichloromethane and 8.5 mg de DMAP were 1129 added The mixture was magnetically stirred at 25 ◦ C and monitored by TLC for h The reaction mixture was poured over ground ice (5 g) and vigorously shaken The organic phase was extracted with AcOEt, washed with 5% HCl solution (1 × 10 mL), brine (2 × 10 mL), followed by neutralization with saturated solution of NaHCO3 , dried over anhydrous Na2 SO4 and concentrated to dryness under vacuum The reaction product was isolated by column chromatography performed over silica gel (230–400 mesh) using hexane/AcOEt (9:1) to give 0.087 g of 11 (92%) 2.6.1 (20S, 25R)-12,23-cyclo-22,26-epoxycholesta-5, 11,22-triene-3ˇ,16ˇ-diyl diacetate (10) ◦ Oil, [˛]25 ¯ max : 2928, 1732, 1682, D −81.5 (c 1.6, CH2 Cl2 ); IR v −1 1634, 1244 cm ; MS, m/z (%): 494 (M+ 10), 419 (6), 314 (100), 254 (6), 239 (97), 184 (4) HRMS calcd m/z for C31 H42 O5 (M+ +1): 495.3105; found: 495.3112 H NMR, ı: 5.38 (1H, d, J = 4.7 Hz, H-6), 5.28 (1H, ddd, J16␣-17␣ = 8.0 Hz, J16␣-15␣ = 7.0 Hz, J16␣-15␤ = 4.0 Hz, H-16␣ ), 4.98 (1H, s, H-11), 4.57 (1H, m, H-3␣ ), 3.94 (1H, dd, Jgem = 10.2 Hz, J26eq-25ax = 3.2 Hz, H-26eq ), 3.29 (1H, dd, Jgem = J26ax-25ax = 10.2 Hz, H-26ax ), 2.46 (1H, m, H-20), 2.38 (1H, ddd, Jgem = 13.0 Hz, J15␣-14␣ = 7.0 Hz, J15␣-16␣ = 7.0 Hz, H-15␣ ), 2.31 (1H, dd, Jgem = 13.3 Hz, J4eq-3ax = 4.6 Hz, H-4eq ), 1.97 and 1.98 (3H each, s, 3-, 16-OCOCH3 ), 1.40 (1H, ddd, Jgem = 13.0 Hz, J15␤-14␣ = 13.0 Hz, J15␤-16␣ = 4.0 Hz, H-15␤ ), 1.02 (3H, d, J21-20 = 6.6 Hz, CH3 -21), 0.97 (3H, s, CH3 -19), 0.92 (3H, d, J27-25 = 6.6 Hz, CH3 -27), 0.86 (3H, s, CH3 -18) 13 C NMR ı: 37.0 (C-1), 27.7 (C-2), 74.1 (C-3), 38.3 (C-4), 140.9 (C-5), 122.8 (C-6), 29.0 (C-7), 31.1 (C-8), 55.7 (C-9), 38.3 (C10), 110.0 (C-11), 144.9 (C-12), 41.4 (C-13), 50.5 (C-14), 35.6 (C-15), 73.9 (C-16), 55.1 (C-17), 17.5 (C-18), 19.6 (C-19), 31.1(C-20), 15.6 (C-21), 154.6 (C-22), 104.6 (C-23), 31.4 (C-24), 27.4 (C-25), 71.6 (C26), 17.2 (C-27), 170.6 and 170.5 (3-, 16-OCOCH3 ), 21.4 and 21.1 (3-,16-OCOCH3 ) 2.6.2 (20S,25R)-12,23-cyclo-22,26-epoxy-5˛cholesta-11,22-diene-3ˇ,16ˇ-diyl diacetate (11) Oil, [˛]25 −27.1◦ (c 0.2, CHCl3 ); IR v¯ max : 2958, 1735, D 1244 cm−1 ; MS, m/z (%): 496 (M+ , 100), 437 (37), 377 (8), 327 (55), 57 (12), 43 (16); HRMS calcd m/z for C31 H44 O5 (M+ + 1): 496.3189; found 496.3194 H NMR ı: 5.31 (1H, ddd, J16␣-17␣ = 7.0 Hz, J16␣-15␣ = 7.0 Hz, J16␣-15␤ = 3.3 Hz, H16␣ ), 5.05 (1H, d, J = 2.0 Hz, H-11), 4.71 (1H, m, H-3␣ ), 3.99 (1H, dd Jgem = 10.4 Hz, J26eq-25ax = 3.2 Hz, H-26eq ), 3.34 (1H, dd, Jgem = J26ax-25ax = 10.4 Hz, H-26ax ), 2.53 (1H, m, H-20), 2.42 (1H, ddd, Jgem = 13.8 Hz, J15␣-14␣ = 7.0 Hz, J15␣-16␣ = 7.0 Hz, H-15␣ ), 2.29 (1H, dd, Jgem = 16.0 Hz, J4eq-3ax = 5.6 Hz, H-4eq ), 2.02 and 2.01 (3H each, s, 3-, 16-OCOCH3 ), 1.06 (3H, d, J21-20 = 6.4 Hz, CH3 -21), 0.96 (3H, d, J27-25 = 6.8 Hz, CH3 -27), 0.90 (3H, s, CH3 -18), 0.83 (3H, s, CH3 -19); 13 C NMR ı: 36.4 (C-1), 27.5 (C-2), 73.6 (C-3), 34.1 (C-4), 44.8 (C-5), 31.2 (C-6), 29.4 (C-7), 34.0 (C-8), 58.9 (C-9), 36.8 (C-10), 110.0 (C-11), 144.4 (C-12), 41.8 (C-13), 50.7 (C-14), 35.5 (C-15), 73.9 (C-16), 55.1 (C-17), 18.1 (C-18), 13.2 (C-19), 31.2 (C-20), 15.8 (C-21), 154.1 (C-22), 104.6 (C-23), 29.0 (C-24), 27.4 (C-25), 71.5 (C-26), 17.3 (C-27), 170.4 and 170.3 (3-, 16-OCOCH3 ), 21.5 and 21.2 (3-, 16-OCOCH3 ) 2.6.3 (20S,25R)-12,23-cyclo-26-hydroxy-22oxocholesta-5,12(23)-diene-3ˇ,16ˇ-diyl diacetate (12) ◦ Oil [˛]25 ¯ max : 3426, 2898, 1734, 1654, D +19.4 (c 0.93, CH2 Cl2 ); IR v 1616, 1246 cm−1 MS, m/z (%): 512 (M+ , 10), 494 (59), 434 (66), 419 (78), 359 (94), 332 (49), 314 (100), 239 (83), 197 (13), 145 (8), 43 (5) HRMS calcd for C31 H45 O6 (M+ + 1): 513.3208; found: 513.3211 H NMR (Py-d5 ) ı: 5.36 (2H, m, H-6 and H-16), 4.75 (1H, m, H-3␣ ), 3.83 (1H, dd, Jgem = 10.4 Hz, J26a-25 = 5.6 Hz, H-26a ), 3.76 (1H, dd, Jgem = 10.4 Hz, J26b-25 = 5.6 Hz, H-26b ), 2.93 (1H, dd, Jgem = 13.0 Hz, J7eq-6 = 5.0 Hz, H-7eq ), 2.83 (1H, dd, Jgem = 16.5 Hz, J11eq-9ax = 5.2 Hz, 1130 J.O.H Pérez-Díaz et al / Steroids 75 (2010) 1127–1136 H-11eq ), 2.67 (1H, dq, J20-21 = J20-17␣ = 7.0 Hz, H-20), 2.44 (1H, ddd, Jgem = 13.5 Hz, J15␣-14␣ = 7.0 Hz, J15␣-16␣ = 7.0 Hz, H-15␣ ), 2.29 (1H, dd, Jgem = 16.5 Hz, J11ax-9ax = 12.6 Hz, H-11ax ), 2.07 and 2.04 (3H each, s, 3-, 16-OCOCH3 ), 1.29 (3H, d, J21-20 = 7.0 Hz, CH3 -21), 1.13 (3H, d, J27-25 = 6.8 Hz, CH3 -27), 1.06 (3H, s, CH3 -19),1.05 (3H, s, CH3 -18) H NMR (CDCl3 ) ı: 5.40 (1H, d, J = 5.0 Hz, H-6), 5.25 (1H, ddd, J16␣-17␣ = J16␣-15␣ = 7.0 Hz, J16␣-15␤ = 3.2 Hz, H-16␣ ), 4.60 (1H, m, H-3␣ ), 3.26 (1H, dd, Jgem = 10.4 Hz, J26a-25 = 5.6 Hz, H-26a ), 3.18 (1H, dd, Jgem = 10.4, J26b-25 = 5.6 Hz, H-26b ), 2.68 (1H, dd, Jgem = 16.5 Hz, J11eq-9ax = 5.2 Hz, H-11eq ), 2.60 (1H, q, J20-21 = 6.6 Hz, H-20), 2.54 (1H, ddd, Jgem = 13.5 Hz, J15␣-16␣ = J15␣-14␣ = 7.0 Hz, H15␣ ), 2.36 (1H, dd, Jgem = 13.5 Hz, J4eq-3ax = 4.7 Hz, 4eq ), 1.46 (1H, ddd, Jgem = J15␤-14␣ = 13.5 Hz, J15␤-16␣ = 3.2 Hz, H-15␤ ), 2.04 and 2.03 (3H each, s, 3-, 16-OCOCH3 ), 1.14 (3H, d, J21-20 = 6.6 Hz, CH3 -21), 1.12 (3H, s, CH3 -18), 1.07 (3H, s, CH3 -19), 0.88 (3H, d, J27-25 = 6.8 Hz, CH3 -27) 13 C NMR (Py-d5 ) ı: 36.8 (C-1), 27.9 (C-2), 73.4 (C-3), 38.4 (C-4), 139.9 (C-5), 122.2 (C-6), 28.5 (C-7), 29.9 (C-8), 51.6 (C-9), 37.2 (C-10), 25.2 (C-11), 164.0 (C-12), 43.1 (C-13), 54.1 (C-14), 35.0 (C15), 73.6 (C-16), 55.5 (C-17), 15.3 (C-18), 19.1 (C-19), 39.1 (C-20), 13.5 (C-21), 201.3 (C-22), 131.3 (C-23), 31.3 (C-24), 37.0 (C-25), 67.4 (C-26), 17.1 (C-27), 170.1, 170.0 (3-, 16-OCOCH3 ), 21.1, 20.8 (3-, 16-OCOCH3 ) 2.6.4 (20S,25R)-12,23-cyclo-26-hydroxy-22-oxo-5˛-cholesta12(23)-ene-3ˇ,16ˇ-diyl diacetate (13) Oil, [˛]25 +59.4◦ (c 0.11, CHCl3 ); IR v¯ max : 3475, 1732, D 1660, 1244 cm−1 ; MS, m/z (%): 514 (M+ , 5), 496 (100), 453 (10), 437 (11), 55 (30), 43 (65); HRMS calcd for C31 H47 O6 (M+ + 1) 515.3367, found 515.3382 H NMR ı: 5.23 (1H, ddd, J16␣-17␣ = 7.0 Hz, J16␣-15␣ = 7.0 Hz, J16␣-15␤ = 3.2 Hz, H-16␣ ), 4.67 (1H, m, H-3␣ ), 3.36 (1H, dd, Jgem = 10.5 Hz, J26a-25 = 5.7 Hz, H26a ), 3.29 (1H, dd, Jgem = 10.5 Hz, J26b-25 = 5.7 Hz, H-26b ), 2.64 (1H, dd, Jgem = 16.0 Hz, J11eq-9ax = 4.8 Hz, H-11eq ), 2.60 (1H, dq, J20-17␣ = 14.0 Hz, J20-21 = 6.9 Hz, H-20), 2.51 (1H, ddd, Jgem = 14.2 Hz, J15␣-16␣ = J15␣-14␣ = 7.0 Hz, H-15␣ ), 2.10 (1H, dd, Jgem = 16.0 Hz, J11ax-9ax = 13.0 Hz, H-11ax ), 1.96 and 1.95 (3H each, s, 3-, 16OCOCH3 ), 1.06 (3H, d, J21-20 = 6.9 Hz, CH3 -21), 0.99 (3H, s, CH3 -18), 0.86 (3H, s, CH3 -19), 0.81 (3H, d, J27-25 = 6.6 Hz, CH3 -27); 13 C NMR ı: 36.1 (C-1), 27.3 (C-2), 73.1 (C-3), 33.9 (C-4), 44.7 (C-5), 31.4 (C-6), 28.0 (C-7), 33.7 (C-8), 53.9 (C-9), 36.7 (C-10), 25.4 (C-11), 165.0 (C-12), 43.4 (C-13), 55.3 (C-14), 34.8 (C-15), 73.3 (C-16), 55.4 (C-17), 15.8 (C-18), 12.2 (C-19), 39.0 (C-20), 13.3 (C-21), 202.6 (C-22), 130.8 (C-23), 27.4 (C-24), 36.4 (C-25), 66.4 (C-26), 17.1 (C-27), 170.3 and 170.1 (3-, 16-OCOCH3 ), 21.4 and 21.1 (3-, 16-OCOCH3 ) 2.6.5 (20S, 25R)-12,23-cyclo-22-oxocholesta-5,12(23)-diene-3ˇ,16ˇ, 26-triyl triacetate (14) Oil, [˛]25 +54.6◦ (c 0.2, CHCl3 ); IR max : 2927, 2858, D 1733, 1657, 1615, 1235 cm−1 ; MS, m/z (%): 554 (M+ , 1), 494 (90), 434 (100), 419 (46), 359 (57), 314 (66); HRMS calcd For C31 H47 O6 (M+ -60) 494.3032, found 494.3025 H NMR ı: 5.41 (1H, d, J = 5.0 Hz, H-6), 5.25 (1H, ddd, J16␣-15␣ = 10.6 Hz, J16␣-17␣ = 7.0 Hz, J16␣-15␤ = 3.7 Hz, H-16␣ ), 4.61 (1H, m, H-3␣ ), 3.93 (1H, dd, Jgem = 10.5 Hz, J26a-25 = 5.9 Hz, H-26a ), 3.90 (1H, dd, Jgem = 10.5 Hz, J26b-25 = 5.9 Hz, H-26b ), 2.62 (1H, dd, Jgem = 13.5 Hz, J11eq-9ax = 6.0 Hz, H-11eq ), 2.53 (1H, q, J20-21 = 7.0 Hz, H-20), 2.47 (1H, dd, Jgem = 14.0 Hz, J15␣-14␣ = 7.0 Hz, H-15␣ ), 2.39 (1H, dd, Jgem = 13.5 Hz, J4eq-3ax = 6.0 Hz, H-4eq ), 2.24 (1H, dd, Jgem = 13.5 Hz, J11ax-9ax = 7.9 Hz, H-11ax ), 2.06, 2.05 and 2.04 (3H each, s, 3-, 16-, 26-OCOCH3 ), 1.46 (1H, ddd, Jgem = 14.0 Hz, J15␤-14␣ = 11.6 Hz, J15␤-16␣ = 3.7 Hz, H-15␤ ), 1.13 (3H, d, J21-20 = 7.0 Hz, CH3 -21), 1.11 (3H, s, CH3 -19), 1.08 (3H, s, CH3 -18), 0.83 (3H, d, J27-25 = 6.9 Hz, CH3 -27) 13 C NMR ı: 36.9 (C-1), 27.7 (C-2), 73.5 (C-3), 38.1 (C4), 139.6 (C-5), 122.1 (C-6), 28.4 (C-7), 31.2 (C-8), 51.5 (C-9), 37.1 (C-10), 25.0 (C-11), 164.5 (C-12), 43.2 (C-13), 54.2 (C-14), 35.0 (C-15), 73.5 (C-16), 55.6 (C-17), 15.6 (C-18), 19.3 (C-19), 39.0 (C-20), 13.2 (C-21), 201.6 (C-22), 130.6 (C-23), 30.0 (C-24), 32.8 (C-25), 69.0 (C-26), 16.7 (C-27), 171.3 and 170.5 (3-, 16-, 26OCOCH3 two signals are overlapped), 21.4, 21.2 and 21.0 (3-, 16-, 26- OCOCH3 ) 2.6.6 (20S,25R)-12,23-cyclo-22-oxo5˛-cholesta-12(23)-ene-3ˇ,16ˇ, 26-triyl triacetate (15) ◦ Oil, [˛]25 ¯ max : 1737, 1660, 1244 cm−1 ; D +54.2 (c 0.2, CHCl3 ); IR v + MS, m/z (%): 556 (M , 1), 496 (100), 438 (27), 421 (51), 56 (39), 43 (63); H NMR ı: 5.25 (1H, ddd, J16␣-15␣ = J16␣-17␣ = 7.0 Hz, J16␣-15␤ = 3.0 Hz, H-16␣ ), 4.66 (1H, m, H-3␣ ), 3.86 (1H, dd, Jgem = 10.4 Hz, J26a-25 = 6.0 Hz, H-26a ), 3.84 (1H, dd, Jgem = 10.4, J26b-25 = 6.0, H-26b ), 2.62 (1H, dd, Jgem = 16.0 Hz, J11eq-9ax = 5.0 Hz, H-11eq ), 2.54 (1H, dq, J20-17␣ = 12.2 Hz, J20-21 = 6.8 Hz, H-20), 2.51 (1H, ddd, Jgem = 14.1 Hz, J15␣-16␣ = J15␣-14␣ = 7.0 Hz H-15␣ ), 2.03, 2.01 and 2.00 (3H each, s, 3-, 16-, 26-OCOCH3 ), 1.04 (3H, s, CH3 -18), 1.10 (3H, d, J21−20 = 6.8 Hz, CH3 -21), 0.91 (3H, s, CH3 -19), 0.81 (3H, d, J27-25 = 7.2 Hz, CH3 -27); 13 C NMR ı: 36.1 (C-1), 27.3 (C-2), 73.1 (C-3), 33.9 (C-4), 44.7 (C-5), 31.4 (C-6), 28.0 (C-7), 33.7 (C-8), 54.0 (C-9), 36.5 (C-10), 25.3 (C-11), 164.4 (C-12), 43.4 (C-13), 55.2 (C14), 34.9 (C-15), 73.4 (C-16), 55.4 (C-17), 15.8 (C-18), 12.2 (C-19), 39.0 (C-20), 13.2 (C-21), 201.7 (C-22), 130.5 (C-23), 28.2 (C-24), 32.7 (C-25), 69.0 (C-26), 16.6 (C-27), 171.2, 170.6 and 170.5 (3-, 16-, 26- OCOCH3 ), 21.4, 21.1 and 20.9 (3-, 16-, 26-OCOCH3 ); Anal calcd for C33 H48 O7 : C 71.19, H 8.69, O 20.12 Found: C 71.19, H 8.98 2.6.7 (25R)-23-acetyl-22,26-epoxy-12-oxocholesta5,22-diene-3ˇ,16ˇ-diyl diacetate (16) Colorless crystals, m.p 111–113 ◦ C (hexane/ethyl acetate) [˛]25 D +30.9◦ (c 1.0, CH2 Cl2 ); IR v¯ max : 1733, 1720, 1664, 1570, 1372, 1246 cm−1 MS, m/z (%): 554 (M+ , 7), 479 (32), 451 (17), 206 (100), 205 (54), 179 (53), 163 (45), 116 (19), 43 (8) HRMS calcd m/z for C33 H47 O7 (M+ +1): 555.3320; found: 555.3316 H NMR, ı: 5.39 (1H, d, J = 5.8 Hz, H-6), 5.16 (1H, ddd, J16␣-17␣ = J16␣-15␣ = 7.8 Hz, J16␣-15␤ = 3.5 Hz, H-16␣ ), 4.58 (1H, m, H-3␣ ), 4.04 (1H, dd, Jgem = 10.3 Hz, J26eq-25ax = 3.2 Hz, H-26eq ), 3.96 (1H, dq, J20-17␣ = 11.2, J20-21 = 7.1 Hz, H-20), 3.45 (1H, dd, Jgem = J26ax-25ax = 10.3 Hz, H26ax ), 2.77 (1H, dd, J17␣-16␣ = 7.8 Hz, J17␣-20 = 11.2 Hz, H-17␣ ), 2.71 (1H, dd, Jgem = J11ax-9ax = 12.8 Hz, H-11ax ), 2.21 (1H, dd, Jgem = 12.8 Hz, J11eq-9ax = 8.2 Hz, H-11eq ), 2.17 (3H, s, 23-COCH3 ), 2.03 (3H, s, 3-OCOCH3 ), 1.85 (3H, s, 16-OCOCH3 ), 1.71 (1H, ddd, Jgem = 13.5 Hz, J1eq-2eq = J1eq-2ax = 3.5 Hz, H-1eq ), 1.49 (1H, ddd, J9ax-11ax = 12.8 Hz, J9ax-11eq = 8.2 Hz, J9ax-8ax = 11.3 Hz, H-9ax ) 1.26 (3H, s, CH3 -19), 1.17 (1H, dd, Jgem = 13.5 Hz, J1ax-2eq = 3.5 Hz, H-1ax ), 1.14 (3H, s, CH3 -18), 1.09 (3H, d, J21-20 = 7.1 Hz, CH3 -21), 0.97 (3H, d, J27-25 = 6.2 Hz, CH3 -27) 13 C NMR, ı: 36.7 (C-1), 27.6 (C-2), 73.5 (C-3), 37.9 (C-4), 139.7 (C-5), 122.1 (C-6), 31.3 (C-7), 31.7 (C-8), 53.9 (C-9), 37.6 (C-10), 38.0 (C-11), 213.4 (C-12), 56.8 (C-13), 55.9 (C-14), 34.8 (C-15), 73.9 (C-16), 46.9 (C-17), 12.6 (C-18), 19.1 (C-19), 33.0 (C-20), 19.4 (C-21), 171.2 (C-22), 106.9 (C-23), 31.9 (C-24), 26.6 (C-25), 71.7 (C-26), 16.9 (C-27), 198.0 (23-COCH3 ), 170.4 (3-, 16OCOCH3 , two signals overlapped), 29.6 (23-COCH3 ), 21.4 and 21.1 (3-, 16-OCOCH3 ) Anal calcd for C33 H46 O7 : C 71.45, H 8.36, O 20.19 Found: 71.49, H 8.60 2.6.8 (25R)-23-acetyl-22,26-epoxy-12-oxo-5˛-cholesta-22-ene3ˇ,16ˇ-diyl diacetate (5) ◦ Colorless crystals m.p 195 ◦ C (MeOH), [˛]25 D + 37.6 (c 0.62, CHCl3 ); IR¯␯max : 1732, 1708, 1665 cm−1 MS, m/z (%): 556 (M+ ) H NMR ı: 5.13 (1H, ddd, J16␣-17␣ = J16␣-15␣ = 8.2 Hz, J16␣-15␤ = 4.1 Hz, J.O.H Pérez-Díaz et al / Steroids 75 (2010) 1127–1136 H-16␣ ), 4.66 (1H, m, H-3␣ ), 4.03 (1H, dd, Jgem = 10.4 Hz, J26eq-25ax = 3.6 Hz, H-26eq ), 3.98 (1H, m, H-20), 3.45 (1H, dd, Jgem = J26ax-25ax = 10.4 Hz, H-26ax ), 2.75 (1H, dd, J17␣-16␣ = 8.2 Hz, J17␣-20 = 11.2, H-17␣ ), 2.59 (1H, dd, Jgem = J11ax-9ax = 12.8 Hz, H11ax ), 2.17 (3H, s, 23-COCH3 ), 2.01 (3H, s, 3-OCOCH3 ), 1.84 (3H, s, 16-OCOCH3 ), 1.23 (3H, s, CH3 -18), 1.08 (3H, d, J20-21 = 7.0 Hz, CH3 -21), 0.97 (3H, d, J27-25 = 6.8 Hz, CH3 -27), 0.93 (3H, s, CH3 -19) 13 C NMR ı: 36.3 (C-1), 27.2 (C-2), 73.1 (C-3), 33.7 (C-4), 44.4 (C5), 28.3 (C-6), 31.8 (C-7), 35.0 (C-8), 57.1 (C-9), 36.3 (C-10), 38.3 (C-11), 213.2 (C-12), 57.0 (C-13), 55.5 (C-14), 34.7 (C-15), 73.8 (C16), 46.8 (C-17), 12.7 (C-18), 11.9 (C-19), 33.0 (C-20), 19.4 (C-21), 171.0 (C-22), 106.6 (C-23), 31.1 (C-24), 26.5 (C-25), 71.5 (C-26), 16.9 (C-27), 197.7 (23-COCH3 ), 170.3 and 170.1 (3-, 16-OCOCH3 ), 29.6 (23-COCH3 ), 21.4 and 21.1 (3-, 16-OCOCH3 ) Anal calcd for C33 H48 O7 : C 71.19, H 8.69, O 20-12 Found C 71.03, H 8.69 2.6.9 (E)-(20S, 25R)-20,23-diacetyl-12-oxofurosta5,22-diene-3ˇ,26-diyl diacetate (17) ◦ Oil, [˛]25 ¯ max : 2928, 1732, 1708, 1667, D −5.1 (c 2.7, CH2 Cl2 ); IR v 1374, 1243, 1034 cm−1 MS, m/z (%): 596 (M+ , 7), 554 (24), 494, (19), 451 (28), 391 (27), 266 (42), 223 (100), 205 (84), 163 (44), 121 (16), 43 (23) HRMS calcd for C35 H48 O8 (M+ + 1): 597.3421; found: 597.3437 H NMR ı: 5.42 (1H, d, J = 5.0 Hz, H-6), 4.70 (1H, ddd, J16␣-15␣ = J16␣-17␣ = 6.9 Hz, J16␣-15␤ = 4.0 Hz, H-16␣ ), 4.53 (1H, m, H3␣ ), 3.95 (2H, d, Jgem = 6.1 Hz, H-26), 2.76 (1H, d, J17␣-16␣ = 6.9 Hz, H-17␣ ), 2.55 (1H, dd, Jgem = J11ax-9ax = 13.3 Hz, H-11ax ), 2.45 (3H, s, 20-COCH3 ), 2.13 (3H, s, 23-COCH3 ), 2.01 and 2.00 (3H each, s, 3-, 26-OCOCH3 ), 1.72 (1H, dd, Jgem = 13.2 Hz, J1eq-2eq = 3.4 Hz, H-1eq ), 1.56 (3H, s, CH3 -21), 1.18 (3H, s, CH3 -18), 1.10 (3H, s, CH3 -19), 0.92 (3H, d, J27-25 = 6.8 Hz, CH3 -27) 13 C NMR ı: 36.6 (C-1), 27.5 (C-2), 73.4 (C-3), 37.9 (C-4), 139.6 (C-5), 121.8 (C-6), 31.1 (C7), 30.8 (C-8), 52.6 (C-9), 37.4 (C-10), 37.2 (C-11), 212.6 (C-12), 56.2 (C-13), 57.4 (C-14), 31.7 (C-15), 83.4 (C-16), 56.4 (C-17), 15.1 (C-18), 19.0 (C-19), 61.8 (C-20), 17.0 (C-21), 174.6 (C-22), 109.6 (C-23), 31.9 (C-24), 33.3 (C-25), 68.8 (C-26), 17.3 (C-27), 206.9 (20-COCH3 ), 199.0 (23-COCH3 ), 170.5 and 170.3 (3-, 26OCOCH3 ), 28.7 (23-COCH3 ), 26.1 (20-COCH3 ), 21.4 and 21.0 (3-, 26-OCOCH3 ) 2.7 (E)-(20S, 25R)-20,23-diacetyl-12-oxo-5˛-furosta22-ene-3ˇ, 26-diyl diacetate (18) ◦ Colorless crystals m.p 172–174 ◦ C, [˛]25 D −10.7 (c 1.6, CHCl3 ); IR v¯ max : 1735, 1707, 1667, 1362 cm−1 MS, m/z (%): 598(M+ , 23), 556 (52), 496 (33), 453 (71), 43 (100) H NMR ı: 4.68 (2H, m, H-3, H16␣ ), 3.97 (2H, m, H-26), 2.74 (1H, d, J17␣-16␣ = 7.0 Hz, H-17␣ ), 2.46 (3H, s, 20-COCH3 ), 2.17 (3H, s, 23-COCH3 ), 2.05 and 2.03 (3H each, s, 3-, 26-OCOCH3 ), 1.57 (3H, s, CH3 -21), 1.17 (3H, s, CH3 -18), 0.94 (3H, d, J27-25 = 7.0 Hz, CH3 -27), 0.92 (3H, s, CH3 -19) 13 C NMR ı: 36.2 (C-1), 27.0 (C-2), 72.9 (C-3), 33.5 (C-4), 44.2 (C-5), 27.9 (C-6), 30.8 (C7), 33.0 (C-8), 57.0 (C-9), 36.0 (C-10), 37.4 (C-11), 212.4 (C-12), 56.3 (C-13), 55.7 (C-14), 31.3 (C-15), 83.2 (C-16), 56.3 (C-17), 14.9 (C18), 11.7 (C-19), 61.6 (C-20), 16.8 (C-21), 174.5 (C-22), 109.3 (C-23), 31.7 (C-24), 34.1 (C-25), 68.6 (C-26), 17.1 (C-27), 206.7 (20-COCH3 ), 198.7 (C, 23-COCH3 ), 171.0 and 170.5 (3-, 26- OCOCH3 ); 28.5 (CH3 , 23-COCH3 ), 25.9 (20-COCH3 ), 21.2 and 20.8 (3-, 26-OCOCH3 ) Anal calcd for C35 H50 O8 , C 69.55, H 8.58, O 21.61 Found C 70.07, H 8.16 2.8 (E)-(25R)-23-acetyl-12-oxofurosta-5,22-diene-3ˇ, 26-diyl diacetate (19) ◦ Oil, [˛]25 ¯ max : 2856, 1732, 1706, 1656, D +36.7 (c 1.2, CH2 Cl2 ) IR v 1240, 1030 cm−1 MS, m/z (%): 554 (M+ , 16), 494 (25), 451 (27), 391 (28), 223 (58), 205 (100), 206 (47), 163 (52), 97 (24), 71(25), 60 (26), 43 (34) HRMS calcd for C33 H46 O7 (M+ +1): 555.3316; 1131 found: 555.3341 H NMR ı: 5.40 (1H, d, J = 5.0 Hz, H-6), 4.91 (1H, ddd, J16␣-15␣ = J16␣−17␣ = 7.2 Hz, J16␣-15␤ = 4.2 Hz, H-16␣ ), 4.57 (1H, m, H-3␣ ), 3.92 (1H, dd, Jgem = 11.5 Hz, J26a-25 = 5.0 Hz, H-26a ), 3.86 (1H, dd, Jgem = 11.5 Hz, J26b-25 = 5.0 Hz, H-26b ), 3.74 (1H, q, J20-21 = 7.1 Hz, H-20), 2.63 (1H, d, J17␣-16␣ = 7.2, H-17␣ ), 2.55 (1H, dd, Jgem = 13.0 Hz, J11ax-9ax = 8.0 Hz, H-11ax ), 2.24 (1H, dd, Jgem = 13.0 Hz, J11eq-9ax = 6.3 Hz, H-11eq ), 2.19 (3H, s, 23-COCH3 ), 2.04 and 2.02 (3H each, s, 3-, 26-OCOCH3 ), 1.24 (3H, d, J21-20 = 7.1 Hz, CH3 -21), 1.10 (3H, s, CH3 -19), 0.94 (3H, s, CH3 -18), 0.93 (3H, d, J27-25 = 7.1 Hz, CH3 27) 13 C NMR ı: 36.7 (C-1), 27.6 (C-2), 73.4 (C-3), 38.0 (C-4), 139.8 (C-5), 122.1 (C-6), 31.3 (C-7), 30.8 (C-8), 52.2 (C-9), 37.4 (C-10), 36.9 (C-11), 212.0 (C-12), 55.6 (C-13), 54.9 (C-14), 33.2 (C-15), 84.3 (C16), 53.9 (C-17), 13.4 (C-18), 19.0 (C-19), 39.0 (C-20), 19.4 (C-21), 178.0 (C-22), 108.1 (C-23), 31.6 (C-24), 33.3 (C-25), 69.0 (C-26), 17.3 (C-27), 198.0 (23-COCH3 ), 171.3 and 170.6 (3-, 26-OCOCH3 ), 29.3 (23-COCH3 ), 21.5 and 21.1 (3-, 26-OCOCH3 ) 2.9 (E)-(25R)-23-acetyl-12-oxo-5˛-furosat-22-ene-3ˇ, 26-diyl diacetate (20) ◦ Colorless crystals m.p 144–145 ◦ C, [˛]25 D +83.56 (c 1.0, CHCl3 ); IR v¯ max : 1732, 1707, 1665, 1243 cm−1 MS, m/z (%): 556(M+ , 24), 496 (30), 453 (52), 205 (48), 163 (33), 43 (100) H NMR ı: 4.84 (1H, m, H-16␣ ), 4.61 (1H, m, H-3), 3.84 (2H, m, H-26), 3.69 (1H, m, H-20), 2.54 (1H, d, J17␣-16␣ = 7.3 Hz, H-17␣ ), 2.14 (3H, s, 23-COCH3 ), 1.99 and 1.96 (3H each, s, 3-, 26-OCOCH3 ), 1.18 (3H, d, J21-20 = 7.0 Hz, CH3 -21), 0.88 (3H, d, J27-25 = 7.0 Hz, CH3 -27), 0.87 (3H, s, CH3 -19), 0.85 (3H, s, CH3 -18), 13 C NMR ı: 36.3 (C-1), 27.2 (C-2), 73.1 (C-3), 33.8 (C-4), 44.5 (C-5), 28.1 (C-6), 31.3 (C-7), 33.3 (C-8), 55.6 (C9), 36.3 (C-10), 37.3 (C-11), 212.5 (C-12), 55.8 (C-13), 54.7 (C-14), 33.0 (C-15), 84.2 (C-16), 54.0 (C-17), 11.9 (C-18), 13.5 (C-19), 38.9 (C-20), 19.3 (C-21), 177.8 (C-22), 108.1 (C-23), 31.4 (C-24), 34.2 (C25), 68.9 (C-26), 17.3 (C-27), 198.4 (23-COCH3 ), 171.2 and 170.6 (3-, 26-OCOCH3 ); 29.2 (23-COCH3 ), 21.4 and 21.1 (3-, 26-OCOCH3 ) Anal calcd for C33 H48 O7 , C 71.05, H 8.67, O 20.32 Found C 71.22, H 8.63 Results and discussion The one pot BF3 ·OEt2 catalyzed reaction of botogenin (1b) and hecogenin (2b) acetates in acetic anhydride using dichloromethane as solvent proceeded under mild conditions to give hexa- and pentacyclic compounds (10–15) with retention of configuration of all asymmetric centers (Scheme 1) The effect of the concentration of acetic anhydride and the use of CH2 Cl2 on the regioselectivity of the reaction was evaluated The use of strong mineral acids and high temperatures was avoided to retain the stereochemistry of all chiral centers, since previous reports describe that these conditions promote inversion of configuration and epimerization [29] Tables and summarize the product distribution and the reaction conditions for the formation of derivatives 5, 10–20 The results indicate that formation of penta- and hexacyclic compounds is favored under low acetic anhydride concentrations, while conditions similar to those described in the literature direct mainly to the 22,26-epoxy (16 and 5) and furostene (17–20) derivatives, as in entries 2, 3, 5, and [24] The selectivity to obtain the hexacyclic derivatives was not increased even at low temperatures (−30 or ◦ C) The preference for the E ring opening has been documented and attributed to: (a) the higher stability of the tetrahydropyran ring under several acidic reaction conditions [32–34], (b) the use of hard Lewis acids (as compared to the use of softer BBr3 or Ph3 P·I2 where the cleavage is directed toward the F ring) [21,29] and (c) the higher reactivity based on the larger basicity of the tetrahydrofuran oxygen [21,35] 1132 J.O.H Pérez-Díaz et al / Steroids 75 (2010) 1127–1136 Scheme Acetolysis of botogenin (1b) and of hecogenin (2b) acetate Table Product distribution in the spiroketal cleavage of botogenin acetate (1b) at 25 ◦ C Entry Lewis acid (mmol) Ac2 O (mmol) Time (min) 10 (%) 12 (%) 14 (%) 16 (%) 17 (%) 19 (%) 1a BF3 (4.25) BF3 (4.25) ZnCl2 (2.1) 6.36 12.7 105 45 40 40 h 27 22 0 0 73 81 29 10 10 5 a Reaction in mL of CH2 Cl2 To explain the formation of compounds 10–15, we propose a reaction mechanism (Scheme 2) in which the ring E opening is activated by the Lewis acid The first step involves the E ring opening promoted by the BF3 ·OEt2 producing the oxocarbenium ion A The elimination of a proton from position 23, leads to the dihydropyranic intermediate B Rotation of the ␴ bond between C-17 and C-20 allows to attain an adequate disposition of the enol ether to attack the ␲ system of the carbonyl group at C-12, directing to the tertiary alcoholate C The loss of the proton in 23 allows the tetrahydropyranic oxygen atom to recover its electronic pair, to afford D Elimination of the fluoroborate from D provides the oxocarbenium ion E, which is the common intermediate; when the H-11 is eliminated compound 10 or 11 are obtained, while attack by the acetate nucleophile yields 14 or 15 During the work-up, the intermediate E can be attacked by H2 O directing to 12 or 13 The pentacyclic compounds (12 and 13) can undergo acid or base-catalyzed cyclization (Scheme 3), a process which is extremely fast and nearly quantitative When compound 13 is allowed to stand in AcOH/CH2 Cl2 or is treated with catalytic amounts of 4-dimethylaminopyridine, compound 11 is obtained in almost quantitative yields The formation of the epoxycholestanes (5 and 16) has been visualized from the intermediate B, under a mechanism already proposed [24] On the other hand, the generation of furostenes 17–20 implies the regioselective cleavage of ring F [26] 3.1 Spectroscopic analysis The structures of the compounds were unambiguously established using two-dimensional NMR experiments The assignments were accomplished by combined utilization of DEPT, COSY, HETCOR experiments at 400 MHz, as well as comparison with previously reported data on steroidal sapogenins [24–26,36,37] ROESY and long-range connectivity from HMBC experiments were performed at 500 MHz (see Supplementary information) Additional information was obtained from IR, and mass spectroscopy Some characteristic H and 13 C NMR signals are shown in Tables and In H NMR, compounds 10 and 11 show a characteristic signal around ı = 4.99 ( ı 0.1 ppm) for the vinylic proton (H-11), that is confirmed by the signal at 110.0 ppm in the 13 C NMR spectra In the HMBC spectrum H-11 shows a three bond correlation with C13 (41.4 ppm) and C-10 (38.3 ppm) which are quaternary carbons In addition, the HMBC experiment allows the assignment of C-22 from the three bonds correlation with Me-21; C-12 correlates with Me-18 and C-5 with Me-19 The AMX system gives signals at 3.94 ( ı 0.01 ppm) and 3.29 ( ı 0.04 ppm) which correspond to the diastereotopic protons of H-26eq and H-26ax respectively coupled with H-25, with a ı = 0.65 ppm indicative of a cyclic system In 13 C NMR there are carbon signals for C-22, C-23, C-12 and C-11 at (ı 154, 104, 144 and 110 respectively), these chemical shifts agree with the conjugated diene system present in the molecule, as is Table Product distribution in the spiroketal cleavage of hecogenin acetate (2b) at 25 ◦ C Entry Lewis acid (mmol) Ac2 O (mmol) Time (min) 11 (%) 13 (%) 15 (%) (%) 18 (%) 20 (%) 4a BF3 (4.25) BF3 (4.25) ZnCl2 (2.1) 6.35 25.0 50.0 45 60 40 h 12 0 57 0 65 65 10 25 20 10 a Reaction in mL of CH2 Cl2 J.O.H Pérez-Díaz et al / Steroids 75 (2010) 1127–1136 1133 Scheme Reaction pathway for the formation of compounds 10–15 corroborated by the IR spectra that shows two bands in 1682 and 1634 cm−1 Additionally, in H NMR spectra the singlet signal for Me-18 is slightly shielded ( ı 0.14 ppm) due to the proximity of the ␲ system Derivatives 12 and 13 show a distinctive carbonyl signal at 202.2 ( ı 1.0 ppm) in 13 C NMR which corresponds to C-22, also the signals at ı = 164.7 ± 0.7 ppm (C-12) and 131.2 ± 0.1 ppm (C-23) for the ␣,␤-unsaturated system On the other hand the H NMR spectra show a distinctive ABX system for the diastereotopic protons of C-26 around 3.6 ppm, this is characteristic for open chain steroids systems The ROESY experiment allowed establishing the configuration at C-20 as (S), furthermore the scalar coupling between H-20 and H-17 is 7.0 Hz evidences the same orientation for both protons IR spectroscopy proved the presence of the 26-hydroxyl group from the broad band around 3500 cm−1 , two strong bands at 1730 and 1655 cm−1 are distinctive frequencies for the acetate and conjugated carbonyls groups, respectively The NMR spectra of steroidal derivatives 14 and 15 are very similar to analogs 12 and 13, except for the signals for CH3 COO around 2.03, the signals in the 13 C NMR at 201, 130 and 164 are assigned to the ketone and ␣,␤-unsaturated system, respectively Derivatives and 16–20 show distinctive signals for the diastereotopic protons at C-11, which in these compounds are easily observed, due to the deshielding effect on the H-11ax with respect to the H-11eq This difference in chemical shifts can be Scheme Acid- or base-catalyzed cyclization of compounds 12 and 13 1134 J.O.H Pérez-Díaz et al / Steroids 75 (2010) 1127–1136 Table Selected H chemical shifts ı (ppm) of 5, 10–20 in CDCl3 Compd H-16 H-26eq 10 11 12a 13 14 15 16 17 18 19 20 5.28 (ddd) 5.31 (m) 5.25 (m) 5.23 (m) 5.25 (m) 5.25 (ddd) 5.16 (ddd) 5.13 (ddd) 4.70 (ddd) 4.68 (m) 4.91 (ddd) 4.84 (m) 3.94 (ddd) 3.29 (dd) 3.99 (ddd) 3.34 (dd) 3.83 (dd) 26a 3.76 (dd) 26b 3.36 (dd) 26a 3.29 (dd) 26b 3.93 (dd) 26a 3.90 (dd) 26b 3.86 (dd) 26a 3.84 (dd) 26b 4.04 (dd) 3.45 (dd) 4.03 (dd) 3.45(dd) 3.95 (d) 3.97 (m) 3.92 (dd) 26a 3.86 (dd) 26b 3.84 (m) a H-26ax H-11 H-20 H-21 H-27 H-18 H-19 4.98 (s) 5.05 (d) 2.83 (dd) 11eq 2.64 (dd) 11eq 2.62 (dd) 11eq 2.62 (dd) 11eq 2.71 (dd) 11ax 2.59 (dd) 11ax 2.55 (dd) 11ax Overlapped 2.55 (dd) 11ax Overlapped 2.46 (m) 2.53 (m) 2.67 (dq) 2.60 (dq) 2.53 (dq) 2.54 (dq) 3.96 (dq) 3.98 (m) – – 3.74 (q) 3.69 (m) 1.02 (d) 1.06 (d) 1.29 (d) 1.06 (d) 1.13 (d) 1.10 (d) 1.09 (d) 1.08 (d) 1.56 (s) 1.57 (s) 1.24 (d) 1.18 (d) 0.92 (d) 0.96 (d) 1.13 (d) 0.81 (d) 0.83 (d) 0.81 (d) 0.97 (d) 0.97 (d) 0.92 (d) 0.94 (d) 0.93 (d) 0.88 (d) 0.86 (s) 0.90 (s) 1.05 (s) 0.99 (s) 1.08 (s) 1.04 (s) 1.14 (s) 1.23 (s) 1.18 (s) 1.17 (s) 0.94 (s) 0.85 (s) 0.97 (s) 0.83 (s) 1.06 (s) 0.86 (s) 1.11 (s) 0.91 (s) 1.26 (s) 0.93 (s) 1.10 (s) 0.92 (s) 1.10 (s) 0.87 (s) 2.29 (dd) 11ax 2.10 (dd) 11ax 2.24 (dd) 11ax 2.21 (dd) 11eq 2.24 (dd) 11eq Determined in Py-d5 Fig Some ROESY interactions observed in 10, 12, 14 and 17 Table 13 C chemical shifts ı (ppm) for representative signals of 5, 10–20 Carbon C-3 C-11 C-12 C-16 C-17 C-18 C-19 C-20 C-21 C-22 C-23 C-25 C-26 C-27 a Compound 10 11 12a 13 14 15 16 17 18 19 20 74.1 110.0 144.9 73.9 55.1 17.5 19.6 31.1 15.6 154.6 104.6 27.4 71.6 17.2 73.6 110.0 144.4 73.9 55.1 18.1 13.2 31.2 15.8 154.1 104.6 27.4 71.5 17.3 73.4 25.2 164.0 73.6 55.5 15.3 19.1 39.1 13.5 201.3 131.3 37.0 67.4 17.1 73.1 25.4 165.0 73.3 55.4 15.8 12.2 39.0 13.3 202.6 130.8 36.4 66.4 17.1 73.5 25.0 164.5 73.5 55.6 15.6 19.2 39.0 13.2 201.6 130.6 32.8 69.0 16.7 73.1 25.3 164.4 73.4 55.4 15.8 12.2 39.0 13.2 201.7 130.5 32.7 69.0 16.6 73.5 38.0 213.4 73.9 46.9 12.6 19.01 33.0 19.4 171.2 106.9 26.6 71.7 16.9 73.1 38.3 213.2 73.8 46.8 12.7 11.9 33.0 19.4 171.0 106.6 26.5 71.5 16.9 73.4 37.2 212.6 83.4 56.4 15.1 19.0 61.8 17.0 174.6 109.6 33.3 68.8 17.3 72.9 37.4 212.4 83.2 56.3 14.9 11.7 61.6 16.8 174.5 109.3 34.1 68.6 17.1 73.4 36.9 212.0 84.3 53.9 13.4 19.0 39.0 19.4 178.0 108.1 33.3 69.0 17.3 73.1 37.3 212.5 84.2 54.0 11.9 13.5 38.9 19.3 177.8 108.1 34.2 68.9 17.3 Determined in Py-d5 J.O.H Pérez-Díaz et al / Steroids 75 (2010) 1127–1136 1135 Fig X-ray of (25R)-23-acetyl-22,26-epoxy-12-oxocholesta-5,22-diene-3␤,16␤-diyl diacetate (16) Ellipsoids are drawn at the 35% probability level explained by: (a) the carbonyl effect, which deshields the alpha axial protons 0.3 ppm more than the equatorial ones [38–40] and (b) through steric compression (also called van der Waals deshielding) [40–43] Besides, there is a considerable deshielding ( ı 1.2 ppm) of H-17␣ due to the ␥ effect of the ␣,␤-conjugated system [44] In the H NMR spectra of and 16 the distinctive signal of the AMX system for the diastereotopic protons H-26 at 3.75 ± 0.3 ppm is characteristic of a cyclic system in the F ring The 13 C NMR spectra of and 16 confirmed the presence of a carbonyl from the signal at 213 ppm (C-12), the ␣,␤-unsaturated system in the F ring gives signals at 198, 106, 171 ppm, this is corroborated in the IR spectra from the stretching bands for C O in 1733 (acetate) and 1664 cm−1 (conjugated carbonyl) The signals H and 13 C NMR are in accordance with those analogs previously reported [24–26] As expected, the H NMR spectra of furostenes 17–20 show signals at 4.80 ± 0.1 ppm (H-16) where the E ring remains unaffected, as compared to those of compounds and 10–16, this proton appears deshielded (5.22 ppm) due to the acetate group from the E ring cleavage In 13 C NMR the signal at 84 ppm (C-16) provides evidence that the cleavage did not occurred at the E ring, this is in contrast with compounds and 10–16, where the E ring is cleaved and C-16 appears at 74 ppm Particularly in compounds 17 and 18, the H-26 protons give a doublet signal in the region of 3.95 ppm (J = 6.1 Hz), which is a distinctive signal for open side chain steroid systems The signal at 1.56 ppm corresponds to the ␤-Me-21 which shows a singlet signal due the presence of the ␣-acyl substituent in C-20 The H and 13 C NMR spectra of derivatives 17–20 perfectly fit the expected chemical shifts previously reported [24–26] The configuration at C-20 in representative compounds (10, 12, 14 and 17) was established by ROESY experiments which allowed to detect the vicinal interaction between CH3 -18 and CH3 -21 (Fig 3); additionally, in compound 12 a correlation between H-20 and H-17, both placed in the ␣ face was observed, confirming the configuration at C-17 as S, being the same of the starting material An interaction in compound 17 between H-17 and H-16, both with an alpha orientation confirms this configuration at C-17 ROESY experiments also allowed confirming the configuration at C-16 in compounds 10, 12–15 from the correlation between H-16␣ and H-15␣; in addition the large J values of H-16␣, evidences the pseudo-axial orientation as reported previously [45] The J values of the H-16␣ in compound 12 were obtained from the experiment H determined in CDCl3 , because in Py-d5 and in Py-d5 + D2 O the signal is overlapped with H-6 Suitable crystals for X-ray analysis of the (25R)-23-acetyl22,26-epoxy-12-oxocholesta-5,22-diene-3␤,16␤-diyl diacetate (16) were obtained by slow evaporation of a mixture of hexane- ethyl acetate (9:1), crystallizing in the orthorhombic system, space group P 21 21 21 The X-ray crystal structure analysis established that the stereochemistry at C-20 and C-25 is S, and R respectively (Fig 4).The steroidal nucleus in 16 shows that rings A and C adopt a chair conformation The C-5–C-6 (Csp2 -Csp2 ) distance of 1.307(7) A˚ confirms [46] a double bond, therefore ring B has a half-chair conformation because of the rigidity of the olefinic bond at The D ring displays an envelope conformation as well, and the F pyran ring shows a twisted conformation because of the double bond between C-22 and C-23, as can be corroborated with the ˚ besides the Me-27 C-22–C-23 (Csp2 -Csp2 ) distance of 1.359(6) A, shows an equatorial orientation [24–26,47] In conclusion, we have developed a new, efficient and accessible route for the preparation of polycyclic steroidal skeletons via regioselective cleavage of the E ring of 12-oxosteroidal sapogenins under acid conditions in dichloromethane, catalyzed by BF3 etherate at room temperature High concentration of Ac2 O directs the reaction to the epoxycholestene (5,16) and furostene (17–20) derivatives, while the use of low Ac2 O concentration, favors the intramolecular reaction and the new hexa- and pentacyclic steroidal (10–15) derivatives are obtained as major products This reaction is appealing because it involves the regioselective opening of ring E, followed by aldol condensation with the carbonyl at C-12 without the loss of asymmetric centers The polycyclic steroidal derivatives were synthesized in one pot under mild conditions Further investigation is ongoing regarding the application of this method to the synthesis of versatile frameworks, as brassinosteroids analogs Acknowledgments The authors thank CONACYT, VIEP and SEP-PADES (project No 2009-01-09-006-220) for financial support and scholarships to J.O.H.P.D and J.L.V Also we thank V González for the NMR spectra, G Cuéllar for the mass spectra and M.A Leyva for X-ray data collection Appendix A Supplementary data Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.steroids.2010.07.008 References [1] Harvey AL Natural products in drug discovery Drug Discov Today 2008;13:894–901 [2] Pettit GR, Inoue M, Kamano Y, Herald DL, Arm C, Dufresne C, et al Isolation and structure of the powerful cell growth inhibitor cephalostatin J Am Chem Soc 1988;110:2006–7 1136 J.O.H Pérez-Díaz et al / Steroids 75 (2010) 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