The first synthesis of Krempene B

Steroids 77 (2012) 1398–1402

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The first synthesis of Krempene B Li-Qun Shen ⇑, Su-Yu Huang, Yong Tang, Fu-Hou Lei College of Chemistry and Chemical Engineering, Guangxi University for Nationalities, Key Laboratory of Development and Application of Forest Chemicals of Guangxi, Nanning 530006, China

a r t i c l e

i n f o

Article history: Received 22 June 2012 Received in revised form 19 August 2012 Accepted 24 August 2012 Available online 31 August 2012

a b s t r a c t The synthesis of Krempene B, which can be isolated from the marine soft coral Cladiella krempfi, is achieved in 23.9% overall yield from commercially available 3b-acetoxy-5-pregnen-20-one by 11 steps. Key transformations include the dienone–phenol rearrangement of steroids and Wittig reaction. Ó 2012 Elsevier Inc. All rights reserved.

Keywords: Krempene B Rearrangement Witting reaction Synthesis

1. Introduction Krempenes B (1) was isolated in 2006 by Xinping Huang et al. [1], from the marine soft coral Cladiella krempfi. It is a member of marine pregnene family which occur naturally in organisms that dwell in the sea and contains a 20-double bond (Fig. 1) [2]. These marine pregnenes exhibit a number of impressive biological activities, including anticancer activity [3]. Due to their biological activity and wide range of application in psychology and pharmacology [4], we devoted to the synthesis of marine pregnenes and their analogues for use in biological studies. In this paper, we describe in Scheme 1, an efficient procedure the first synthesis Krempenes B (1).

2. Experimental 2.1. General Melting points were determined using a WRS-1B apparatus and were uncorrected. IR spectra of the compound were recorded in the range of 400–4000 cm 1 with a Magna 550 FT-IR spectrometer using KBr pellets. The 1H and 13C NMR spectra were recorded on 600/150 MHz NMR spectrometers (a Bruker AV600 spectrometer), using tetramethylsilane (TMS) as internal standard and CDCl3 as solvent. High resolution mass (HRMS) spectra were obtained in ESI mode on a Finnigan MAT95XP HRMS system (Thermo Electron Corporation). All chemicals were of reagent grade and used as com⇑ Corresponding author. Tel./fax: +86 771 3260558. E-mail address: [email protected] (L.-Q. Shen). 0039-128X/$ - see front matter Ó 2012 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.steroids.2012.08.006

mercially purchased without further purification. All solvents were dried and distilled before use. THF and dioxane were distilled from sodium/benzophenone ketyl; dichloromethane was distilled from calcium hydride; methanol was distilled from Mg/I2; The reactions were monitored by TLC on silica gel 60 F254. Chromatography was conducted by using 200–300 mesh silica gel.

2.2. Synthesis 2.2.1. 3b-Hydroxy-5-androstene-17b-carboxylic acid (3) To a stirred solution of sodium hydroxide (21.00 g, 0.525 mol) in water (360.0 mL) at 5 °C was added bromine (21.50 g, 0.134 mol), followed after 30 min by addition of dioxane (120 mL). The reaction mixture was kept at 0 °C until required. To a stirred solution of 3b-acetoxy-5-pregnen-20-one (14.00 g, 39.1 mmol) in dioxane (534.7 mL) and water (155.5 mL) at 5 °C was added dropwise the pre-cooled hypobromite solution. The reaction mixture was stirred for an additional 3 h at 8 °C and was quenched with 10% aqueous sodium sulfite (50.0 mL). The mixture was then heated under reflux for 15 min, and the solution, while still hot (90 °C), was acidified by the cautious addition of concentrated hydrochloric acid (25 mL). The clear yellow solution was kept at 5 °C for 24 h. The precipitate was collected by suction filtration, washed with water and dried to give 3 as a white solid. Yield: 11.80 g (95%); m.p. 273–274.5 °C (Lit [5]. 274–276 °C). IR (KBr): 3095, 1791, 1721, 1581, 1484, 1457, 1424, 1065, 419 cm 1.1H NMR (600 MHz, CDCl3): d = 11.91 (s, 1H), 5.25 (d, J = 8.4 Hz, 1H), 4.61 (s, 1H), 3.32–3.22 (m, 1H), 2.25 (t, J = 18.0 Hz, 1H), 2.12–2.06 (m, 2H), 1.96–1.90 (m, 3H), 1.78–1.60 (m, 4H), 1.55–1.52 (m, 2H), 1.42–1.32 (s, 3H), 1.27–1.22 (m, 2H), 1.12–1.09 (m, 1H),

L.-Q. Shen et al. / Steroids 77 (2012) 1398–1402

OH

H H

H H

HO

S H H

H

3-methoxy -19-norpregna -1,3,5(10),20-tetraene

Krempene B (1)

O

H

MeO

S

O H

H

O

H Stereonsteroid B

H H

H Krempene A

Fig. 1. Krempenes B and related marine pregnene products.

O

COOH a

AcO

3 COOCH3

OAc

COOCH3

OAll

k

OAll

Krempenes B (1)

12

OH

5

COOCH3 h

CH2OH OAll

9

8

4 COOCH3

O

6

COOCH3 g

c

d O

f OH

HO

e

7

2.2.3. Methyl 3-oxo-androst-4-ene-17b-carboxylate (5) A solution of 4 (8.00 g, 24.0 mmol) in toluene (160 mL) and cyclohexanone (20 mL) was distilled under argon until 10 mL of distillate was collected. Then the stock solution of aluminum isopropoxide (1.47 g, 7.20 mmol) in toluene (10 mL) was added dropwise to the refluxing solution and the mixture was refluxed for 3 h, followed by reduced distillation in vacuum to exhaustively remove the solvent. After cooling to room temperature, a little methanol was added. Filtration and washing with acetone gave 5. Yield: 6.52 g (82%); m.p. 127–129 °C (Lit [6]. 128–130 °C). IR (KBr): 3030, 2947, 1730, 1671, 1435, 1060 cm 1.1H NMR (600 MHz, CDCl3): d = 5.73 (s, 1H), 3.66 (s, 3H), 2.59–2.45 (m, 2H), 2.28–2.20 (m, 2H), 2.15 (m,1H), 2.05–1.96 (m, 2H), 1.81–1.70 (m, 3H), 1.68–1.57 (m, 3H),1.41 (m, 1H), 1.37–1.24 (m, 2H), 1.18 (s, 3H),1.10–1.05 (m, 2H), 0.98–0.91 (m, 2H), 0.69 (s, 3H).13C NMR (150 MHz, CDCl3): d = 199.6, 174.5, 124.1, 118.1, 56.2, 55.2, 54.9, 53.5, 44.0, 42.7, 38.1, 37.8, 35.7, 34.0, 32.3, 31.7, 24.4, 23.6, 20.9, 19.1, 13.5.

b

HO

2

COOCH3

1399

i j

10 CHO

OAll

11

Scheme 1. Reagents and conditions: (a) NaOBr, dioxane, H2O, conc. HC1, 95% (b) H2SO4, MeOH, reflux, 3 h, 86% (c) Al(i-PrO)3, cyclohexanone, toluene, 110 °C, 3 h, 82% (d) DDQ, dioxane, reflux, 48 h, 81.2% (e) acetic anhydride, PTS, r.t., 24 h, 85% (f) NaHCO3, CH3OH, THF, H2O, r.t. 12 h, 97.3% (g) NaH, allyl bromide, DMF, r.t. 15 h, 92% (h) LiAlH4, THF, r.t., 24 h, 86%; (i) DMP, CH2Cl2, r.t., 30 min, 90%; (j) methyltriphenylphosponium bromide, BuLi, THF, 78 °C, then 0 °C, 6 h; 83% (k) 5% Pd/C, PTS, CH3OH, 72 h 90%.

0.98–0.92 (m, 2H). 0.96 (s, 3H), 0.64 (s, 3H). 13C NMR (150 MHz, CDCl3): d = 175.3, 142.3, 120.8, 70.6, 56.1, 55.2, 50.3, 43.7, 42.8, 38.3, 37.7, 37.5, 32.2, 32.1, 31.9, 24.7, 23.9, 21.1, 19.8, 13.7.

2.2.2. Methyl 3b-hydroxy-5-androstene-17b-carboxylate (4) To a stirred solution of 3 (10.00 g, 31.4 mmol) in methanol (100 mL) was added a few drops of concentrated sulfuric acid. The reaction mixture heated under reflux for 3 h and then allowed to cool to room temperature. The crystalline product which formed upon cooling was collected by filtration to provide 4. Yield: 8.98 g (86%); m.p 173–175 °C (Lit [6]. 173–176 °C). IR (KBr): 3462, 2923, 1730, 1437, 1210 cm 1.1H NMR (600 MHz, CDCl3): d = 5.32 (d, J = 6.0 Hz, 1H), 3.65 (s, 3H), 3.53–3.48 (m, 1H), 2.32 (t, J = 9.6 Hz 1H), 2.27–2.26 (m, 1H), 2.24–2.22 (m, 1H), 2.15–2.09 (m,1H), 2.01–1.96 (m, 2H), 1.80–1.70 (m, 4H), 1.65–1.60 (m, 1H), 1.53– 1.33 (m, 5H), 1.27–1.24 (m, 2H), 1.09–1.05 (m, 2H), 0.98 (s, 3H), 0.98–0.95 (m, 1H), 0.65 (s, 3H).13C NMR (150 MHz, CDCl3): d = 174.7, 141.0, 121.5, 71.8, 56.2, 55.3, 51.4, 50.0, 44.1, 42.3, 38.3, 37.4, 36.7, 32.1, 31.9, 31.7, 24.7, 23.7, 21.0, 19.5, 13.4.

2.2.4. Methyl 3-oxo-androst-1,4-diene-17b-carboxylate (6) A mixture of 5 (1.000 g, 3.0 mmol), and DDQ (0.825 g, 3.6 mmol) in dry dioxane (10 mL) was refluxed for 48 h. The suspension was cooled, filtered, and the filtrate washed three times with 1% sodium hydroxide solution, then with water. The solution was dried, concentrated in vacuo. The residue was flash-chromatographed (10–20% ethyl acetate in petroleum ether) to give 6 as a white solid. Yield: 0.807 g (81.2%); m.p. 147.3–149.2 °C. IR (KBr): 3037, 2940, 1723, 1664, 1296 cm 1. 1H NMR (600 MHz, CDCl3): d = 7.05 (d, J = 10.2 Hz, 1H), 6.23 (dd, J = 10.2, 2.4 Hz, 1H), 6.06 (s, 1H), 3.66 (s, 3H), 2.45 (m, 1H), 2.36–2.32 (m, 2H), 2.13 (m, 1H), 2.03 (m, 1H), 1.95 (m, 1H), 1.81 (m, 1H), 1.73 (m, 1H), 1.70–1.61 (m, 3H), 1.36–1.24 (m, 2H), 1.22 (s, 3H), 1.11–1.01 (m, 3H), 0.72 (s, 3H). 13C NMR(150 MHz, CDCl3): d = 186.5, 174.4, 169.1, 155.9, 128.0, 124.0, 55.1, 55.0, 52.4, 51.5, 44.2, 43.7, 38.0, 35.7, 33.6, 32.9, 24.7, 23.6, 22.8, 18.8, 13.7. 2.2.5. 1-Acetoxy-4-methyl-1,3,5(10)-estratriene-17b-carboxylate (7) To a stirred solution of 6 (0.500 g, 1.5 mmol) in acetic anhydride (5 mL) was added p-toluenesulfonic acid (0.107 g, 0.6 mmol). The reaction mixture was stirred at room temperature for 24 h. The solution was hydrolyzed by swirling with water, the precipitate was filtered, dried and recrystallized from ethyl acetate to yield 7. Yield: 0.479 g (85%); m.p. 143.5–145 °C. IR (KBr): 2936, 2874, 1762, 1734, 1602, 1581, 1470, 1442, 1220 cm 1. 1H NMR (600 MHz, CDCl3) d = 7.00 (d, J = 8.4 Hz, 1H), 6.72 (d, J = 7.8 Hz, 1H), 3.68 (s, 3H), 2.70–2.58 (m, 3H), 2.43 (t, J = 9.0 Hz, 1H), 2.34 (t, J = 9.0 Hz, 1H), 2.28 (s, 3H), 2.19 (s, 3H), 2.04 (dt, J = 12.6, 3 Hz, 1H), 1.89–1.83 (m, 2H), 1.77 (m, 1H), 1.55 (m, 1H), 1.49 (ddd, J = 13.2, 12.6, 3.6 Hz, 1H), 1.44–1.19 (m, 5H), 0.71 (s, 3H). 13C NMR(150 MHz, CDCl3) d = 205.1, 156.3, 138.3, 133.9, 129.0, 127.5, 116.7, 109.6, 68.9, 63.3, 55.6, 53.5, 45.8, 45.3, 39.8, 39.4, 29.3, 26.5, 26.3, 24.6, 21.3, 19.3, 14.7. HRMS: [M+] calcd for C23H30O4: 370.2144; found: 370.2138. 2.2.6. Methyl 1-hydroxy-4-methyl-1,3,5(10)-estratriene-17bcarboxylate (8) To a stirred solution of 7 (0.500 g, 1.3 mmol) in methanol (10 mL) and THF (1 mL) was added saturated sodium bicarbonate solution (2.2 mL). The mixture was stirred at room temperature for 12 h under argon. The mixture was acidified with 2% hydrochloric acid and extracted with ethyl acetate (3  10 mL). The solution was dried, concentrated in vacuo. The residue was flashchromatographed (20–40% ethyl acetate in petroleum ether) to give 8 as a white solid. Yield: 0.431 g (97.3%); m.p. 120–122 °C. IR (KBr): 3456, 2927, 1711, 1586, 1489, 1462, 1439, 1217 cm 1. 1 H NMR (600 MHz, CDCl3): d = 6.84 (d, J = 7.8 Hz, 1H), 6.47 (d,

1400

L.-Q. Shen et al. / Steroids 77 (2012) 1398–1402

J = 7.8 Hz, 1H), 4.62 (broad s, 1H), 3.68 (s, 3H), 3.04 (dd, J = 13.8, 3.6 Hz, 1H), 2.64 (m, 1H), 2.58 (m, 1H), 2.48–2.43 (m, 2H), 2.17 (m, 1H), 2.14 (s, 3H), 2.03 (dt, J = 13.2, 2.4 Hz, 1H), 1.86–1.79 (m, 3H), 1.54 (dd, J = 13.2, 3.6 Hz, 1H),1.47–1.42 (m, 3H), 1.28–1.21 (m, 2H), 0.72 (s, 3H). 13C NMR (150 MHz, CDCl3): d = 174.9, 153.1, 138.7, 128.8, 127.7, 126.9, 113.0, 55.6, 55.2, 51.4, 45.03, 45.01, 40.4, 39.4, 29.3, 26.43, 26.40, 24.4, 23.9, 19.4, 14.3. HRMS: [M+] calcd for C21H28O3: 328.2038; found: 328.2032. 2.2.7. Methyl 1-allyloxy-4-methyl-1,3,5(10)-estratriene-17b– carboxylate (9) To a stirred suspension of sodium hydride (0.177 g, 7.4 mmol of a 60% dispersion in mineral oil) in DMF (25 mL) at 0 °C was added a solution of 8 (0.605 g, 1.8 mmol) in DMF (10 mL), and the reaction mixture was stirred at 10 °C for 3 h. The reaction solution was again cooled to 0 °C, allyl bromide (0.500 g, 4.1 mmol) was added, and the stirring was continued at room temperature for 12 h. The mixture was then poured into ice water (200 mL), and the aqueous layer was extracted with ethyl acetate (3  100 mL). The organic phase was washed, dried and concentrated in vacuo. The residue was flash-chromatographed (5–10% ethyl acetate in petroleum ether) to give 9. Yield: 0.625 g (92%). IR (KBr): 3080, 2925, 1726, 1581, 1516, 1467, 1449, 1437, 1215 cm 1. 1H NMR (600 MHz, CDCl3): d = 6.93 (d, J = 8.4 Hz, 1H), 6.63 (d, J = 8.4 Hz, 1H), 6.06 (m, 1H), 5.42 (dd, J = 17.4, 1.8 Hz, 1H), 5.26 (dd, J = 10.8, 1.8 Hz, 1H), 4.50 (m, 2H), 3.68 (s, 3H), 3.08 (dd, J = 13.8, 3.6 Hz, 1H), 2.67–2.57 (m, 2H), 2.5–2.54 (m, 2H), 2.20 (m, 1H), 2.16 (s, 3H), 2.02 (dt, J = 13.2, 3 Hz, 1H), 1.85–1.82 (m, 3H), 1.55 (dd, J = 13.2, 3.6 Hz, 1H), 1.50–1.39 (m, 3H), 1.26–1.17 (m, 2H), 0.73 (s, 3H). 13 C NMR (150 MHz, CDCl3): d = 174.9, 156.4, 138.4, 133.9, 129.3, 128.9, 127.4, 116.7, 109.6, 68.9, 55.6, 55.3, 51.4, 45.3, 45.1, 40.5, 39.5, 29.4, 26.8, 26.3, 24.4, 23.9, 19.3, 14.3. HRMS: [M+] calcd for C24H32O3: 368.2351; found: 368.2353. 2.2.8. 1-Allyloxy-4-methyl-17b-hydroxymethyl-1,3,5(10)-estratriene (10) To a stirred suspension of lithium aluminum hydride (0.077 g, 2.0 mmol) in THF(10 mL) was added dropwise a solution of 9 (0.500 g, 1.4 mmol) in THF (10 mL) at 0 °C under argon, and the reaction mixture was stirred at room temperature for 24 h. The reaction mixture was cooled 0 °C, and the reaction was carefully quenched with water (0.5 mL), 15% aqueous potassium hydroxide (0.5 mL), and water (1.5 mL). After being stirred for 45 min, the mixture was diluted with ethyl acetate (50 mL), filtered, and concentrated in vacuo. The residue was flash-chromatographed (20– 40% ethyl acetate in petroleum ether) to give 10 as a white solid. Yield: 0.397 g (86%); m.p. 110–111.5 °C. IR (KBr): 3389, 2935, 1649, 1584, 1474, 1257 cm 1. 1H NMR (600 MHz, CDCl3): d = 6.93 (d, J = 7.8 Hz, 1H), 6.63 (d, J = 8.4 Hz, 1H), 6.06 (m, 1H), 5.42 (dd, J = 17.4, 1.8 Hz, 1H), 5.25 (dd, J = 10.8, 1.8 Hz, 1H), 4.51 (m, 2H), 3.77 (m, 1H), 3.60 (m, 1H), 3.04 (dd, J = 13.2, 3.6 Hz, 1H), 2.68– 2.58 (m, 2H), 2.49 (t, J = 10.2, 1H), 2.17 (s, 3H), 1.91–1.90 (m, 2H), 1.85–1.73 (m, 3H), 1.49–1.43 (m, 4H), 1.33–1.20 (m, 3H), 0.72 (s, 3H). 13C (150 MHz, CDCl3): d = 156.3, 138.5, 133.9, 129.5, 128.8, 127.3, 116.6, 109.5, 68.9, 64.9, 55.3, 53.4, 45.5, 43.0, 40.0, 39.8, 29.4, 26.7, 26.3, 25.8, 24.4, 19.3, 13.3. HRMS: [M + 1+] calcd for C23H33O2: 341.2480; found: 341.2476. 2.2.9. 1-Allyloxy-4-methyl-17b-formyl-1,3,5(10)-estratriene (11) To a stirred solution of 10 (0.200 g, 0.6 mmol) in dichloromethane (20 mL) was added Dess-Martin periodinane (0.750 g, 1.8 mmol) at room temperature. After 45 min, saturated aqueous NaHCO3 (5 mL) and 10% Na2S2O3 (5 mL) were added to the reaction mixture, and the aqueous layer was extracted with dichloromethane (3  20 mL). The organic phase was washed with 10 mL of 1:1 10% Na2S2O3: saturated aqueous NaHCO3, followed by 10 mL of

H2O and 10 mL of brine, dried and concentrated in vacuo. The residue was flash-chromatographed (5–10% ethyl acetate in petroleum ether) to give 11. Yield: 0.179 g (90%); yellow oil. 1H NMR (600 MHz, CDCl3): d = 9.80 (s, 1H),6.92 (d, J = 7.8 Hz, 1H), 6.61 (d, J = 7.8 Hz, 1H), 6.04 (m, 1H), 5.41 (dd, J = 17.4, 1.8 Hz, 1H), 5.25 (dd, J = 10.8, 1.8 Hz, 1H), 4.48 (d, J = 4.8 Hz, 2H), 3.10 (dd, J = 10.8, 3.0 Hz, 1H), 2.66–2.55 (m, 2H), 2.50 (t, J = 9.6, 1H), 2.37 (t, J = 9.6, 1H), 2.19–2.17 (m, 1H), 2.15 (s,1H),2.03 (dt, J = 10.8, 3.6 Hz, 1H), 1.84–1.68 (m, 4H), 1.48–1.35 (m, 3H), 1.29–1.23 (m, 4H), 0.80 (s, 3H). 13C NMR (150 MHz, CDCl3): d = 205.1, 156.3, 138.3, 133.9, 129.0, 128.9, 127.5, 116.7, 109.6, 68.9, 63.3, 55.7, 45.8, 45.3, 39.8, 39.4, 29.3, 26.5, 26.3, 24.7, 21.3, 19.3, 14.7. HRMS: [M + 1+] calcd for C23H31O2: 339.2321; found: 339.2325. 2.2.10. 1-Allyloxy-19-nor-4-methylpregna-1,3,5(10),20-tetraene(12) To a stirred suspension of methyltriphenylphosponium bromide (0.533 g, 1.5 mmol) in THF (10 mL) was added dropwise n-BuLi (0.6 mL, 1.5 mmol) at 78 °C under argon. The reaction mixture was warmed to 0 °C and stirred for 1 h. The reaction mixture was again cooled to 78 °C, a solution of 11 (0.350 g, 1.1 mmol) in THF (8 mL) was added dropwise, and the stirring was continued at 0 °C for 6 h. The reaction was quenched by of saturated aqueous ammonium chloride at 0 °C, and the mixture was diluted with ethyl acetate (50 mL). The phase were separated, the aqueous layer was extracted with ethyl acetate (3  20 mL). The organic phase was washed, dried and concentrated in vacuo. The residue was flash-chromatographed (5–10% ethyl acetate in petroleum ether) to give 12. Yield:0.289 g (83%). IR (KBr): 3082, 2950, 1646, 1586, 1464,1260 cm 1. 1H NMR (600 MHz, CDCl3): d = 6.91 (d, J = 7.8 Hz, 1H), 6.60 (d, J = 7.8 Hz, 1H), 6.03 (m, 1H), 5.80 (m, 1H), 5.39 (dd, J = 17.4, 1.8 Hz, 1H), 5.23 (dd, J = 10.8, 1.8 Hz, 1H), 5.0–4.97 (m, 2H), 4.47 (m, 2H), 3.02 (dd, J = 10.8, 3.0 Hz, 1H), 2.64–2.59 (m, 2H), 2.46 (t, J = 12, 1H), 2.16 (s,3H),2.03 (m, 1H), 1.84–1.79 (m, 2H), 1.77–1.69 (m, 1H), 1.60–1.58 (m, 1H), 1.46–1.18 (m, 7H), 0.66 (s, 3H). 13C NMR (150 MHz, CDCl3): d = 156.4, 140.2, 138.6, 134.0, 129.6, 128.9, 127.3, 116.6, 114.6, 109.6, 68.9, 55.7, 55.0, 45.6, 44.7, 40.4, 38.5, 29.5, 27.5, 26.6, 26.4, 24.6, 19.4, 13.7. HRMS: [M+] calcd for C24H32O: 336.2453; found: 336.2449. 2.2.11. 19-nor-4-Methylpregna-1,3,5 (10), 20-tetraen-1-ol (Krempenes B (1)) To a stirred solution of 12 (0.110 g, 0.3 mmol) in methanol (10 mL) is added 5% Pd/C (0.52 g) and p-toluenesulfonic acid (0.025 g, 0.1 mmol). The reaction mixture was refluxed for 72 h. After removal of the catalyst by filtration, the filtration was concentrated in vacuo. The resulting residue was flash-chromatographed (20% ethyl acetate in petroleum ether) to give 1 as a white crystalline solid. Yield: 0.087 g (90%); m.p. 148.5–150 °C. [a]D25 = + 82.9 (c = 0.7, CH3OH). IR (KBr): 3493, 3072, 2923, 2856, 1635, 1590,1457, 1270, 1159, 1109 cm 1. 1H NMR (600 MHz, CDCl3): d = 6.84 (d, J = 7.8 Hz, 1H), 6.47 (d, J = 7.8 Hz, 1H), 5.82 (ddd, J = 7.8, 11.4, 18 Hz, 1H), 5.01 (dd, J = 17.4, 1.8 Hz, 1H), 4.99 (dd, J = 10.8, 1.8 Hz, 1H), 4.56 (broad s, 1H), 2.97 (dddd, J = 16.2, 4.2, 4.2, 3.6 Hz, 1H), 2.65 (ddd, J = 16.8, 16.2, 4.2, 1H),2.59 (ddd, J = 12, 12, 5.4 Hz, 1H), 2.43 (dd, J = 10.8, 10.2 Hz, 1H), 2.15 (s, 3H), 2.08 (ddd, J = 9, 9, 8.4 Hz, 1H), 1.84– 1.80 (m, 2H), 1.77–1.74 (m, 1H), 1.72 (ddd, J = 13.8, 3.6, 3.6 Hz 1H), 1.60–1.57 (m, 1H), 1.45(dddd, J = 12, 10.8, 10.2, 1.8 Hz, 1H), 1.35 (ddd, J = 12, 10.8, 6 Hz, 1H), 1.34–1.32 (m, 1H),1.28 (dd, J = 12.0, 10.8, Hz, 1H), 1.21(dddd, J = 16.2, 4.2, 4.2, 3.6 Hz, 1H), 1.22–1.18 (m, 1H), 0.67 (s, 3H). 13C NMR(150 MHz, CDCl3): d = 153.1, 140.1, 138.8, 128.8, 127.6, 127.2, 114.6, 113.0, 55.7, 55.0, 45.3, 44.6, 40.3, 38.5, 29.4, 27.5, 26.5, 26.2, 24.7, 19.4, 13.6. HRMS: [M+] calcd for C21H28O: 296.2140; found: 296.2134.

L.-Q. Shen et al. / Steroids 77 (2012) 1398–1402

1401

Fig. 3. ORTEP view of compound Krempenes B.

Fig. 2. ORTEP view of compound 7.

3. Results and discussion The synthesis of Krempenes B (1) from 3b-acetoxy-5-pregnen20-one 2 is summarized in Scheme 1. Treatment of pregnenolone acetate 2 with sodium hypobromite solution in dioxane, and subsequent acidified by concentrated hydrochloric acid afforded the required 3b-hydroxy-5-and -rostene-17b-carboxylic acid (3) in 95% yield [5]. Then, the COOH group was protected as the methyl ester to give 86% yield of methyl 3b-hydroxy-5-androstene-17bcarboxylate (4). Oppenauer oxidation of 4 provided methyl 3-oxo-androst-4-ene-17b-carboxylate (5) in 82% yield, which was then dehydrogenated with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) in dioxane at 110 °C to afford a satisfying yield (81.2%) of the methyl 3-oxo-androst-1,4-diene-17b-carboxylate (6) [7]. With 6 in hand (Scheme 1), introduction of the steroids containing ring A aromatic was achieved by carrying out the dienone-phenol rearrangement [8] of steroids 6 with p-toluenesulfonic acid in acetic anhydride to yield the methyl 1-acetoxy-4-methyl1,3,5(10)-estratriene-17b-carboxylate (7) in 85% yield, the structure of which was confirmed by X-ray crystal structure analysis (Fig. 2) [9]. Compound 7 was easily hydrolyzed with saturated sodium bicarbonate in metnanol at room temperature to gain methyl 1-hydroxy-4-methyl-1,3,5(10)-estratrien-17b-carboxylate (8) in 97.3% yield. We investingated a protecting group for the phenolic oxygen of compound 8. We examined the feasibility of employing the tertbutyldimethylsilyl ether and aryl toluenesulfonate protecting groups [10], but it was unsuccessful possibly for the phenolic hydroxy possessing very weak acid and steric hindrance offered by the C10 methylene protons of the ring B. In the end, the allyl group emerged as the suitable protecting group for the phenolic oxygen. Deprotonation of compound 8 with sodium hydride in dimethylformamide (DMF) at room temperature followed by alkylation of the intermediate phenoxide anion with allyl bromide provided methyl 1-allyloxy-4-methyl-1,3,5 (10)-estratriene-17b-carboxylate (9) in 92% yield [11]. The subsequent reduction of compound 9 with lithium aluminum hydride in THF gave 1-allyloxy-4methyl-17b-hydroxymethyl-1,3,5(10)-estratriene (10) in 86% yield. Oxidation of the primary alcohol group of compound 10 was brought about by means of Dess–Martin periodinane (DMP) in CH2Cl2 at room temperature and produced the required 1-allyloxy-4-methyl-17b-formyl-1,3,5(10)-estratriene (11) in 90% yield. No epimerization of the side chain at C (17) was observed under these conditions, since 1H NMR and 13C NMR spectra of aldehyde hydrogen displayed a single set of signals. The next step was to construct pregn-20-ene. The reaction of estratriene 11 with Wittig reagent prepared from methyltriphenyl-

phosphonium bromide in dry THF at 78 °C afforded 1-allyloxy19-nor-4-methylpregna-1,3,5(10), 20-tetraene (12, 83%) [12]. The construction pregn-20-ene had also been described in the literature by the treatment of pregnenolone tosylhydrazone with metal lithium reagent and pregnenolone hydrazone with iodine, triethylamine treated and sodium [13]. However, the reports based on pregna-4,20-diene-3-one was dehydrogenated with 2,3-dichloro5,6-dicyano-1,4-benzoquinone (DDQ) to give pregna-1,4,20trien-3-one in the low yield (22%) [14]. With precursor 12 in hand (Scheme 1), the cleavage of allyl group in 12 with lithium and a catalytic amount of naphthalene (4 mol%) in THF at low temperature or under basic conditions in the presence of a catalytic amounts of Pd(PPh3)4 was attempted, but without success [15]. Fortunately, in the end, the removle of the allyl group in 12 with 5% Pd/C and p-toluenesulfonic acid (PTS) in methanol at 65.5 °C for 72 h afford the desired Krempenes B (1) in 90% yield [16]. The analytical and spectral data of the synthetic Krempenes B (1) were consistent with that of the natural Krempenes B (1). The structure was also determined by X-ray diffraction of Krempenes B (1) (Fig. 3) [17].

4. Conclusions We have completed the first synthesis of Krempenes B (1), in eleven steps from commercially available 3b-acetoxy-5-pregnen20-one 2 with an overall yield of 23.9%. Furthermore, our sequence is well suited to the generation of structural analogues. The biological activity of 1 and analogues of 1 are currently investigated and will be reported in due course.

Acknowledgements This work was supported by National Natural Science Foundation of China (No. 21062002), the Scientific Research Program of Guangxi University for Nationalities (Project No. 2010QD018 and 2010ZD010).

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