Biooxidation of Cycloketones
overnight at room temperature. The ion-exchange resin was
separated by filtration over a bed of Celite, and the solvent was
evaporated. Crude product was purified by column chromatography
(SiO2 ) 1:50, LP/EtOAc ) 6:1), which gave pure 7g21 as a colorless
solid in 53% yield.
endo-Tricyclo[5.2.1.02,6]decan-10-one (7a). Ketone 7a was
prepared according to the general procedure for catalytic hydro-
genation using endo-tricyclo[5.2.1.02,6]dec-8-en-10-one 5a (1.00 g,
6.70 mmol) as the starting material. Crude product was purified
by column chromatography (SiO2, LP/EtOAc ) 100:1) to give 7a35
as a yellow oil (0.86 g) in 85% yield. 1H NMR (CDCl3): δ 1.63-
1.75 (m, 12H), 2.45-2.51 (m, 2H); 13C NMR (CDCl3): δ 27.7 (t),
29.2 (t), 29.9 (t), 38.2 (d), 43.6 (d), 214.9 (s).
endo-Tricyclo[6.2.1.02,7]-undecan-11-one (7b). Ketone 7b was
prepared according to the general procedure for catalytic hydro-
genation using endo-tricyclo[6.2.1.02,7]undec-9-en-11-one (5b) (1.00
g, 6.2 mmol) as the starting material. The crude product was purified
by column chromatography (SiO2, LP/EtOAc ) 100:1) to give 7b
as a yellow oil (0.89 g) in 87% yield. 1H NMR (CDCl3): δ 0.81-
1.89 (m, 14H), 2.06-2.14 (m, 2H); 13C NMR (CDCl3): δ 17.2 (t),
18.1 (t), 19.5 (t), 32.9 (d), 43.4 (d), 216.7 (s). Calcd C 80.44%, H
9.82%, found C 80.49%, H 9.43%.
was extracted 2 times with dichloromethane. The combined organic
phases were washed with saturated bicarbonate and brine, dried
over sodium sulfate, and evaporated
Oxa-tricyclo[5.2.2.02,6]undecan-9-one (8a). endo-Tricyclo-
[5.2.1.02,6]decan-11-one 7a (100 mg, 0.67 mmol) was oxidized with
E. coli overexpressing CHMORhodo2 and CHMOBrevi2 according to
the general procedure. The crude product was purified via column
chromatography (SiO2, LP/EtOAc ) 24:1), and 8a was isolated as
colorless crystals in the following yields: CHMORhodo2 in 63% yield
D
(70 mg) and CHMOBrevi2 in 67% yield (74 mg). [R]20 (CHCl3)
D
(CHMORhodo2): -17.30, c ) 1.70 g/100 mL; [R]20 (CHCl3)
1
(CHMOBrevi2): +13.04, c ) 1.43 g/100 mL. mp: 78-80 °C. H
NMR (CDCl3): δ 1.66-1.90 (m, 12H), 2.49-2.55 (m, 1H), 4.54-
4.59 (m, 1H); 13C NMR (CDCl3): δ 16.7 (t), 20.6 (t), 27.8 (t),
27.9 (t), 28.1 (t), 37.7 (d), 39.2 (d), 41.2 (d), 78.9 (d), 177.3 (s).
Calcd C 72.26%, H 8.49%, found C 72.48%, H 8.78%.
(1R,4R,5R,8R)-1,4-Ethano-5,8-methano-1,4,4a,5,6,7,8,8a-oc-
tahydro-3H-2-benzopyran-3-one (8c). Tetracyclic ketone 7c (100
mg, 0.57 mmol) was oxidized with E. coli overexpressing CHMOA
-
cineto according to the general procedure. The crude product was
purified by column chromatography (SiO2, LP/EtOAc ) 20:1), and
8c (56 mg, 51% yield) was isolated as colorless crystals. mp: 52-
D
54 °C. [R]20 (CHCl3) (CHMOAcineto): -39.63; c ) 0.965 g/100
(1R,4R,5â,8â)-1,2,3,4,4a,5,6,7,8,8a-Decahydro-dimethanon-
aphthalen-10-one (7c). Compound 6c (1.71 g, 7.71 mmol) was
converted according to the general procedure for ketal hydrolysis
using pTSA. Chromatographic purification of crude product (SiO2,
LP/EtOAc ) 100:1) provided 7c as a colorless oil (0.85 g) in 63%
mL. 1H NMR (CDCl3): δ 1.10-1.22 (m, 3H), 1.58-2.25 (m, 11H),
2.63-2.68 (m, 1H), 4.61-4.66 (m, 1H); 13C NMR (CDCl3): δ
18.4 (t), 22.7 (t), 30.3 (t), 30.6 (t), 34.7 (t), 36.2 (d), 37.9 (d), 38.9
(d), 42.1 (d), 46.3 (d), 78.6 (d), 176.8 (s). Calcd C 74.97%, H
8.39%; found C 74.88%, H 8.34%.
1
yield. H NMR (CDCl3): δ 1.11-1.20 (m, 2H), 1.54-1.72 (m,
4,8-Dioxa-tricyclo[5.2.2.02,6]undecan-9-one (8g). 4-Oxatricyclo-
[5.2.1.02,6]decan-10-one 7g (100 mg, 0.57 mmol) was oxidized with
E. coli overexpressing CHMOAcineto and CPMOComa according to
the general procedure. The crude product was purified by column
chromatography (SiO2, LP/EtOAc ) 3:1), and the colorless crystals
were isolated in the following yields: CHMOAcineto in 53% yield
4H), 1.79-2.06 (m, 8H), 2.29-2.36 (m, 2H); 13C NMR (CDCl3):
δ 18.3 (t), 31.3 (t), 36.4 (t), 37.2 (d), 43.0 (d), 46.9 (d), 215.3 (s).
Calcd C 81.77%, H 9.15%; found C 81.52%, H 9.12%.
General Procedures for Screening Experiments in Plates.
Plates with either 12 or 24 wells were utilized. Each well (2 or 1
mL, respectively) was charged with LBamp medium (5.0 g of
peptone, 2.5 g of yeast extract, 5.0 g of NaCl, 500 mL of deionized
water, 2 mL of ampicilline stock solution -(50 mg/mL)) and
inoculated with 1% of an overnight preculture of recombinant E.
coli overexpressing BVMOs from different bacterial origin. A plate
was incubated at 120 rpm at 37 °C on an orbital shaker for 2 h.
Isopropyl-â-D-galactopyranoside (IPTG) was added (final concen-
tration of 0.025 mM) together with substrate (final concentration
of 3-6 mM). The plate was shaken at room temperature. After 24
h, samples for GC measurement were taken (700 µL of culture
was centrifuged, and the supernatant was extracted with 700 µL of
EtOAc supplemented with a defined concentration of methyl
benzoate as the internal standard).
General Procedures for Biotransformations on Preparative
Scale. Fresh LBamp medium (250 mL) was inoculated with 1% of
an overnight preculture of recombinant E. coli strains overexpress-
ing the appropriate BVMO in a baffled Erlenmeyer flask. The
culture was incubated at 120 rpm at 37 °C on an orbital shaker for
2 h, and then IPTG was added to a final concentration of 0.025
mM. The substrate (100 mg) was added neat along with â-cyclo-
dextrin (1 equiv). The culture was incubated at room temperature
until GC showed complete conversion of the ketone (24-72 h).
General Procedure for Chemical Baeyer-Villiger Oxidation.
mCPBA (1.5 equiv, 70% purity) was added to a solution of the
corresponding ketone in dry dichloromethane (10% solution), and
the mixture was stirred overnight. Complete conversion was
determined by TLC and gas chromatography. Excess triethylamine
was added, and the mixture was stirred for 15 min. Afterward, water
was added, and the organic layer was separated. The aqueous layer
D
(58 mg) and CPMOComa in 49% yield (54 mg). [R]20 (CHCl3)
D
(CHMOAcineto): -17.68, c ) 0.57 g/100 mL; [R]20 (CHCl3)
(CPMOComa): +15.66, c ) 0.49 g/100 mL. mp: 116-118 °C. 1H
NMR (CDCl3): δ 1.55-2.13 (m, 4H), 2.54-2.58 (m, 1H), 2.64-
2.80 (m, 2H), 3.49-3.65 (m, 2H), 3.78 (d, J ) 10.2 Hz, 1H), 3.96
(d, J ) 9.8 Hz, 1H), 4.56-4.60 (m, 1H); 13C NMR (CDCl3): δ
17.3 (t), 21.1 (t), 38.1 (d), 38.6 (d), 41.3 (d), 69.2 (t), 70.5 (t), 77.9
(d), 175.9 (s). Calcd C 74.97%, H 8.39%; found C 74.88%, H
8.34%.
Acknowledgment. This project was funded by the Austrian
Science Fund (FWF, Project I19-B10), the Deutsche Fors-
chungsgemeinschaft (Ba 1372/11), and the Fonds der Chemis-
chen Industrie. The authors thank Dr. Pierre E. Rouviere (E.I.
DuPont Company) for supporting this project by a generous
donation of six E. coli expression systems for BVMOs and Prof.
Margaret M. Kayser (University of New Brunswick) for
providing the strain of CPMOComa
.
Note Added after ASAP Publication. At the top of page 4,
CHMOAcineto was missing from the sentence in the version
published ASAP on November 15, 2007; the corrected version
was published November 16, 2007.
Supporting Information Available: 1H, 13C, and DEPT NMR
1
spectra of ketones 7a-c,h and lactones 8a-c,f-h and H NMR
spectra of previously reported compounds; CIF file for X-ray
structure determination of (-)-8g. This material is available free
(35) Neff, J. R.; Nordlander, J. E. J. Org. Chem. 1976, 41, 2590-2596.
JO701704X
J. Org. Chem, Vol. 72, No. 25, 2007 9603