3676
R. N. Patel et al. / Tetrahedron: Asymmetry 14 (2003) 3673–3677
was stirred at room temperature (22–25°C) and moni-
tored by TLC (silica gel; ethyl acetate/hexane 2:1; RF
(2)=0.42; RF (3)=0.2) for the disappearance of the
acetoxy b-lactam. After the reaction was complete the
pH of the reaction mixture was adjusted to 7 using an
acidic resin (AG 50W×8; 50–100 mesh size, hydrogen
form). The reaction mixture was then filtered through a
pad of Celite to remove the resin. Methanol was then
removed on the rotary evaporator at ambient tempera-
ture under reduced pressure. The crude hydroxy b-lac-
tam was then dried in a vacuum oven at 50°C (house
vacuum with a flow of nitrogen) to give 39.7 g of
material. To a 1-L, 3-necked flask equipped with a
magnetic stirrer, temperature probe and reflux con-
denser was charged the crude hydroxy b-lactam 4 (37.4
g) followed by 561 mL of reagent grade ethyl acetate.
The mixture was heated to reflux to dissolve the b-lac-
tam. The reaction mixture was then cooled to 60–65°C
and seeded with the enantiomerically pure hydroxy
b-lactam 4. The mixture was stirred at 60–65°C for 4 h
during which time the product started to crystallize.
The pot temperature was lowered to room temperature
over 4–5 h and the mixture was stirred at room temper-
ature (22–24°C) for 16–18 h. The reaction mixture was
cooled to 0°C and stirred for 1 h. Filtration followed by
washing with 2×50 mL of cold ethyl acetate and drying
under reduced pressure at 40–50°C gave 29.7g of 4 as a
white solid (87% recovery; >99.9% ee as determined by
chiral HPLC). 1H NMR (300 MHz; DMSO-d6): l
8.0.92 (s, 9H), 3.20 (d, J=4.9 Hz, 1H), 4.67–4.70 (m,
1H), 5.91 (d, J=6.8 Hz, 1H), 8.11 (s, 1H) ppm. 13C
NMR (75 MHz; DMSO-d6): l 24.3, 30.9, 60.7, 74.7,
169.2 ppm. Anal. calcd for C7H13NO2: C, 58.72; H,
9.15; N, 9.78. Found: C, 58.72; H, 9.11; N, 9.81.
HRMS: calcd for C7H14NO2: 144.1023 (M+H)+.
Found:144.1025 (M+H)+. A similar procedure was used
to isolate 60 g of 4 from the 7.5-L reaction mixture
obtained using the immobilized BMS lipase.
Richard Mueller and David Kronenthal for reviewing
this manuscript and providing valuable suggestions.
References
1. Wani, M. C.; Taylor, H. L.; Wall, M. E.; Coggon, P.;
McPhail, A. T. J. Am. Chem. Soc. 1971, 93, 2325–2327.
2. Suffness, M.; Wall, M. E. In Taxol: Science and Applica-
tion; Suffness, M., Ed. Discovery and development of
taxol; CRC Press: New York, 1995; pp. 3–25.
3. Holton, R. A.; Biediger, R. J.; Boatman, P. J. In Taxol:
Science and Application; Suffness, M., Ed. Semisynthesis
of taxol and taxotere; CRC Press: New York, 1995; pp.
97–123.
4. Kingston, D. G. I. In Taxol: Science and Application;
Suffness, M., Ed. Natural taxoids: structure and chem-
istry; CRC Press: New York, 1995; pp. 287–317.
5. Synder, J. A.; McIntosh, R. J. Ann. Rev. Biochem. 1976,
45, 699–715.
6. Schiff, P. B.; Fant, J.; Horwitz, S. B. Nature 1979, 277,
665–667.
7. Patel, R. N. Ann. Rev. Microbiol. 1995, 98, 361–395.
8. Rose, W. C.; Long, B. H.; Fairchild, C. R.; Lee, F. Y.;
Kadow, J. F. Clin. Cancer Res. 2001, 7, 2016–2021.
9. Liu, J. PCT Int. Appl. 2001, 23 pp. A2 20010802 CAN
135:137629 AN 2001:565021.
10. Guillemard, V.; Saragovi, H. Cancer Res. 2001, 61, 694–
699.
11. Lin, S.; Ojima, I. Expert Opinion on Therapeutic Patents
2000, 10, 869–889.
12. Chai, K. B.; Moon, Y. H.; Kim, N. D.; Ha, T. H.; Shin,
J. A.; Lim, C. G.; Kim, W. J.; Lee, G. S.; Suh, K. H.
PCT Int. Appl. 1999, 64 pp. CODEN: WO 9937631 A1
19990729 CAN 131:116387 AN 1999:487282.
13. Greenwald, R. B.; Bolikal, D. 1997, 6 pp., US 5622986 A
19970422 CAN 126:330727 AN 1997:287180.
14. Rothenberg, M. L. Current Opinion in Investigational
Drugs 1993, 2, 1269–1277.
4.5. Analytical methods
15. Holton, R. A.; Nadizadeh, H.; Beidiger, R. J. Eur. Pat.
Appl. 1993, 17 pp. EP 534708 A1 19930331 CAN
119:49693 AN 1993:449693.
16. Mathew, A. E.; Mejillano, M. R.; Nath, J. P.; Himes, R.
H.; Stella, V. J. J. Med. Chem. 1992, 35, 145–151.
17. Gou, D.-M.; Liu, Y.-C.; Chen, C.-S. J. Org. Chem. 1993,
58, 1287–1289.
18. Brieva, R.; Crich, J. Z.; Sih, C. J. Org. Chem. 1993, 58,
1068–1072.
19. Patel, R. N.; Banerjee, A.; Howell, J. M.; McNamee, C.
G.; Brzozowski, D.; Nanduri, V.; Thottathil, J. K.;
Szarka, L. J. Tetrahedron: Asymmetry 1993, 4, 2069–
2084.
The racemic acetate and racemic alcohol were analyzed
by HPLC. A Nova Pak C18 reverse-phase column
(4×150 mm) was used. The mobile phase was 15% (v/v)
acetonitrile in water, and the flow rate was 1 mL/min.
The detection wavelength was 227 nm. The retention
times for the acetate and alcohol were 15.2 and 3.2 min,
respectively. The separation of enantiomers of acetate
and alcohol was carried out by chiral HPLC. A Chiral-
cel AD column was used (Diacel Chemical Industries
Inc. Ltd, Chiral Technologies, Easton, PA). The mobile
phase consisted of 1.96% absolute ethanol in hexane
and was used at 1 mL/min at ambient temperature. The
detection wavelength was 210 nm. The retention times
for the two acetate enantiomers were 22 and 50 min,
respectively. The retention times for the two alcohol
enantiomers were 36 and 40 min, respectively.
20. Food & Drug Administration. Chirality, 1992, 4, 338–
340.
21. Buckland, B. C.; Robinson, D. K.; Chartrain, M.
Metabolic Eng. 2000, 2, 42–48.
22. O’Brien, M. K.; Vanasse, B. Curr. Opin. Discov. Dev.
2000, 3, 793–806.
23. Pesti, J. A.; Dicosimo, R. Curr. Opin. Discov. Dev. 2000,
3, 764–782.
Acknowledgements
24. Mahmoudian, M. Biocat. Biotrans. 2000, 18, 105–116.
25. Stewart, D. Curr. Opin. Chem. Biol. 2001, 5, 120–
129.
We would like to acknowledge Dr. John Wasylyk for
development of the HPLC assay system and Drs.