a centrifuge tube, which was shaken at 125 rpm at 25 ◦C for
24 h. Centrifugation (10 000 rpm, 10 min) and drying in vacuo
afforded BCL immobilized on BIO-M (500 mg). The amounts
of enzyme in the solutions before and after immobilization were
determined by the method of Bradford22 to calculate the content
of enzyme bound to BIO-M (4.6% (w/w)).
ee; 1H NMR (CDCl3, 500 MHz) 0.91 (t, J = 7.5 Hz, 3H), 1.91–
2.03 (m, 2H), 2.10 (s, 3H), 5.83 (t, J = 7.0 Hz, 1H), 7.45–7.48
(m, 3H), 7.78–7.84 (m, 4H); HPLC: Chiralcel OJ-H, hexane/i-
PrOH (9 : 1), flow rate 0.5 mL min-1, detection 254 nm, (R) 18.7
min, (S) 25.7 min.
Determination of the specific surface area of the material
Immobilization of CAL. A solution of CAL (Novozymes,
Lipozyme CALB L, 50 mL) was dialyzed as shown above.
This enzyme solution (87 mL) and BIO-M (500 mg) were
put in a centrifuge tube and shaken at 125 rpm at 25 ◦C for
24 h. Centrifugation (10 000 rpm, 10 min) and drying in vacuo
afforded CAL immobilized on BIO-M (489 mg). The content of
enzyme bound to BIO-M was determined as shown above (3.1%
(w/w)).
Nitrogen adsorption isotherms at 77 K were measured in liquid
nitrogen baths with Belsorp mini-II (Bel Japan, Inc.). Prior to
the measurements, samples were degassed in vacuo at 100 ◦C
for 4 h. The specific surface areas were calculated by using the
Brunauer–Emmett–Teller (BET) equation.23
Acknowledgements
Kinetic resolution of secondary alcohols with lipase immobilized
on chemically modified BIO
This work was supported by a grant from the Ministry of
Education, Culture, Sports, Science and Technology (MEXT) of
Japan. We thank Dr S. Hayakawa (Okayama University) for the
measurement of solid-state NMR. We also thank Dr Y. Ikeda
(Kyoto University) and Dr Y. Kusano (Kurashiki University
of Science and the Arts) for valuable discussions. CAL was
provided from Novozymes Japan, and synthetic g-Fe2O3 was
a gift from Toda Kogyo Corporation.
Typical procedure. A mixture of 1 (1.00 mmol), BIO-M-
immobilized lipase (10.0 mg), and molecular sieves 3A (three
pieces) in dry i-Pr2O (5 mL) was stirred at 30 ◦C for 30 min.
The reaction was started by addition of vinyl acetate (185 mL,
2.00 mmol). The progress of the reaction was monitored by
thin layer chromatography (TLC). The mixture was filtered and
concentrated. Alcohol (S)-1 and ester (R)-2 were separated by
silica gel column chromatography. The enantiomeric purities
were determined by chiral GC or HPLC.
References
1 Book and reviews: (a) Immobilized Catalysts: Solid Phases, Immobi-
lization and Applications, ed. A. Kirschning, Springer, Berlin, 2004;
(b) T. Matsumoto, M. Ueno, N. Wang and S. Kobayashi, Chem.–
Asian J., 2008, 3, 196–214; (c) Y. Uozumi, Bull. Chem. Soc. Jpn.,
2008, 81, 1183–1195.
2 Books: (a) C.-H. Wong and G. M. Whitesides, Enzymes in Synthetic
Organic Chemistry, Elsevier, Oxford, 1994; (b) Enzymatic Reactions
in Organic Media, ed. A. M. P. Koskinen and A. M. Klibanov,
Blackie Academic, Glasgow, 1996; (c) Enzyme Catalysis in Organic
Synthesis, ed. K. Drauz and H. Waldmann, Wiley-VCH, Weinheim,
2nd edn, 2002; (d) K. Faber, Biotransformations in Organic Chemistry,
Springer-Verlag, Berlin, 5th edn, 2004; (e) A. S. Bommarius and
B. R. Riebel, Biocatalysis, Wiley-VCH, Weinheim, 2004; (f) U. T.
Bornscheuer and R. J. Kazlauskas, Hydrolases in Organic Synthesis,
Wiley-VCH, Weinheim, 2nd edn, 2006; (g) Industrial Biotransfor-
mations, ed. A. Liese, K. Seelbach and C. Wandrey, Wiley-VCH,
Weinheim, 2nd edn, 2006.
3 For immobilized enzymes, see: (a) M. T. Reetz, A. Zonta, V.
Vijayakrishnan and K. Schimossek, J. Mol. Catal. A: Chem., 1998,
134, 251–258; (b) M. Kamori, T. Hori, Y. Yamashita, Y. Hirose and Y.
Naoshima, J. Mol. Catal. B: Enzym., 2000, 9, 269–274; (c) M. Suzuki,
C. Nagasawa and T. Sugai, Tetrahedron, 2001, 57, 4841–4848; (d) T.
Sakai, K. Hayashi, F. Yano, M. Takami, M. Ino, T. Korenaga and
T. Ema, Bull. Chem. Soc. Jpn., 2003, 76, 1441–1446; (e) M. T. Reetz,
P. Tielmann, W. Wiesenho¨fer, W. Ko¨nen and A. Zonta, Adv. Synth.
Catal., 2003, 345, 717–728; (f) U. Drechsler, N. O. Fischer, B. L.
Frankamp and V. M. Rotello, Adv. Mater., 2004, 16, 271–274; (g) H.
M. R. Gardimalla, D. Mandal, P. D. Stevens, M. Yen and Y. Gao,
Chem. Commun., 2005, 4432–4434; (h) H. R. Hobbs, B. Kondor, P.
Stephenson, R. A. Sheldon, N. R. Thomas and M. Poliakoff, Green
Chem., 2006, 8, 816–821; (i) S. van Pelt, S. Quignard, D. Kuba´c, D.
Y. Sorokin, F. van Rantwijk and R. A. Sheldon, Green Chem., 2008,
10, 395–400; (j) W. Wang, Y. Xu, D. I. C. Wang and Z. Li, J. Am.
Chem. Soc., 2009, 131, 12892–12893; (k) R. A. Sheldon, Org. Process
Res. Dev., 2011, 15, 213–223.
Kinetic resolution of 1a. The products were characterized
according to the literature.14b (S)-1a: 31% yield; 83% ee; 1H
NMR (CDCl3, 300 MHz) 1.51 (d, J = 6.3 Hz, 3H), 1.77 (d,
J = 3.0 Hz, 1H), 4.87–4.95 (m, 1H), 7.28–7.41 (m, 5H); GC: CP-
cyclodextrin-b-2,3,6-M-19, Inj. 250 ◦C, Col. 95 ◦C, Det. 220 ◦C,
(R) 30.0 min, (S) 32.6 min. (R)-2a: 23% yield; 98% ee; 1H NMR
(CDCl3, 300 MHz) 1.54 (d, J = 6.8 Hz, 3H), 2.07 (s, 3H), 5.88
(q, J = 6.8 Hz, 1H), 7.27–7.36 (m, 5H); GC: CP-cyclodextrin-b-
2,3,6-M-19, Inj. 250 ◦C, Col. 95 ◦C, Det. 220 ◦C, (S) 24.6 min,
(R) 27.5 min.
Kinetic resolution of 1b. (S)-1b: 56% yield; 69% ee; [a]1D9 -32.6
(c 0.96, CHCl3); lit.16d [a]D26 -11.5 (c 1.03, CHCl3) for (S)-1b with
25% ee; 1H NMR (CDCl3, 500 MHz) 1.59 (d, J = 6.0 Hz, 3H),
1.88 (s, 1H), 5.08 (q, J = 6.0 Hz, 1H), 7.45–7.53 (m, 3H), 7.82–
7.85 (m, 4H); HPLC: Chiralcel OJ-H, hexane/i-PrOH (9 : 1),
flow rate 0.5 mL min-1, detection 254 nm, (S) 30.2 min, (R) 39.8
min. (R)-2b: 40% yield; >99% ee; [a]2D0 +117 (c 1.04, CHCl3);
5
lit.16d [a]2 +88.1 (c 1.18, CHCl3) for (R)-2b with >99% ee; 1H
D
NMR (CDCl3, 500 MHz) 1.63 (d, J = 8.3 Hz, 3H), 2.10 (s, 3H),
6.05 (q, J = 8.3 Hz, 1H), 7.48–7.49 (m, 3H), 7.81–7.85 (m, 4H);
HPLC: Chiralcel OJ-H, hexane/i-PrOH (9 : 1), flow rate 0.5 mL
min-1, detection 254 nm, (R) 25.1 min, (S) 28.9 min.
Kinetic resolution of 1c. (S)-1c: 59% yield; 77% ee; [a]2D2 -28.8
(c 1.03, CHCl3); lit.16d [a]D28 -4.14 (c 1.11, CHCl3) for (S)-1c
1
with 7% ee; H NMR (CDCl3, 500 MHz) 0.95 (t, J = 7.0 Hz,
3H), 1.83–1.95 (m, 3H), 4.78 (t, J = 5.3 Hz, 1H), 7.45–7.49
(m, 3H), 7.79–7.85 (m, 4H); HPLC: Chiralcel OJ-H, hexane/i-
PrOH (9 : 1), flow rate 0.5 mL min-1, detection 254 nm, (S) 24.4
min, (R) 33.5 min. (R)-2c: 40% yield; 97% ee; [a]2D2 +96.8 (c 1.11,
CHCl3); lit.16d [a]D29 +43.7 (c 0.65, CHCl3) for (R)-2c with >99%
4 For shape-controlled materials produced by living organisms, see for
example: (a) S. Mann, Biomineralization: Principles and Concepts in
Bioinorganic Materials Chemistry, Oxford University Press, Oxford,
2001; (b) Handbook of Biomineralization: Biological Aspects and
Structure Formation, ed. E. Ba¨uerlein, Wiley-VCH, Weinheim,
2007.
3194 | Green Chem., 2011, 13, 3187–3195
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