V. P. Bui et al. / Tetrahedron Letters 43 (2002) 2839–2841
2841
9910412), and TDC Research Inc., as well as to the
Agricultural University of Norway (T.V.H. and Y.S.)
and the Norwegian Research Council (T.V.H.) for gen-
erous financial support.
to ambient temperature, ampicillin (100 mg L−1) was
added, and each flask inoculated with a single colony of
E. coli JM109 (pDTG 602) from a fully grown plate. The
inoculum was grown overnight on an orbital shaker
(35°C, 150 rpm) and added to sterilized production
medium (8 L) in a 15 L fermentor. The production
medium consists of an aqueous mixture of KH2PO4 (60
g), citric acid (16 g), MgSO4·7H2O (40 g), trace metal
solution (16 mL) including: [Na2SO4 (1 g L−1), MnSO4 (2
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g
L−1), ZnCl2 (2 L−1), CoCl2·6H2O (2 L−1),
g g
CuSO4·5H2O (0.3 g L−1), FeSO4·7H2O (10 g L−1), pH
1.0], conc. H2SO4 (9.6 mL) and ferric ammonium citrate
(9.6 mL, 270 g L−1). The mixture was neutralized with
conc. ammonia and supplemented with ampicillin (800
mg) and thiamine hydrochloride (2.69 g). The culture was
grown for 24 h with concentrated glucose solution (720 g
L−1) as the carbon source, then induced with 80 mg
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8. General procedure for the large-scale (15 L) fermentation
with E. coli JM109 (pDTG602): A mineral salts broth
(MSB, 600 mL) containing K2HPO4 (9.6 g), KH2PO4 (8.4
g), (NH4)2SO4 (3 g), yeast extract (9 g), glucose (18 g),
and MgSO4·7H2O (1.2 g) was divided into two 2.8 L
Fernbach flasks and sterilized. After the broth had cooled