Sulfate Ion → (R)-2-Octanol

Candida utilis IFO 0988 strain was cultured in 3.6 L of medium B and the microbial cells were prepared by centrifugation. Obtained microbial cells were suspended in 100 mM potassium phosphate buffer (pH 8.0), 10% glycerol, 1 mM ethylenediaminetetraacetic acid 2Na (EDTA·2Na), 0.01% 2-mercaptoethanol, and 2 mM phenyl methane sulfonylfluoride (PMSF) and homogenized with bead beater (Biospec). After that, microbial cell debris were removed by centrifugation and cell-free extract was obtained. Protamine sulphate was added to this cell-free extract, and supernatant was obtained by centrifugation to remove nucleic acid. Ammonium sulfate was added to the supernatant, and the supernatant which was crude enzymatic solution was obtained in 30% saturation. The solution was dialyzed with standard buffer including 30% saturated ammonium and added to phenyl-sepharose HP (2.6 cm×10 cm) equilibrated with the same buffer. The gradient elution was conducted with 30% to 0% saturated ammonium sulfate. The fraction having selectivity of (R)-2-octanol among extracted fractions was recovered and concentrated by ultrafiltration. Composition of medium B used are followings:

1. customthe microbial cells were prepared by centrifugation
2. customObtained microbial cells
3. customAfter that, microbial cell debris were removed by centrifugation
4. extractioncell-free extract
5. customwas obtained
6. customsupernatant was obtained by centrifugation
7. customto remove nucleic acid
8. additionAmmonium sulfate was added to the supernatant
9. customwas obtained in 30% saturation
10. customThe solution was dialyzed with standard
11. additionadded to phenyl-sepharose HP (2.6 cm×10 cm)
12. washThe gradient elution
13. extractionamong extracted fractions
14. customwas recovered
15. concentrationconcentrated by ultrafiltration

ORD record

ord-60e38230c8a049f1bead7b52682bb6ba

Source dataset

uspto-grants-2007_04

DOI

10.6084/m9.figshare.5104873.v1

Patent number

US07202069B2