(3R)-3-aminoazepan-2-one

反応条件: temp:120.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

実験手順: To a solution of (R)-lysine HCl salt (100 g, 0.68 mol) in xylene (1 L) was added hexamethyldisilazane (1000 mL, 4.76 mol) and chlorotrimethylsilane (10 mL, 78.8 mmol). The mixture was heated to 120° C. for 24 h, then to 180° C. for 48 h. The solvent was then removed under reduced pressure and the crude material was purified by column chromatography (20:1 to 10:1 CH2Cl2/MeOH) to afford (R)-3-aminoazepan-2-one (I-1a). The solvent was then removed under reduced pressure the crude material was purified by column chromatography (20:1 to 10:1 CH2Cl2/MeOH)

反応条件: temp:120.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

実験手順: To a solution of (R)-lysine HCl salt (100 g, 0.68 mol) in xylene (1 L) was added hexamethyldisilazane (1000 mL, 4.76 mol) and chlorotrimethylsilane (10 mL, 78.8 mmol). The mixture was heated to 120° C. for 24 h, then to 180° C. for 48 h. The solvent was then removed under reduced pressure and the crude material was purified by column chromatography (20:1 to 10:1 CH2Cl2/MeOH) to afford (R)-3-aminoazepan-2-one (I-1a). The solvent was then removed under reduced pressure the crude material was purified by column chromatography (20:1 to 10:1 CH2Cl2/MeOH)

反応条件: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

実験手順: Then D-α-amino-ε-caprolactam and pyridoxal-5-phosphate were added to the disintegrated cell suspensions of each monoculture to concentrations of 5 g/l and 0.01 mM respectively. After 1.5 h of incubation at 20° C. and 20 rpm, 2 ml samples were transferred into 1 ml of 1 M H3PO4. After removal of all particulates by centrifugation followed by filtration through a 0.22 μM filter, concentrations D- and L-α-amino-ε-caprolactam in the samples were determined by HPLC using the protocol as described in Example 1. Finally, monocultures that formed L-α-amino-ε-caprolactam from the D-α-amino-ε-caprolactam substrate were sent to the NCIMB for strain determination. After removal of all particulates by centrifugation followed by filtration through a 0.22 μM filter concentrations D- and L-α-amino-ε-caprolactam in the samples

反応条件: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

実験手順: Then 100 mg of these lyophilized cells from each monoculture were added to a reaction mixture consisting of 10 mM HEPES-NaOH buffer (pH 7.7), toluene (2,5 v/v %), pyridoxal-5-phosphate (20 μM), and L-α-amino-ε-caprolactam (2.5 m %). Chemical blanks (with the assay mixture without the lyophilized cells) were used as a negative control. All reaction mixtures (including the blanks) were incubated for approximately 72 hours at 40° C. before the reaction was stopped by the addition of 9 volumes of 1 M H3PO4. After removal of the cells by filtration through a 0.45 μM filter, these reaction mixtures were analyzed by the HPLC method described above to determine the concentration of D- and L-α-amino-ε-caprolactam. Finally, monocultures that formed D-α-amino-ε-caprolactam from the L-α-amino-ε-caprolactam substrate were sent to the NCIMB for strain determination. Nb. no D-α-amino-ε-caprolactam could be determined in the chemical blanks. Then 100 mg of these lyophilized cells from each monoculture were added to a reaction mixture All reaction mixtures ( After removal of the cells by filtration through a 0.45 μM filter these reaction mixtures

反応条件: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

実験手順: Then 100 mg of these lyophilized cells from each monoculture were added to a reaction mixture consisting of 10 mM HEPES-NaOH buffer (pH 7.7), toluene (2,5 v/v %), pyridoxal-5-phosphate (20 μM), and L-α-amino-ε-caprolactam (2.5 m %). Chemical blanks (with the assay mixture without the lyophilized cells) were used as a negative control. All reaction mixtures (including the blanks) were incubated for approximately 72 hours at 40° C. before the reaction was stopped by the addition of 9 volumes of 1 M H3PO4. After removal of the cells by filtration through a 0.45 μM filter, these reaction mixtures were analyzed by the HPLC method described above to determine the concentration of D- and L-α-amino-ε-caprolactam. Finally, monocultures that formed D-α-amino-ε-caprolactam from the L-α-amino-ε-caprolactam substrate were sent to the NCIMB for strain determination. Nb. no D-α-amino-ε-caprolactam could be determined in the chemical blanks. Then 100 mg of these lyophilized cells from each monoculture were added to a reaction mixture All reaction mixtures ( After removal of the cells by filtration through a 0.45 μM filter these reaction mixtures

反応条件: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

実験手順: Then 100 mg of these lyophilized cells from each monoculture were added to a reaction mixture consisting of 10 mM HEPES-NaOH buffer (pH 7.7), toluene (2,5 v/v %), pyridoxal-5-phosphate (20 μM), and L-α-amino-ε-caprolactam (2.5 m %). Chemical blanks (with the assay mixture without the lyophilized cells) were used as a negative control. All reaction mixtures (including the blanks) were incubated for approximately 72 hours at 40° C. before the reaction was stopped by the addition of 9 volumes of 1 M H3PO4. After removal of the cells by filtration through a 0.45 μM filter, these reaction mixtures were analyzed by the HPLC method described above to determine the concentration of D- and L-α-amino-ε-caprolactam. Finally, monocultures that formed D-α-amino-ε-caprolactam from the L-α-amino-ε-caprolactam substrate were sent to the NCIMB for strain determination. Nb. no D-α-amino-ε-caprolactam could be determined in the chemical blanks. Then 100 mg of these lyophilized cells from each monoculture were added to a reaction mixture All reaction mixtures ( After removal of the cells by filtration through a 0.45 μM filter these reaction mixtures

反応条件: temp:120.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

実験手順: To a solution of (R)-lysine HCl salt (100 g, 0.68 mol) in xylene (1 L) was added hexamethyldisilazane (1000 mL, 4.76 mol) and chlorotrimethylsilane (10 mL, 78.8 mmol). The mixture was heated to 120° C. for 24 h, then to 180° C. for 48 h. The solvent was then removed under reduced pressure and the crude material was purified by column chromatography (20:1 to 10:1 CH2Cl2/MeOH) to afford (R)-3-aminoazepan-2-one (I-1a). The solvent was then removed under reduced pressure the crude material was purified by column chromatography (20:1 to 10:1 CH2Cl2/MeOH)