NADPH → 3-hydroxy-L-kynurenine
反応式
C1N=C(N)C2N=CN([C@@H]3[O:14][C@H](COP(OP(OC[C@H]4O[C@@H](N5C=C(C(N)=O)CC=C5)[C@H](O)[C@@H]4O)(O)=O)(O)=O)[C@@H](O)[C@H]3OP(O)(O)=O)C=2N=1.[CH:49]1[CH:50]=[CH:51][C:52]([NH2:63])=[C:53]([C:55]([CH2:57][C@H:58]([NH2:62])[C:59]([OH:61])=[O:60])=[O:56])[CH:54]=1>>[CH:49]1[CH:50]=[C:51]([OH:14])[C:52]([NH2:63])=[C:53]([C:55]([CH2:57][C@H:58]([NH2:62])[C:59]([OH:61])=[O:60])=[O:56])[CH:54]=1参加物
3
実験手順
The reaction mixture contained: 75 μl of suspended homogenate; 100 μl of substrate solution containing 50 mM K-phosphate buffer pH 7.5, 2 mM MgCl2, 0.4 mM NADPH, 50 μM L-kynurenine (final concentration), and 25 μl of different concentrations of inhibitor solutions. The reaction was stopped by addition of 200 μl of 1 M HClO4 after 60 min incubation. L-3-hydroxykynurenine formed was quantified by HPLC with coulometric detection at a working voltage of +0.2 V. The column was a 10 cm C18 reversed phase (3 μm. The mobile phase consisted of 950 ml distilled water, 20 ml acetonitrile, 9 ml triethylamine, 5.9 ml phosphoric acid, 100 mg sodium EDTA and 1.5 g heptanesulfonic acid. The flow rate was 1 ml/min.
後処理
- concentrationconcentration), and
- concentration25 μl of different concentrations of inhibitor solutions
- addition