反应 #1925204

反应方程式

N-ethylmorpholine

4-carboxybenzaldehyde dimethylacetal

dansyl-lysine

N-ethylmorpholine

Nε-dansyl-L-lysine

→

Nε-dansyl-Nα-(m-formylbenzoyl)-lysine

溶剂

acetic acid

dimethylformamide

dimethylformamide

反应条件

后处理

1. 1
   其他30 minutes
2. 2
   workup.ADDITIONwas added
3. 3
   workup.WAITto continue for a further 4 hours
4. 4
   其他The reaction
5. 5
   洗涤The major fluorescent peak eluting on the gradient (around 40% acetonitrile)
6. 6
   workup.ADDITIONTo this stock solution (600 μl) were added 6 μl M HCl
7. 7
   workup.ADDITIONthe solution was diluted ten-fold with 10−3M HCl
8. 8
   浓缩the concentration to 0.7 mM
9. 9
   其他absorption spectrum with that
10. 10
    浓缩of a solution of dansyl-lysine of known concentration

实验过程

Nε-dansyl-L-lysine (Sigma, 9.5 mg) was dissolved in 100 μl dimethylformamide under gentle warming. N-ethylmorpholine (3 μl) was added, so that the apparent pH, as judged by spotting onto moist pH paper, was between 8 and 8.5. This solution was mixed with a solution of 7.3 mg of the hydroxysuccinimido ester of 4-carboxybenzaldehyde dimethylacetal in 50 μl dimethylformamide. The apparent pH was checked in the same way as before, and adjusted with N-ethylmorpholine (1 μl at a time) if necessary. After 4 hours and 30 minutes (the apparent pH having been checked from time to time and adjusted if necessary), a similar solution of 7.3 mg of the active ester was added and the reaction allowed to continue for a further 4 hours. The reaction, as judged by t.l.c., was then essentially complete. The t.l.c. system was that described in Example 1(c) and the Rf values were approx. 0.3 and 0.6 for dansyl-lysine and the product, respectively. The mixture was then diluted to 1 ml with acetic acid (1%, v/v), centrifuged, and the supernatant subjected to HPLC (system of Example 4(c)). The major fluorescent peak eluting on the gradient (around 40% acetonitrile) was pooled, and the acetonitrile was driven off in a current of air. The product was characterized by measurement of its electrophoretic mobility at pH 6.5 (predicted and observed, 0.5 [mAsp=1.0]). To this stock solution (600 μl) were added 6 μl M HCl, and the solution was diluted ten-fold with 10−3M HCl. This brought the concentration to 0.7 mM, as judged by direct comparison of the u.v. absorption spectrum with that of a solution of dansyl-lysine of known concentration. It was assumed that the optical density of the dansyl chromophore would be approximately the same in the two substances.

来源