Summary
Type: SPE
Application #: 300510
Catalog#: 730015 - SPE Cartridges, Chromabond, C18 ec, 6ml, PP, 1000mg, 30/pk
Detection:
Conditions: Step 1:
Column conditioning:
force or aspirate 12 ml methanol, 6 ml dist. water, then 6 ml 0.05 N formic
acid through the column
Sample application:
force or aspirate the prepared sample through the column
Column washing:
force or aspirate 8 ml 1 N formic acid/ acetonitrile (3:1, v/v), then 4 ml
dist. water through the column; then dry the column in a stream of nitrogen
Elution:
4 x 1 ml methanol, then evaporate in a stream of nitrogen, redissolve in
t-butyl methyl ether, evaporate again; finally dissolve in 100 μl ethyl
acetate/formic acid conc. (9:1, v/v), heat to 45 °C for 30 min and dry again in
a stream of nitrogen
Step 2: derivatization with BSTFA with subsequent SPE
Derivatization:
Heat sample from step 1 with 0.5 g methoxyamine hydrochloride in 9.5 ml
N,N-dimethylformamide at 45 °C for 30 min and evaporate; redissolve in 50 μl
acetonitrile, 20 μl N,N-diisopropylethylamine and 20 μl
pentafluorobenzylbromide solution (33%), heat 25 min to 45 °C and evaporate;
then add 50 μl BSTFA and react 2 h at 45 °C and overnight at room temperature;
evaporate in a stream of nitrogen and redissolve in 1 ml hexane
Column conditioning:
force or aspirate 2 ml dichloromethane, 2 ml dichloromethane/hexane (1:1, v/v),
then 2 ml hexane through the column
Sample application:
force or aspirate the derivatized sample through the column
Column washing:
force or aspirate 2 ml hexane, 2 ml dichloromethane/ hexane (1:1, v/v), then 2
ml dichloromethane through the column
Elution:
2 x 1 ml dichloromethane/methanol (100:1, v/v), evaporate in a stream of
nitrogen, redissolve in 50 μl heptane
Authors: Wübert J
Source: Anal Chem 69 (1997) 2143 - 2146