Liquid chromatographic/electrospray mass spectrometric determination (LC/ESI-MS) of chelerythrine and dihydrochelerythrine in near-critical CO(2) extracts from real and spiked plasma samples.
Two polar benzo[c]phenanthridine alkaloids, chelerythrine (CHE) and dihydrochelerythrine (DHCHE), were extracted at 35 degrees C and 10MPa (15MPa for real samples) from real and spiked plasma samples with acceptable recoveries (95.1% and 81.0%, respectively) using near-critical CO(2) modified with aqueous (1:1, v/v) methanol. The alkaloids were quantified by a liquid chromatographic/electrospray mass spectrometric (LC/ESI-MS) method on a Zorbax SB-CN column (75mmx4.6mm, 3.5mum particle size) using methanol (organic phase) and 50mM ammonium formiate (aqueous phase) as a mobile phase. A linear gradient 0-1min, isocratic at 60% organic phase (v/v); from 1.0 to 7.0min, 60-71% organic phase (v/v); from 7.0 to 18.0min, 71-60% organic phase (v/v) was applied. The limit of detection was 1.22ng (3.50pmol) for CHE and 0.95ng (2.72pmol) for DHCHE per 1ml of the sample. The linearity of the calibration curves was satisfactory as indicated by coefficients of determination 0.9979 and 0.9995 for CHE and DHCHE, respectively. Repeatability and intermediate precision (average R.S.D.s) were 1.0-1.5%, accuracy was in the range 99.7-100.3%. Average recovery was 100.1% for both, standard solutions and spiked plasma extracts. Three samples of real rat plasma were extracted and analysed to test the method.
Klejdus
Borivoj
B
Department of Chemistry and Biochemistry, Mendel University of Agriculture and Forestry Brno, Zemedelská 1, CZ-613 00 Brno, Czech Republic.
Lojková
Lea
L
Kosina
Pavel
P
Ulrichová
Jitka
J
Simánek
Vilím
V
Kubán
Vlastimil
V