On-line SPE sample treatment as a tool for method automatization and detection limits reduction: Quantification of 25-hydroxyvitamin D3/D2
- Resource Type
- Authors
- Kalliopi Pazaitou-Panayiotou; Evangelia Bekou; Victoria F. Samanidou; Andreas Tsakalof; Dimitrios Palaiogiannis
- Source
- Journal of Chromatography B. 1043:219-227
- Subject
- Detection limit
Chromatography
Chemistry
Six port
Sample (material)
010401 analytical chemistry
Clinical Biochemistry
Analytical chemistry
030209 endocrinology & metabolism
Cell Biology
General Medicine
01 natural sciences
Biochemistry
0104 chemical sciences
Analytical Chemistry
Reduction (complexity)
03 medical and health sciences
0302 clinical medicine
Gradient system
Solid phase extraction
Total Vitamin D
- Language
- ISSN
- 1570-0232
The development and approbation of new, automated UHPLC-DAD method for the quantification of 25-hydroxyvitamin D3/D2 (25OH-D3/D2) metabolites in plasma/serum for the evaluation of patient's vitamin D status are presented. The method was developed on the Ultimate 3000 UHPLC dual gradient system supplied with the on-line SPE-concentration column coupled through six port switching valve to analytical column. This configuration and materials selected enable large volume sample injection (500μL) and on-line sample preconcentration, clean up and subsequent selective metabolites transfer onto the analytical column. The new method abrogates main conventional time consuming and error source off-line steps of analysis and thus simplifies analysis. The large volume injection increases the sensitivity of instrumental analysis by about ten-fold on-line pre-concentration of metabolites. The instrument response is linear (R>0.99) in the investigated concentration range 10-100ngmL-1 which covers all the possible vitamin D status from serious deficiency (