Application Note: Extraction of vitamins B1 and B6 from whole blood using ISOLUTE PLD+ Protein and Phospholipid Removal Plate
21 March 2023

Vitamin B1 (Thiamine Diphosphate, TDP) and B6 (Pyridoxal-5- Phosphate, PLP) in whole blood is measured clinically to help measure various metabolic functions. TDP and PLP are highly water soluble (logP= -5.9, -2.2 respectively) and are extremely light-sensitive. Traditionally, TDP and PLP are measured through fluorescence detection, requiring the use of hazardous reagents for derivatization. This additional derivatization step is also time consuming. In this application note, Biotage describes a simple ISOLUTE® PLD+ protocol for the extraction of vitamin B1 and B6 (TDP/PLP) from whole blood and LC-MS/MS analysis. High reproducible recoveries and low protein and phospholipid coextraction are demonstrated. ISOLUTE PLD+ Protein and Phospholipid Removal plates and columns offer a fast and extremely simple sample preparation approach. Proteins are efficiently removed from the sample using an in-plate, ‘solvent first’, protein crash and filter procedure, which is easily automated.