Basic Information
Poster Number: 1110-11    
Author Name: Ritu Arora Affiliation: Agilent Technologies
Session Title: Bioanalytical LC-MS I
Event Type: Poster
Event Title: Effect of Hematocrit on Analyte Quantification Using Dried Blood Spot Technology for Pharmaceutical Bioanalysis
Presider(s):   Start Time: ( Slot # 11 )
Date: Tuesday, March 13th, 2012 Location: Red Area on Exposition Floor, Aisles 1300-1500
Keywords: Bioanalytical, Liquid Chromatography/Mass Spectroscopy, Sample Handling/Automation, Sample Preparation

Boguszewski, PaulAgilent Technologies
Hudson, WilliamAgilent Technologies
Yong, BenAgilent Technologies

Abstract Content
The surge in interest in Dried Blood Spot (DBS) technology for supporting pharmaceutical bioanalysis is due to the many advantages it offers over conventional plasma sampling. These include the small volumes of blood samples required, various associated cost and ethical advantages, ease of collection, reduced sample shipping costs, and versatile storage conditions.The hematocrit (Ht or HCT) or packed cell volume (PCV) is the percentage of blood volume that is occupied by red blood cells. It is normally about 45% for men and 40% for women. Its levels change with age, sex, and general health. Changes in blood hematocrit results in changes to the viscosity, high hematocrit levels being the most viscous. This variability in viscosity leads to differences in flux and diffusion properties of blood through different substrates used for DBS sample collection. Consequently, assay bias is affected, i.e. the physical characteristics of DBS samples (spot area), the accurate quantification of analytes within these samples (recoveries), and ion-suppression.

A novel, non-cellulose based dried blood spotting material has been developed and used to investigate the effect of hematocrit levels on some of the elements affecting assay bias. Six pharmaceutical compounds were used for this study representing acids, bases, and neutrals. Hematocrit levels in human blood were adjusted by adding or removing plasma to yield HCT levels of 30, 40, 45, 50, 60, and 70. Spiked blood was aliquoted on DMS cards, and worked up. The compound set was analyzed separately in positive and negative modes LC-MS/MS using either totally porous sub-2Ám or superficially porous sub-3 Ám columns. The area of spotted blood increases with increasing hematocrit levels on non-cellulose based DBS cards, in contrast to the trend seen on cellulose-based substrate available commercially. Data will be presented on the effect of hematocrit on analyte quantification and spot areas, and its overall effect on assay bias.