Pretreatment of plasma samples by a novel hollow fiber centrifugal ultrafiltration technique for the determination of plasma protein binding of three coumarins using acetone as protein binding releasing agent.
A novel and practical sample pretreatment method based on hollow fiber centrifugal ultrafiltration (HFCF-UF) was developed to determine plasma protein binding by using HPLC. The samples for analyzing unbound and total concentrations could be prepared in parallel simultaneously by the same device. It only required centrifugation for a short time and the filtrate could be injected directly for HPLC analysis without further treatment. Coumarins were selected as the model drugs. Acetone was chosen as the releasing agent to free the binding drug from the drug-protein complex for the total drug concentration determination. Non-specific bindings (NSBs) between the analytes and hollow fiber membrane materials were investigated. The type and volume of protein binding releaser were optimized. Additionally, centrifugal speed and centrifugal time were considered. Under the optimized conditions, the absolute recovery rates of the unbound and total concentrations were in the range of 97.5-100.9% for the three analytes. The limits of detection were in the range of 0.0135-0.0667μgmL(-1). In vitro plasma protein binding of the three coumarins was determined at three concentrations using the validated method and the relative standard deviations (RSDs) were less than 3.4%. Compared with traditional method, the HFCF-UF method is simple to run, no specialized equipment requirement and is a more accurate plasma pretreatment procedure with almost excellent drug-protein binding equilibrium. Therefore, this method can be applied to determine the plasma protein binding in clinical practice. It also provides a reliable alternative for accurate monitoring of unbound or total drug concentration in therapeutic drug monitoring (TDM).
School of Pharmacy, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang 050017, PR China.
Junmei
J
Li
School of Pharmacy, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang 050017, PR China. Electronic address: shiqingwen11@163.com.
Qingwen
Q
Shi
School of Pharmacy, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang 050017, PR China. Electronic address: jiangye@hebmu.edu.cn.
Ye
Y
Jiang
School of Pharmacy, Hebei Medical University, 361 East Zhongshan Road, Shijiazhuang 050017, PR China.
Yan
Y
Liu
D000096:Acetone / Q000378:metabolism*; D001798:Blood Proteins / Q000378:metabolism*; D002498:Centrifugation; D002851:Chromatography, High Pressure Liquid; D003374:Coumarins / Q000378:metabolism*; D006801:Humans; D011485:Protein Binding; D014462:Ultrafiltration / Q000379:methods*
D001798:Blood Proteins; D003374:Coumarins; D000096:Acetone
Acetone; Coumarins; HPLC; Hollow fiber centrifugal ultrafiltration; Plasma protein binding; Protein binding displacer