Bioanalytical High-Throughput Selected Reaction Monitoring-LC/MS Determination of Selected Estrogen Receptor Modulators in Human Plasma: 2000 Samples/Day
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- 作者：Jerry Zweigenbaum and Jack Henion
- 刊名：Analytical Chemistry
- 出版年：2000
- 出版时间：June 1, 2000
- 年：2000
- 卷：72
- 期：11
- 页码：2446 - 2454
- 全文大小：113K
- 年卷期：v.72,no.11(June 1, 2000)
- ISSN：1520-6882
文摘
The high-throughput determination of small molecules inbiological matrixes has become an important part of drugdiscovery. This work shows that increased throughput LC/MS/MS techniques can be used for the analysis ofselected estrogen receptor modulators in human plasmawhere more than 2000 samples may be analyzed in a 24-hperiod. The compounds used to demonstrate the high-throughput methodology include tamoxifen, raloxifene,4-hydroxytamoxifen, nafoxidine, and idoxifene. Tamoxifenand raloxifene are used in both breast cancer therapy andosteoporosis and have shown prophylactic potential forthe reduction of the risk of breast cancer. The describedstrategy provides LC/MS/MS separation and quantitationfor each of the five test articles in control human plasma.The method includes sample preparation employing liquid-liquid extraction in the 96-well format, an LC separation of the five compounds in less than 30 s, and selectedreaction monitoring detection from low nano- to microgram per milliter levels. Precision and accuracy aredetermined where each 96-well plate is considered atypical "tray" having calibration standards and qualitycontrol (QC) samples dispersed through each plate. Aconcept is introduced where 24 96-well plates analyzedin 1 day is considered a "grand tray", and the method iscross-validated with standards placed only at the beginning of the first plate and the end of the last plate. Usingidoxifene-d5 as an internal standard, the results obtainedfor idoxifene and tamoxifen satisfy current bioanalyticalmethod validation criteria on two separate days where2112 and 2304 samples were run, respectively. Methodvalidation included 24-h autosampler stability and onefreeze-thaw cycle stability for the extracts. Idoxifeneshowed acceptable results with accuracy ranging from0.3% for the high quality control (QC) to 15.4% for thelow QC and precision of 3.6%-13.9% relative standarddeviation. Tamoxifen showed accuracy ranging from 1.6%to 13.8% and precision from 7.8% to 15.2%. The lineardynamic range for these compounds was 3 orders ofmagnitude. The limit of quantification was 5 and 50 ng/mL for tamoxifen and idoxifene, respectively. The othercompounds in this study in general satisfy the morerelaxed bioanalytical acceptance criteria for modern drugdiscovery. It is suggested that the quantification levelsreported in this high-throughput analysis example areadequate for many drug discovery and related earlypharmaceutical studies.