AUTOMATED SOLID-PHASE EXTRACTION continued Sorbents for QuEChERS
Remove more problematic pigment and lipid interferences than traditional phases
Supel™ QuE Z-Sep
Assay validation For SPE method validation, the linear calibration curve ranged from 5 to 500 ppb (µg/L). To verify the method, DMEM containing 200 ppb of cyclophosphamide and 800 ppb of caffeine was used. One sample was quantified 10 times to assess measuring precision. To determine repeatability precision, 25 samples were analyzed in one day. The limit of detection (LOD) and limit of quantification (LOQ) were determined on the basis of 10 blanks measured on the same day. LOD was calculated based on the signal-to-noise ratio. LOQ is the lowest measurable concentration of the analyte.
To test the within-day precision, 10 samples for the manual procedure and 24 or 96 samples for the automated process were produced and analyzed each day for five days. For the between-day precision, samples were analyzed on five different days. The samples were split into five parts and refrigerated and measured over five consecutive days. Hence, 210 samples were quantified using both manual and automated SPE. Data are presented as mean value (MV%) and relative standard deviation (RSD%).
Results The calibration curve is linear in the range from 5 to 500 ppb (y = 0.0015 · x +
0.0022) with a correlation coefficient of R = 0.998 (Figure 2). To validate the methods, 200 ppb was used in both procedures and set as 100% for final comparison of the manual and automated processes. The manual process was first validated and set as the comparison reference. The between-day precision was averaged at 97.18 ± 3.13 % (Table 2). The LOD was 6.85 µg/L and the LOQ was 13.12 µg/L for the manual process. For the automated process, the LOD was 6.26 µg/L and the LOQ was 6.77 µg/L (Table 3). The measuring precision for the manual process was 91.58 ± 2.61% and 96.09 ± 1.44% for the automated procedure. In addition, the manual sample preparation demonstrated a repeatability precision of 95.26 ± 4.14%; the repeatability precision of the automated process was 97.63 ± 4.39% with the 24-column adapter plate and 74.35 ± 5.89% with the 96-deep-well plate, respectively. The automated process with the 24-col- umn adapter plate had a higher repeatability precision than the manual process, but the opposite was true with the 96-deep-well plate. Possible
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Figure 2 – Calibration curve. Calibration function: y = 0.0015 · x + 0.0022; R2
= 0.998; analysis via HPLC/MS. AMERICAN LABORATORY • 30 • MARCH 2015
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