AOAC RI Chemical Cont. Methods-2018 Awards

668  S chneider & A ndersen : J ournal of AOAC I nternational V ol . 98, N o . 3, 2015

would not produce statistically relevant determinations of MDL and LOQ. The method accuracy generally yielded analyte trueness greater than 90%. In order to determine the extent of correction provided by the combination of extracted matrix calibrants and internal standard correction, participants were requested to concurrently analyze a set of solvent calibrants and a single post-extraction fortified matrix calibrant (QC, 1.0 µg/kg) along with the study samples. Internal standard corrected peak area ratios for the 0.90 ng/g fortified sample results generated by 10 laboratories were converted to concentrations using the three different methods of calibration: ( 1 ) extracted matrix calibration curves (the quantitative method used in this collaborative study), ( 2 ) single point calibration against the post-extraction fortified matrix QC calibrant, and ( 3 ) solvent calibration curve (Table 5). For the single point QC calibration method, the 0.90 µg/kg fortified samples were normalized relative to the 1.0 µg/kg QC calibrant. There was generally good agreement among the three calibration methods for all analytes except for BG, where the post-extraction fortified QC matrix calibrant and solvent calibration curve overpredicted the concentration of the residue level and often led to poorer repeatability. Differences in method performance betweenBGandMG-D5may account for variations in the analyte accuracy for BG. BG quantification was studied in greater detail with respect to internal standard correction and matrix effects in a complementary single-laboratory validation study of this method (18). While the post-extraction fortified QC matrix calibrant produced the highest calculated recoveries generated by the three methods, there was generally little difference between the average recoveries generated by the extracted matrix calibration curves and the solvent calibration curves for the analyteswithmatched isotopically-labeled internal standards (MG, LMG, CV, and LCV). The variance in analyte recovery using solvent calibration curves was also generally the largest. A more extensive investigation of this phenomenon has been reported (18). In that work, method accuracy determined from data collected by a single laboratory for MG, LMG, CV, and LCV was found to be generally comparable regardless of which calibration method was used, as long as internal standard correction was applied. The post-extraction fortified calibrant trueness for BG matched the collaborative study results with enhanced recoveries (129–163%); however, in the single- laboratory validation, very low recoveries were found for BG using solvent based calibrants (0–64%). From the results of both studies, and the procedure described by First Action 2012.25  (8), it is clear that acceptable method trueness for all analytes is achieved only when extracted matrix calibrants with internal standard correction are used for quantitative analysis. Analyte identification was achieved by comparison of peak area ratios of the qualitative:quantification product ion transitions of test samples to the average value of the ratios obtained from extracted calibrant samples (0.25–5 µg/L). These results are summarized in Table 6. Acceptability criteria for both the EU (±20–50%, based on ratio found; 10) and the FDA (±10% absolute; 11) were applied to the data, and the results were compared. It is interesting to note that while there were individual cases where one or the other approach provided higher Qualitative Results

identification percentages, on the whole, the two approaches provided comparable results. Evaluation of retention times for identification revealed two laboratories that had some difficulty in meeting the stricter EU standard (±2.5%) on a total of six or 13 samples, respectively. All samples, however, met the FDA retention time standard (±5%). In general, identification was successful for the overwhelming majority of samples. Blank samples did occasionally meet identification criteria, particularly for CV and LCV, as evidenced by the higher percentage of identifications listed for those blank samples (Table 6). Although these samples did meet the requirement of having signals greater than three times the instrument noise, the calculated concentrations for most blank samples was below 0.05 µg/kg. Thirteen of the 84 blank samples met identification criteria and had a calculated analyte concentration >0.05 µg/kg; calculated concentrations for those individual blank samples are reported in Table 6. Five of those blank samples have analytes with concentrations at or above the MDL for the particular analyte/matrix pair: 0.08 μg/kg LMG in salmon, 0.14 μg/kg CV in salmon, 0.30 μg/kg CV in shrimp, and 0.12 and 0.13 μg/kg LCV in catfish. None of the blank samples have analyte concentrations that exceed the CCα for the analyte/matrix pair. In general, concentrations of the identified analytes in the blank samples are well below the 1 µg/kg level of concern. True false-positive samples may be a result of instrument carryover or trace contamination with ink from commonly used laboratory marking pens. For best results, it is advisable to inject water samples between test samples to identify and minimize interference (8), and to avoid the use of laboratory marking pens when labeling samples. The authors of the first action method proposed that this method could be used as a screening method by estimating the concentration of residues in an unknown sample by comparison to a single point extracted matrix calibrant spiked at 0.5 µg/kg. In that analytical strategy, unknown samples that yielded corrected peak areas greater than those generated for the 0.5 matrix calibrant would require a secondary analysis with a full calibration curve (8). From the results of the 14 participating laboratories, peak area data (internal standard corrected) for each 0.5 µg/kg extracted matrix calibrant was tabulated and compared to the appropriate (matching analyte and matrix) corrected peak area for the 10 blinded unknown samples analyzed by each laboratory. The percentage of blinded samples that yielded peak areas greater than the peak area of the 0.5 µg/kg calibrant is summarized in Table 7. None of the negative control samples yielded peak areas greater than the 0.5 µg/kg calibrant, and all of the 1.75 µg/kg fortified samples had responses greater than the 0.5 µg/kg calibrant. For the 0.42 µg/kg fortified samples, 14% (58 of the 420 analyte measurements; five analytes × three matrixes × duplicate samples × 14 laboratories) yielded peak areas greater than the 0.5 µg/kg calibrant. Of these, 22 of the analyte measurements (5%) yielded peak areas >20% of the 0.5 µg/kg calibrant, corresponding to concentrations of 0.62 to 1.14 µg/kg based on the single calibrant estimation. Table 7 highlights the screen results/analyte; however, it should be noted that one sample often yielded incorrect screen results for more than one residue. For example, catfish at the 0.42 µg/kg level yielded several samples Use for Screening