6. AOACSPIFANMethods-2018Awards

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Jing et al.: J ournal of AOAC I nternational V ol. 98, N o . 5, 2015  1399

Figure 1. Representative chromatograms of SRM 1849a (total analysis). Secondary transitions were monitored for both choline and carnitine, including their internal standards (only the primary transitions are given in Table 2014.04D).

( d ) The method is valid for analytical solution concentrations between 50% of WS1 and 10% above WS3. I. Calculations ( a ) For each of the three WS, the software plots each relative response (analyte/internal standard) versus its corresponding working standard concentration to obtain two separate calibration curves for choline and carnitine (two data points for each concentration, one from the beginning of the analysis and one from the end). It applies a linear regression model to the data and obtains an equation for the best-fit line. ( b ) For each sample injected, the instrument measures the response (analyte/internal standard) for choline and carnitine and uses the linear regression equation to calculate the resulting concentration in the analytical solution. ( c ) The concentration in the analytical solution (ng/mL) is multiplied by a dilution factor (DF) to project the results back to the original sample, on a µg/g basis:

additives (recommended by column supplier). After verifying equilibration of the UPLC system, inject the mid-level working standard four times to verify system suitability. RSD of the peak areas from these injections should be <5%. Once system suitability has been established, inject working standards (WS 1–3), followed by a reagent blank, control sample, and samples. Reinject working standards approximately every 4 h (e.g., enough time for 16 samples with analysis cycle time of 15 min). ( a ) The RSD of the four standard injections to prove equilibration prior to run must be <5%. ( b ) Calibration curve residuals must be ≤4%. Samples should be bracketed by two sets of such valid calibration curves. ( c ) A suitable control sample is National Institute of Standards and Technology Standard Reference Material (NIST SRM) 1849a, reconstituted as a normal sample powder (each packet contains about 10 g). A control sample must be run concurrently with every sample set and a corresponding control chart set up. The control chart RSD of the means of choline and carnitine must be <4.0%. H. System Suitability

(1)

C x

= C s

× DF

where C x and C s

is the concentration of the analyte in the product (µg/g) is the concentration in the analytical solution measured

by the instrument (ng/mL).