SimultaneousDetermination of SevenWater SolubleVitamins inProducts byLC-MS/MS
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Performance vs. SMPR
Precision
Methodprecision and accuracy for themethod is presented in table 4. Sevenmatrices designed for theSPIFAN
initiativewere evaluated induplicate over sixdays of analysis. To supplement that data set, data from four
additional products are included. These datawere generated in support of an in-housemethod validation effort.
For these samples datawere collected induplicate over eight dayswith two analysts andon two instruments.
Of the eleven fortified formulas tested allmatricesmet or exceeded theRepeatability (RSD
r
) criteria of the
appropriate SMPR except for the analysis of thiamine in soybased infant formula.
Recovery (Accuracy)
TheSMPR for biotin requires a recoveryof 90-110% for concentrations greater than1µg / 100 g, and80-120%
for concentrations greater between 0.1 and 1µg / 100g. The SMPRs for folic acid andpantothenic acid require a
recoveryof 90 to110%over the range of the assay. Table 5 shows the recoveryof all seven compounds spiked
into theSPIFANmilkbased infant formula placeboproduct. Each level was fortified in triplicate and analyzed
on a single dayof analysis. Fortification rateswere targetedbased on the analyticalmethod’s calibration range.
The low spike represents approximately2X the low calibration standardwhile the high standard is approximately
90% of the high calibrant. All overspike analyses for all compoundsmet the 90 to110%SMPR criteria.
Specificity
As part of themethod feasibility four placebo productswere analyzed in duplicate on sixdays of analysis. No
interferingpeakswere observed for any analysis in anyof the fourmatrices. Additionally a reagent blankwas
analyzed in duplicate on eachdayof analysis. These samples included the stable isotope internal standard, and all
reagents included in the sample preparation. No peak for the native compoundwas observed that represented
more than 10% of the lowest calibrant on anydayof analysis.
Method specificity is further verified by themonitoringof twoproduct ions for both the native and internal
standard. An ion ratio for the compound is establishedusing the calibration standards analyzed during that days
analysis. All analyte peaksmust demonstrate an ion ratio consistent with the standards. A tolerance for the ion
ratio analysis has been established for each analyte based on the overall systemperformance. Any component
failing the ion ratio for either the native compound or the internal standard is flagged to identify a possible
integration error or a possible co-eluting interferent. No analysis of biotin during the feasibility analysis failed
the ion ratio analysis.
BIO-03
FORWORKINGGROUP/ERPUSEONLY
DONOTDISTRIBUTE
