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Thompson et al.:

J

ournal of

AOAC I

nternational

V

ol.

98, N

o.

6, 2015 

1715

the standard’s nominal concentration). The inclusion of a PB

(run as a sample; its measured concentration must be <1/2 of

the lowest calibration standard), a duplicate sample (relative

difference within 10% for Cr, 7% for Se, and 5% for all other

elements), and known reference materials serving as control

samples (recovery check within control or certified limits) are

mandatory for good method performance. If any of these QC

checks fails, results should be considered invalid.

(c) 

The order of analysis should be calibration standards,

followed by rinse, blank check (PB run as a sample), check

standard, control sample, sample, sample duplicate (up to

10 samples), and finally a repeated check standard.

G. Calculations

Sample concentrations in ng/g are automatically calculated

by the software using a nonweighted least-squares linear

regression calibration analysis to produce a best-fit line:

= +

a blank

Y x

Note that for the Agilent software used in this work, the

sample blank is identical to the Cal Blk and is essentially zero

because high purity reagents are used.

The analyte concentration in the sample is then calculated:

= − ×

x y blank

a

DF

where

x

= analyte concentration (ng/g);

y

= analyte to ISTD

intensity ratio, which is the measured count of each analyte’s

standard solution data point in the calibration curve divided by

the counts of the ISTD at the same level; similarly, the blank =

analyte to ISTD intensity ratio, which is the measured count of

the blank standard solution data point in the calibration curve

divided by the counts of the ISTD at the same level as the blank

standard solution;

a

= slope of the calibration curve (mL/ng);

and DF = volume of the sample solution (mL) divided by

sample weight (g).

H. Method Validation

This method has undergone a thorough single-laboratory

validation (SLV) using AOAC guidelines to probe its linearity,

LOQ, specificity, precision, accuracy, and ruggedness/

robustness. Accuracy has also been affirmed by comparison

to ICP-atomic emission spectrometry (AES) results generated

in the authors’ own laboratory. In addition, reproducibility

was estimated during a limited multilaboratory testing (MLT)

study employing six laboratories and four different ICP/MS

instruments. Both the SLV and MLT results are summarized in

a concurrent publication (6).

Results and Discussion

Specificity

The specificity of the method was determined using a single

element standard at 50 mg/L for each analyte and checking for

apparent signal from the other analytes. None of the standards

produced a response above the PLOQ for any of the other 11

analytes (data not shown), demonstrating that each response is

specific for that analyte. The ISTDs were not tested since they

are used at a low concentration of 50 μg/L.

Linearity

Linearity was demonstrated by analyzing various independent

standards (made from the same stock) as samples against

the normal calibration curve. Linearity standards at nine

concentrations of each analyte spanning the range from 50%

of the lowest calibration standard to 50% above the highest

calibration standard were analyzed twice on each of 3 days using

freshly made standards each day. The means of all six analyses

are reported in Table 2. At the lowest level, 50% of the lowest

calibration standard, all analytes demonstrated acceptable

agreement (95–105%, with rounding) with the nominal value.

Therefore, 50% of the lowest calibration standard concentration

is set as the PLOQ. Overall, the recoveries varied from 91 to

107%, and RSDs varied from 0.3 to 9.3%. The recoveries were

nearly all within a desired 95–105% range, though there are no

specific criteria in the SMPR for linearity. The only elements that

presented any linearity issues were P and Fe, which were routinely

under-recovered (P) or over-recovered (Fe) by about 5–6% across

the calibration curve. Possibly, the linearity could be improved

by adjusting some factors for the analysis of these elements, as

they both have relatively low mass with significant background

interferences that must be handled by the CRC. In practice, no

accuracy issues were observed except for some apparent bias in

P results relative to SRM 1849a (

see

below). Typical correlation

coefficients were 0.9995 or better for all analytes.

LOQ

The PLOQ values from the linearity experiment were

converted from a solution concentration (mg/L) to a weight

basis (mg/100 g for a typical dilution of 1.0 g RTF to 50 mL)

and compared to the SMPR (

see

Table 3). The PLOQs meet

the SMPR for all elements except Fe, Cu, and Mn. In these

cases, the test portion size could be increased to 2–3 g RTF to

improve the PLOQ 2–3-fold lower. The lowest concentrations

of Mn, Cu, and Fe found in the SPIFAN matrixes were

150 ng/g (0.015 mg/100 g), 580 ng/g (0.058 mg/100 g), and

14000 ng/g (1.4 mg/100 g), respectively, all in the SPIFAN

control milk. SMPR for LOQ for Mn, Cu, and Fe are 0.001,

0.001, and 0.01 mg/100 g, respectively, at least 10-fold lower

than observed values.

Precision

SPIFANmatrixes were tested on 8 days (including two analysts

and two instruments) in duplicate, and the results are summarized

in Table 4. The SMPRs require RSD

r

to be ≤5% in all 11 matrixes.

All analytes in all matrixes meet this criterion for the within-day

duplicates (data not shown), typically in the 1–2% range. This

requirement is built into the method due to the criterion that

duplicate results must agree to within 5%. When considering

intermediate reproducibility precision (among days/analysts/

instruments, but in a single laboratory), of the 12 elements and

11 matrixes, there are 11 instances of RSD

iR

>5%. Ten of these

are for the ultratrace elements, Mo and Cr, and there is one

instance for Ca in Adult RTF with high fat. The Adult RTF with

high fat matrix has since been shown to be unstable and perhaps

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