654 / 696
Fonterra Co-operative Group
Page 10
Repeatability for the method in SPIFAN kit samples ranged between 1.5
–
5.2% which complies with the
≤
11.0% limit set with the SMPR [2] The HorRat values were within acceptability criteria for repeatability of
0.3
–
1.3 [4]. Intermediate precision ranged between 3.1
–
7.9% with a mean value of 5.5%.
Limits of Detection and Quantitation
The limit of detection (LOD) is the lowest content that can be detected with reasonable statistical certainty.
This appropriate concentration for a spiked blank sample was determined by evaluating multiple samples
spiked at a range of concentrations and establishing which spike concentration gave equivalent signal to
~10X signal to noise. This was determined to be a spike concentration of 2 ng to a 2 g sample. The LOD
was determined from 10 independent analyses of a blank sample (unfortified whole milk powder) spiked with
vitamin D
2
and vitamin D
3
. The LOD and LOQ are calculated as described in equations 5 and 6.
LOD =
x
+ 3 x SD
equation 5
where:
x¯ = mean of replicates
SD = standard deviation of all replicates
LOQ =
x
+ 10 x SD
equation 6
where:
x¯ = mean of replicates
SD = standard deviation of all replicates
The LOD and LOQ for vitamin D
2
are determined to be 0.12 and 0.15 μg hg
-1
(0.013 and 0.016 μg hg
-1
) and
the LOD and LOQ for vitamin D
3
are determined to be 0.16 and 0.25 μg hg
-1
(0.018 and 0.028 μg hg
-1
)
(Tables 6
–
7). The LOD and LOQ for both vitamin D
2
and vitamin D
3
are lower than those defined in the
vitamin D SMPR [2].
Table 6: Limits of detection and quantitation for vitamin D
2
Vitamin D
3
Sample
Powder (μg hg
-1
)
RTF (μg hg
-1
)
Mean
0.102
0.011
Standard Deviation (SD)
0.005
0.001
3 x SD
0.014
0.002
10 X SD
0.045
0.005
Limit of Detection
0.12
0.013
Limit of Quantitation
0.15
0.016
VitD-18 (February 2016)
FOR ERP USE ONLY
DO NOT DISTRIBUTE