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F
eng
et al
.:
J
ournal of
AOAC I
nternational
V
ol
.
100, N
o
.
2, 2017
517
Table 3. Spike-recovery data obtained in duplicate for
different levels on 1 day
Spike
level
Integrated area
Measured Theoretical
Recovery,
%
Total whey Casein Whey, %
Whey, %
spiked
Whey, %
spiked
0
90447 431555 21.3
95042 450546 21.4
1
467929 454430 59.0 37.7
39.0
96.6
489937 464188 59.6 38.3
39.0
98.1
2
686816 457990 67.7 46.4
46.2
100.5
699775 481562 67.0 45.7
46.2
99.0
3
823437 448464 72.0 50.7
50.8
99.7
847557 455946 72.2 50.9
50.8
100.2
4
1181386 467897 77.9 56.6
56.5
100.2
1179619 504671 76.6 55.3
56.5
97.8
Table 4. Spike-recovery data obtained in singlet for
different levels on 3 different days
Level
Day
Area
Measured
Theoretical
Total
whey Casein
Whey,
%
Whey, %
spiked
Whey, %
spiked
Recovery,
%
0
1 112590 581315 20.0
2 134644 668598 20.6
3 125910 626821 20.6
1
1 544500 636291 54.5 34.5 32.1 107.6
2 528392 647887 53.3 32.7 31.3 104.4
3 544475 625202 54.9 34.4 33.0 104.2
2
1 632182 609700 59.2 39.2 37.3 105.0
2 654331 605001 60.2 39.6 36.6 108.1
3 698583 633572 60.7 40.1 38.3 104.8
3
1 805608 630879 64.1 44.1 41.4 106.6
2 789708 615699 64.2 43.6 40.8 106.9
3 836902 621176 65.4 44.8 42.3 105.8
4
1 939097 642338 67.2 47.2 44.7 105.6
2 930074 631568 67.3 46.7 44.1 105.8
3 906509 594583 68.1 47.5 45.6 104.3
Table 5. Repeatability and intermediate precision for three
infant formulas
Day
Formula/Replicate
1/1
1/2
2/1
2/2
3/1
3/2
1
61.0
60.8
55.9
57.2
64.4
64.2
2
59.7
59.8
56.5
56.1
64.4
64.7
3
60.5
59.3
56.2
56.1
64.4
65.1
4
60.0
59.9
56.1
56.6
64.2
64.2
5
59.5
58.8
56.4
56.5
64.8
65.2
6
59.3
59.7
57.1
57.8
64.9
64.8
Avg.
59.9
56.4
64.7
RSD
r
, %
0.5
0.8
0.3
RSD
R
, % 1.0
0.8
0.7
Table 6. Repeatability and intermediate precision for SMP
and demineralized sweet whey ingredient
Day
SMP
a
SMP
a
DW
b
DW
b
1
21.7
21.2
95.5
98.6
2
20.1
19.8
98.3
97.9
3
21.8
21.4
97.4
97.6
4
21.6
21.9
97.0
98.3
5
22.2
22.4
98.5
99.2
6
22.1
22.4
98.8
97.9
Avg.
21.7
97.9
RSD
r
, %
1.5
0.9
RSD
R
, %
3.8
1.0
a
Because SMP is not processed like infant formula, a CF of 1.29 was
used.
b
Because whey protein contains almost no caseins, no CF was used.
Table 7. Repeatability and intermediate precision for two
infant formulas
Rep. 1 Rep. 2 Rep. 3 Rep. 4 Rep. 5 Rep. 6 Avg.
SD RSD
Day
Formula 4
1
71.2 70.5 70.7 71.1 71.1 70.8 70.9 0.3 0.4
2
71.0 70.7 71.2 70.7 70.7 70.8 70.9 0.2 0.3
3
70.6 70.4 70.1 70.3 70.5 70.3 70.3 0.2 0.2
Total
70.7 0.3 0.5
Formula 5
1
40.7 41.8 41.6 41.3 41.1 41.7 41.4 0.4 1.0
2
41.7 42.4 42.5 41.2 42.2 42.2 42.0 0.5 1.2
3
41.8 42.0 42.0 42.1 41.9 41.6 41.9 0.2 0.4
Total
41.8 0.5 1.1
Accuracy (Spike Recovery)
Accuracy was evaluated in samples where four levels of
sweet whey ingredient were spiked into the same level of SMP
by comparing the theoretically calculated percentage whey
values with the measured values obtained using equations 1
and 2 (for skim milk, where no whey proteins were added, the
CF was set to 1.29). The results are listed in Tables 3 and 4. The
SMPs and whey protein ingredients used in Tables 3 and 4 are
from different lot numbers.
Precision
Two independent sample preparations were tested on each
day for 6 separate days for the following: three infant formula
samples (shown in Table 5), one SMP sample, and one sweet
whey (demineralized) sample (shown in Table 6). Two other
infant formula samples were tested by preparing and analyzing six
replicates each day on 3 different days using this method (shown
in Table 7). The percentage of whey protein was reported to one
decimal place for individual and averaged replicates. The SD and
percentage RSDs were calculated and also reported to one decimal
place. Data are presented in Tables 5–7. The typical e-grams for
each infant formula sample are presented in Figures 10–14.
The molar attenuation (or molar extinction) coefficient,
reflected as the mass-to-area ratio at 220 nm, is an intrinsic
235