234 / 258
516
F
eng
et al
.:
J
ournal of
AOAC I
nternational
V
ol
.
100, N
o
.
2, 2017
still migrate as two groups either before or after the caseins.
Caseins were eluted as a single group between light and heavy
chains of immunoglobulins, where almost no major whey proteins
were found (Figure 8).
Linearity
Linearity data were obtained by spiking seven different levels
of whey protein ingredients into a fixed level of SMP. Whey
protein content was designed to range from 20% (no added
whey) to about 80% in the mixtures of whey and skim milk.
The typical spiking scheme is presented in Table 1.
Analyses were performed in duplicate on a single day, as
well as in single analyses on 3 separate days. The relationship
between the amount of added whey protein and the whey
protein-to-casein ratio proved to be linear, and the coefficients
of determination (R
2
) were all higher than 0.9900 (Table 2).
The relationship between the amount of added whey protein
and the percentage of added whey in the total protein proved
to be logarithmic; the R
2
were also all higher than 0.9900
(Table 2). Typical linearity relationships are shown in
Figure 9.
Figure 8. Typical e-gram of SMP.
Table 1. Testing scheme for linearity and accuracy (whey
protein ranging from 20 to 80%)
Ingredients
a
Weight,
mg
DI water,
mL
b
Protein
Lot No.
% mg/mL
SMP
84.6
2.00
38.5
16.3
DY19
WPC35
162
1.00
35.2
57.0 C22JUL15J1
Experiment
c
SMP, μL WPC35, μL
DI
water, μL Total, μL
Whey
content, %
WPCL0
60
0
60.0
120.0
21.2
WPC35L1
60
5
55.0
120.0
37.6
WPC35L2
60
10
50.0
120.0
48.0
WPC35L3
60
20
40.0
120.0
60.4
WPC35L4
60
30
30.0
120.0
67.5
WPC35L5
60
40
20.0
120.0
72.2
WPC35L6
60
60
0.0
120.0
77.8
a
WPC = Whey protein concentrate.
b
DI = Deionized.
c
L0–6 = Levels 0–6.
Table 2. Summary of R
2
and the equations for seven levels
of whey protein in total proteins for the whey-to-casein ratio
and added whey protein as the percentage of total protein
Day 1: Avg. for
two replicates
Day 2:
Single
Day 3:
Single
Day 4:
Single
Whey/casein
R
2
0.9931
0.9990
0.9980
0.9984
Equation
y
= 0.0378
x
+
0.2573
y
= 0.0421
x
+
0.2119
y
= 0.0429
x
+
0.2013
y
= 0.044
x
+
0.2232
Percentage whey added
R
2
0.9961
0.9942
0.9957
0.9928
Equation
y
= 16.909
ln(
x
) – 12.42
y
= 16.564
ln(
x
) – 9.7994
y
= 17.576
ln(
x
) – 13.359
y
= 16.339
ln(
x
) – 8.6141
Figure 9. Typical linearity relationships between the area ratio of whey to casein and measured whey percentage versus whey protein
amount added.
234