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1040 A. Introduction
Although standard methods are available from many nationally recognized sources, there may be occasions when they
cannot be used or when no standard method exists for a particular constituent or characteristic. Therefore, method development may be required. Method development is the set of exper-
1. Single-Operator Characteristics
Sum
Difference
(1.30)
Squared
Difference
0.07
0.09
0.0
0.29
0.27
0.09
0.23
0.05
0.0049
0.0081
0.0
0.0841
0.0729
0.0081
0.0529
0.0025
0.49
0.2335
Obtain the unknowns from other personnel in the analysts laboratory using either purchased analytical-grade reagents or standards available from National Institute of Standards and Technology (NIST). If available for the particular constituent, performance
evaluation samples from EPA-Cincinnati are particularly useful.
3. Method Ruggedness
4. Equivalency Testing
Nominal
Variation
10 min
5g
1M
100C
5 min
yes
6.0
12 min
10 g
1.1M
95C
10 min
no
6.5
A or a
B or b
C or c
D or d
E or e
F or f
G or g
A
B
C
D
E
F
G
A
B
c
D
e
f
g
A
b
C
d
E
f
g
A
b
c
d
e
F
G
a
B
C
d
e
F
g
a
B
c
d
E
f
G
a
b
C
D
e
f
G
a
b
c
D
E
F
g
Result
SOURCE: YOUDEN, W.J. & E.H. STEINER. 1975. Statistical Manual of AOAC.
Assoc. Official Analytical Chemists, Washington, D.C.
5. References
varying a factor, find the four results where the factor was
nominal (all caps) and the four where it was varied (all lower
case) and compare the averages of the two groups. For example,
to compare the effect of changing C to c, use results (s u w
y)/4 and (t v x z)/4. Calculate all seven pairs to get
seven differences, which can then be ranked to reveal those with
a significant effect on the results. If there is no outstanding
difference, calculate the average and standard deviation of the
eight results s through z. The standard deviation is a realistic
estimate of the precision of the method. This design tests main
effects, not interactions.
1. Variables
Deviation
Laboratory
1
Result
mg/L
32.7
35.2
36.3
32.6
33.7
33.6
30.6
30.6
32.4
32.6
32.5
33.9
32.4
33.4
32.9
Experimental
xs
From
Known
Known
Amount
mg/L
From Grand
Average
34.71.8
2.0
1.7
33.30.6
0.6
0.3
31.21.0
1.5
1.8
33.00.8
0.3
32.60.8
0.1
4.3
11.6
23.4
32.7
AND
BIAS
Amount
Found
mg/L
CV
(% Standard
Deviation)
Bias
%
4.8
12.2
23.8
33
12.5
10.2
5.4
4.5
11.5
5.6
1.9
0.9
P 3 1 6 1 18
1.3
(x)/n33
s1.5
r 1 (30/18) 2.7 or r 3.
0.4
Send each of five laboratories four concentrations of a compound (4.3, 11.6, 23.4, and 32.7 mg/L) with instructions to
analyze in triplicate using the procedure provided. Tabulate
results as shown in Table 1040:IV (the results for only one
concentration are shown). Because there are no obviously aberrant values (use the method in Section 1010B.4 to reject outliers), use all the data.
Calculate the average and standard deviation for each laboratory; use all 15 results to calculate a grand average and standard
deviation. The difference between the average of each laboratory
and the grand average reveals any significant bias, such as that
shown for Laboratories 1 and 3. The difference between the
grand average and the known value is the method bias, e.g., 33.0
32.7 0.3 mg/L, or 0.9%. The relative standard deviation of
the grand average (1.5 mg/L) is 4.5%, which is the method
precision, and the s for each laboratory is the single-operator
precision.
As noted in Table 1040:IV, the sum of the deviations from the
known value for the laboratories was 1.3, so the average deviation
(bias) was 1.3/5 0.26, rounded to 0.3, which is the same as the
difference between the grand average and the known value.
For all four unknowns in this test, the percentage results indicated
increasing bias and decreasing precision as the concentration decreased. Therefore, to describe the method in a formal statement, the
precision would be given by a straight line with the formula y mx
b; where y is the relative standard deviation, m is the slope of the
line, x is the concentration, and b is the relative standard deviation
at concentration 0. The values found from the collaborative test
are shown in Table 1040:V.
These results indicate that the method is acceptable. However,
concentrations of less than about 10 mg/L require greater care in
analysis.
0.2
sources of error, analyze at least two replicates of each concentration per laboratory;
OperatorsTo determine overall precision, involve at least six
analysts with not more than two from each laboratory;
LevelsIf the method development has indicated that the relative standard deviation is constant, test three levels covering the
range of the method. If it is not constant, use more levels spread
uniformly over the operating range.
If matrix effects are suspected, conduct the test in each medium for which the method was developed. If this is not feasible,
use appropriate grades of reagent water as long as this is stipulated in the resulting statement of method characteristics.
2. Number of Replicates
Calculate the number of replicates after the number of variables to be tested has been determined by using the formula:
r 1 (30/P)
where:
r number of replicates and
P the product of several variables.
4. Reference
1. YOUDEN, W.J. & E.H. STEINER. 1975. Statistical Manual of the
AOAC. Assoc. Official Analytical Chemists, Washington, D.C.