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Quality Assurance & Calibration Methods: Testing For A Supiciously High (Or Low) Result
Quality Assurance & Calibration Methods: Testing For A Supiciously High (Or Low) Result
Anal Chem 9/26/2016
To judge whether an extremely, and hence suspiciously high (or low) value in a set of measurements is an
outlier, we have a choice of three different tests:
If the standard deviation is well known beforehand from N > 20 previous measurements (hence s ≈ σ), we can compare
the difference between the suspected value and the mean of all data in the set with the critical Z-value.
If the standard deviation is not known beforehand, but rather is computed from the same limited set, the suspicious value
included, we use the Grubbs' test.
For a small numer of observations (N < 10), we can also apply Dixon's Q-test.
The t-test can not be used for outlier testing: This is because, with increasing separation of the suspected
outlier, the standard deviation increases even more rapidly, such that the t-value becomes even smaller and
never exceeds a critical test value. It is for this reason that the Grubbs and Dixon tests have been
designed. Because they are based on fewer measurements, Grubbs and Dixon are more severe than the Z-test.
Although the 5% level is usually applied in statistical tests, 1% is recommended for outlier testing, to avoid rejecting data
too readily.
Rejection becomes more acceptable if we can propose a possible reason why a particular measurement does not belong to
the same population as the others: have we switched samples, used the wrong reagent…..?
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GIST Anal Chem 9/26/2016
Quality assurance: What we do to get the right answer for our purpose.
The right answer should have sufficient accuracy & precision to support subsequent decisions.
There is no point in spending extra money to obtain a more accurate or more precise answer if it is not
necessary.
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GIST Anal Chem 9/26/2016
Specifications: States how good the numbers need to be & what precautions are required in
the analytical procedure.
Sampling requirements.
Accuracy & precision.
Rate of false results.
Selectivity & sensitivity.
Acceptable blank values.
Recovery of fortification.
Calibration checks.
Terms in specifications
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Suppose you must certify that a contaminant in drinking water is below a legal limit.
False positive: The concentration exceeds the legal limit when, in fact, it is below the limit.
False negative: The concentration is below the limit when it is actually above the limit.
In choosing a method,
Selectivity (specificity): The ability to distinguish analyte from other species in the sample.
Sensitivity: The capability of responding reliably & measurably to changes in analyte concentration.
Blanks: Account for interference by other species in the sample & for traces of analyte found
in reagents used for sample preservation, preparation, & analysis.
Method blank: A sample containing all components except analyte & it is taken through all steps of the
analytical procedure.
Reagent blank: Similar to a method blank, but it has not been subject to all sample preparation procedures.
Field blank: Similar to a method blank, but it has been exposed to the site of sampling.
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GIST Anal Chem 9/26/2016
Spike recovery
: Check the matrix effect on the response to analyte
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Performance test samples (quality control samples or blind samples): Samples of known
composition provided to the analyst as unknowns.
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GIST Anal Chem 9/26/2016
Standard operating procedures (SOP): State what steps will be taken & how they will be
carried out.
Adhering to these procedures guards against the normal human desire to take shortcuts based on false
assumptions.
Assessment: The process of (1) collecting data to show that analytical procedures are
operating within specified limits & (2) verifying that final results meet use objectives.
Documentation is critical for assessment.
Standard protocols: Provide directions for what must be documented & how the documentation is to be
done.
Control chart: A visual representation of confidence intervals for a Gaussian distribution (Box 5-1).
2. Method Validation
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Method validation: The process of proving that an analytical method is acceptable for its
intended purpose.
Specificity, linearity, accuracy, precision, range, limit of detection, limit of quantification, & robustness.
Linearity: How well a calibration curve follows a straight line, showing that response is
proportional to the quantity of analyte.
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Square of correlation coefficient, R2: ( xi x)( yi y )
R =
2
( xi x)2 ( yi y)2
The y-intercept of the calibration curve (after subtraction of the blank) should be close 0.
Range: The concentration over which linearity, accuracy, & precision are all acceptable.
Robustness: The ability of an analytical method to be unaffected by small, deliberate
changes in operating parameters (e.g., solvent composition, pH, buffer concentration,
temperature, injection volume, & detector wavelength in a chromatographic method).
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GIST Anal Chem 9/26/2016
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GIST Anal Chem 9/26/2016
Example
: Detection limit
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GIST Anal Chem 9/26/2016
3. Standard Addition
: Matrix & matrix effect
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Standard addition
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Standard addition (“Spiking” of the sample): Known quantities of analyte are added to the
unknown.
From the increase in signal, we deduce how much analyte was in the original unknown.
Requires a linear response to analyte.
As in titrations, higher precision can be achieved when standards are added by mass, instead of volume.
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GIST Anal Chem 9/26/2016
Standard addition
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Dilution factor
: Ratio of final concentration to initial concentration
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V V
[X]f [X]i O [S]f [S]i S
V V
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GIST Anal Chem 9/26/2016
Example
: Standard addition
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V 5.00
[S]f [S]i s (2.08 M) 0.104 M
V 100.0
V 95.0
[X]f [X]i O [X]i 0.950[X]i
V 100.0
Graphical procedure
: Successive standard addition to a single solution
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V I V
I S X I X X [S]i S
VO [X]i VO
Function to plot Function to plot
on y -axis on x -axis
x-intercept: –[X]i
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GIST Anal Chem 9/26/2016
Graphical procedure
: Standard addition with constant total volume
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x-intercept: –[X]f
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GIST Anal Chem 9/26/2016
4. Internal Standards
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Internal standard
A known amount of a compound, different from analyte, that is added to the unknown.
Signal from analyte is compared with signal from the internal standard to find out how much analyte is
present.
Assumption: Relative response to analyte & standard remains constant over a range of concentrations.
Where to use?
Analyses in which the quantity of sample analyzed or the instrument response varies slightly from run to
run.
Sample loss can occur during sample preparation steps prior to analysis: Add to the unknown prior to any
manipulations, then the ratio of standard to analyte remains constant because the same fraction of each is
lost in any operation.
Response factor, F
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Response factor: Relative response of a detector to analyte (X) & internal standard (S).
If the detector responds equally to standard & analyte: F = 1.
If the detector responds twice as much to analyte as to standard: F = 2.
If the detector responds half as much to analyte as to standard: F = 0.5.
AX A
F S
[X] [S]
Chromatographic separation of
[X] & [S]: The concentrations of analyte & standard after mixing. unknown (X) and internal standard (S).
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GIST Anal Chem 9/26/2016
Example
: Using an internal standard
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Preliminary experiment:
A solution containing 0.083 7 M X & 0.066 6 M S: AX = 423 & AS = 347.
Analysis of the unknown:
10.0 mL of 0.146 M S were added to 10.0 mL of unknown, & the mixture was diluted to 25.0 mL:
AX = 533 & AS = 582.
Unknown?
Find the F first.
AX A 423 347
F S F F = 0.9700
[X] [S] 0.083 7 0.066 6
Example
: Using an internal standard (cont.)
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AX A 533 582
F S 0.0970 0 [X] = 0.057 21 M
[X] [S] [X] 0.058 4
Because X was diluted from 10.0 to 25.0 mL, the original concentration of X in the unknown:
25 .0
(0.057 21 M) 0.143 M
10.0
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