Robustness testing

O. In actual practice, this is a robust test, in the sense that in most types of problems it is not sensitive to the normality assumption when the sample size is 10 or greater. 

Rodriguez LC, Garcia RB, Garcia Campana AM, Bosque Sendra JM (1998) A new approach to a complete robustness test of experimental nominal conditions of chemical testing procedures for internal analytical quality assessment. Chemom Intell Lab Syst 41 57... 

As shown in Figure 11, before extensive validation, the performance of the method is evaluated appropriately. Column durability tests, robustness testing for the chromatographic and sample preparation conditions, analytical method evaluation ring tests (AMERTs), method capability assessments, and pre-validation studies are applied to... 

A tentative list of factors that may be investigated in the robustness test is presented in Table 8. This list is not complete and additional factors may be added. The limits for the factor levels are proposals and should be evaluated case by case. The significance of the effects of the factors on the responses such as the resolution of all peak pairs, the tailing factor, retention times, analysis time, etc., is evaluated. 

A tentative list of factors to be evaluated in robustness testing for sample preparation is presented in Table 9. Additional factors can be added. The limits for the factor levels are typical and should be considered as proposals. Responses to be considered are usually related to the quantitative performance of the method. 

For example, if an HPLC system does not pass system suitability because of a capacity factor (k ) that is too low, the results of robustness testing could tell the analyst that the amount of organic can be changed by 10% without affecting the result. If evaluated at the time of validation, the analyst can make the adjustment with no additional validation. If this particular study has not been performed, the analyst has no other choice but to verify the validity of the method at the new condition. 

The analysis time is adjusted according to the worst case results during robustness testing. 

Additives are often used to increase selectivity. They are paramount in chiral separations, but they are also frequently used in non-chiral separations, e.g., cyclodextrins (CDs). In our lab, BGEs with and without a cyclodextrin are part of our generic protocol. Figure 8 demonstrates that although one can more or less predict interaction with the additive from the chemical structures, it is still difficult to predict separation.Batch-to-batch variability and variability between suppliers can be a problem of (chiral) additives and a check of different batches has to be part of the robustness test (e.g., reference 56). If the additive is charged and has one or more pK s around the pH of the BGE, extra care should be taken to control the pH. Alternatively, better robustness might be obtained with another uncharged additive, even if this results in lower resolution. 

AIMS/OBJECTIVES AND STEPS IN A ROBUSTNESS TEST SELECTION OF FACTORS AND LEVELS... 

In biomedical and pharmaceutical analysis, and particularly in the pharmaceutical industry, much attention is paid to the quality of the obtained analytical results because of the strict regulations set by regulatory bodies. Proper method validation demonstrates the fit of an analytical method for a given purpose. In this context, robustness testing has become increasingly important. 

The setup and treatment of results of such a robustness test are discussed in this chapter. All steps of the test are considered. Finally, a literature review and critique of applications of robustness testing of CE methods has been provided. 

The ICH guidelines define robustness as The robusmess of an analytical procedure is a measure of its capacity to remain unaffected by small, but deliberate variations in method parameters and provides an indication of its reliability during normal usage.Thus, the robustness of a method is a measure of its capacity to be transferred to another laboratory without affecting the analytical result. A robustness test is the experimental setup applied to evaluate the robustness of the method. The ICH guidelines also state that One consequence... 

Initially, robustness testing was performed to identify potentially important factors, which could affect the results of an interlaboratory study.Therefore, the robustness test was executed at the end of the method validation procedure, just before the interlaboratory study. Flowever, a method found to be non-robust should be redeveloped and revalidated, leading to a waste of time and money. For these economical reasons, nowadays, method robusmess is verified at an earlier stage in the lifetime of the method, i.e., at the end of method development or at the beginning of the validation procedure. ... 

A robustness test examines potential sources (factors) of variability in one or more responses of the considered method. In pharmaceutical analysis, chromatographic and electrophoretic separation methods are most commonly evaluated. 

Another aim of performing a robustness test can be to define SST limits.The SST limits are then determined based on the experimental data from a robustness test, while frequently they are chosen arbitrarily based on the experience of the analyst. 

FIGURE I Schematic representation of the different steps in a robustness test. 

These different steps are represented in Figure 1, discussed in more detail helow (Sections III-VIII), and illustrated with two examples, taken from the literature. In Section IX, a critical review of some published case studies, describing robustness tests of CE methods, is given. 

Defining and selecting the factors to be examined should be carefully thought through before starting a robustness test. Usually, the factors in a robusmess test are operational or environmental factors. The former are selected from the operating procedure of the method, while the latter are not necessarily specified in this procedure. Those factors, which are most likely to vary when a method is transferred between different laboratories, analysts or instruments, are selected. 

Table 1 gives an overview of potential factors to be considered when performing a robustness test on a capillary electrophoretic (CE) method. In references 6, 9, 17, and 19, lists of factors for high-performance liquid chromatographic (HPLC), gas chromatographic (GC), and/or thin-layer chromatographic (TLC) methods, can be found. 

Table 1 also contains some peak measurement/analysis parameters that might be investigated in a robustness test (see Section III.C). ° However, except from the detection wavelength, these parameters are hardly ever evaluated in a robustness test, even though they can have a large influence on the electropherogram. 

TABLE I Potential Factors in the Robustness Testing of Capillary Electrophoretic Methods... 

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