Reproducibility? Ruggedness? Robustness? To many people, all of these terms mean the same thing. But in reality, in the words of a popular children's program, "one of these things is not like the other." An earlier "Validation Viewpoint" column (1) touched briefly on this topic, but this column will explore the topic in a little more detail. So, for the purposes of this discussion, the "R-word" we are most interested in is robustness. However, first we need a few clarifications.

Terms and Definitions

There are two guideline documents important to any method validation process (2,3): USP Chapter 1225: Validation of Compendial Methods; and the International Conference on Harmonization (ICH) Guideline: Validation of Analytical Procedures: Text and Methodology Q2 (R1). While the USP is the sole legal document in the eyes of the FDA, this article draws from both guidelines as appropriate for definitions and methodology.

That is, it is a measure of the reproducibility of test results under the variation in conditions normally expected from laboratory to laboratory and from analyst to analyst. The use of the term ruggedness, however, is not used by the ICH, but is certainly addressed in guideline Q2 (R1) under intermediate precision (within-laboratory variations; different days, analysts, equipment, and so forth) and reproducibility (between-laboratory variations from collaborative studies applied to the standardization of the method). It is also falling out of favor with the USP, as evident in recently proposed revisions to chapter 1225, where references to ruggedness have been deleted to harmonize more closely with ICH, using the term "intermediate precision" instead (4).

Both the ICH and the USP guidelines define the robustness of an analytical procedure as a measure of its capacity to remain unaffected by small but deliberate variations in procedural parameters listed in the documentation, providing an indication of the method's or procedure's suitability and reliability during normal use. But while robustness shows up in both guidelines, interestingly enough, it is not in the list of suggested or typical analytical characteristics used to validate a method (again, this apparent discrepancy is changing in recently proposed revisions to USP chapter 1225 [3]).

Robustness traditionally has not been considered as a validation parameter in the strictest sense because usually it is investigated during method development, once the method is at least partially optimized. When thought of in this context, evaluation of robustness during development makes sense as parameters that affect the method can be identified easily when manipulated for selectivity or optimization purposes. Evaluating robustness either before or at the beginning of the formal method validation process also fits into the category of "you can pay me now, or you can pay me later." In other words, investing a little time up-front can save a lot of time, energy, and expense later.

During a robustness study, method parameters are varied intentionally to see if the method results are affected. The key word in the definition is deliberate. In liquid chromatography (LC), examples of typical variations are

• mobile phase composition;
• number, type, and proportion of organic solvents;
• buffer composition and concentration;
• pH of the mobile phase;
• different columns lots;
• temperature;
• flow rate;
• wavelength;
• gradient variations;
• hold times;
• slope; and
• length.