Extractables Analysis With No Substitutions

by Patricia Stancati, Sartorius     
November 2003


It would be difficult to validate the manufacturing process of a pharmaceutical formulation without using the actual formulation itself. However, the use of a substitute solution, when performing filter extractables analysis has been suggested as a "workaround" when interfering substances are present in the formulation. Because of the need to justify the use of substitute solution, the availability of the effective sample preparation techniques, and the inherent uncertainty of the results when not using the exact pharmaceutical formulation, the value of using a substitute solution is questionable.


Pharmaceutical formulations, such as those used to make solid drugs, and injectable and infusion solutions must conform to defined quality standards. A series of validation activities are preformed by the pharmaceutical industry to establish the conformance, and to satisfy the requirements of the FDA. The FDA requires that the purity of a pharmaceutical be closely monitored, and that activities, which may compromise this purity, be thoroughly investigated.

Throughout the manufacturing process, a pharmaceutical formulation is exposed to certain conditions that may compromise purity. One such condition is filtration, which under ideal circumstances removes unwanted particles or bacteria from the formulation while allowing the actual pharmaceutical formulation to remain unadulterated. It is possible, however, that chemical interactions between the formulation and the filter may introduce new impurities while removing others. This possibility becomes more critical in the final steps of downstream processing, where impurities may have a greater effect on the product's safety, purity and efficacy.

It is important, therefore, to conduct filter extractables analysis to determine the presence of contaminating substances. If present, further analysis may be required to demonstrate maintained formulation safety in the presence of filter extractables.


Filter manufacturers perform extractables tests on filters using standard "model" solutions such as water or ethanol. The information from these studies assists in identifying potential filter extractables, but does not fully mimic the actual pharmaceutical formulations filtered by their customers.

When pharmaceutical manufacturers perform validations, the FDA prefers process conditions that are representative of the routine production. To ensure that all possible situations are accounted for, the test conditions are usually the "worst" or the most challenged conditions seen during a production run. Such conditions might include elevated temperatures, extended filtration runs, and extremes of pH and chemical concentrations. All of these conditions might maximize the level of filter extractables introduced into a formulation.


If a typical filtration process is performed at room temperature (18- 25°C) for 30 minutes, a suitable extractables test condition may be 30°C for 1 hour. If the product formulation varies from pH 5.0 to 8.0, it may be appropriate to perform extractables analysis with the formulation at both pH extremes. The chemical interaction (pH) of the formulation with the filter is as likely as the physical interaction (length of exposure) to release filter extractables.


Accepted by FDA?

Complete Interaction Profile?

"Worst-Case" Test Condition?

Actual Pharmaceutical Formulation




Substitute Solution

With Justification




The example presented in the previous section is suitable for a simple formulation, such as a buffer or pure solvent, in which filter extractables are easily analyzed. What should be done in the case of a complex solution containing preservatives, protein, or other substances that might mask potential extractables during analysis? For example, if High Performance Liquid Chromatography (HPLC) is the analysis method, a strong-UV absorbing component in the formulation may hinder the ability to detect minute levels of extractables. To eliminate the interference, a substitute solution could be used, however it must be fully representative of the actual solution. In order to prove this adequately, each chemical component of the formulation that is "missing" from the substitute solution must be accounted for. The validation must describe why that chemical compound will not contribute to the level of filter extractables. To predict possible chemical interactions with any amount of certainty, the materials of the filter and polymer construction must be known. Filter manufacturers rarely disclose these sometime-proprietary polymer constituents. Even when all the constituents are known, it may not be possible to completely predict how the solution will interact. For this reason, it is ideal to use the actual pharmaceutical formulation to perform extractables testing.

Figure 1. Before Sample Preparation

Figure 2. After Sample Preparation


In actuality, there is usually no reason the actual pharmaceutical formulation cannot be used. After exposing the filter to the appropriate test conditions, methods to remove the interfering components(s) of the formulation can be employed without removing the extractables. Sample preparation techniques, such as solid phase extraction and liquid/liquid extraction, make it possible to alter the post-extraction formulation so that interfering compounds are removed and any extractables remain. Even low levels of extractables will be visible after appropriate treatment. Figures 1 and 2 show HPLC chromatograms of a sample before and after preparation techniques are applied. The technique removes compounds that elute in the time range possible extractables, but allows extractables to remain.

The presence of an alloy with a lower corrosion resistance than 316L often increases rouge formation. An example of this is a straight chromium stainless steel component in a high purity water system.

In remote possibilities, where a sample preparation technique is not available, minor alterations can be made to the product formulation with adequate cause. The elimination of one component may allow analysis to be performed. Then a justification for the modified solution includes only the elimination of that one component.


Using a substitute solution in place of an actual pharmaceutical formulation in filter extractables testing introduces an inherent uncertainty in the results. When the validation process is being inspected by the FDA, there must be confidence that the procedure used approximated as closely as possible the conditions seen in routine production. In most cases, absolute confidence can be provided by using the actual formulation for the extractable validation. There is no secondguessing, or bracketing of test conditions. There is no leap of faith. In essence, there is no substitute for the confidence inherent in validating the real thing.

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