A Robust, Automated Karl Fischer Titration System - Pharmaceutical Technology

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A Robust, Automated Karl Fischer Titration System
The authors developed a robust, automated system to conduct Karl Fischer moisture assays for lyophilized products.


Pharmaceutical Technology
Volume 33, Issue 11, pp. 52-60

Sample-preparation techniques

All of Genentech's (South San Francisco, CA) lyophilized products are packaged in glass vials. Many configurations, spanning various compositions, fill volumes, and vial sizes, exist, however. The authors considered the techniques as outlined below for preparing samples.

Method A: extraction in the product vial. Mettler-Toledo recommends this technique for handling freeze-dried substances (3). The procedure calls for injecting a known amount of anhydrous solvent (e.g., methanol or other primary alcohols) directly into the unopened sample vial and allowing time for water to be extracted into the solvent. Agitation or sonication can expedite the extraction. An aliquot of the extract is then withdrawn from the sample vial and introduced into the titration cell for analysis.

Method B: drying oven. In this technique, the sample vial is placed into a small oven and heated to desorb the moisture from the contents. An inert, dry carrier gas is fed into the vial, and the effluent is introduced into the titration cell for analysis. The oven temperature setting and total desorption time often depend on the sample. Higher temperatures allow for faster water desorption but can cause the decomposition of certain substances such as sugars, which can yield erroneous moisture results (2).

Method C: direct extraction of crushed sample in titration cell. Genentech has traditionally used this technique. The product vial is uncapped, and the contents are crushed inside the vial using a smooth metal rod. A known amount of powdered sample is then transferred directly into the titration vessel for analysis.

All of the authors' historical data were collected using Method C. The following study was undertaken to determine the comparability between Method A and Method C only. The experimental setup for Method B was not available. Furthermore, many of Genentech's lyophilized products are formulated with large amounts (e.g., as high as 85% wt.) of sugars, which limits drying ovens to low temperatures. Other products contain high amounts of salts and require high oven temperature for efficient moisture desorption. Because it was undesirable to develop a separate method for each product, the authors did not use the moisture-analysis technique that relies on drying ovens.

A Mettler-Toledo DL31 volumetric Karl Fischer titrator with a one-component Hydranal Composite 2 titrant was used in all cases. Sodium tartrate dihydrate was used as the standard to determine the reagent titer at the beginning of each day, as recommended by the instrument manufacturer.

The authors used active lyophilized product configured as a 0.88-mL fill volume in 5-cm3 vials in the study. Two sets of samples, spanning the typical range of moisture normally observed for this product (i.e., 0.5–5% wt.), were tested.

The authors investigated the following two extraction schemes for Method A:

I Introduction of anhydrous methanol into the product vial, 5-min sonication, withdrawal of an aliquot and analysis. This is the instrument manufacturer's recommended procedure.

II Introduction of anhydrous methanol into the product vial, 5-min sonication, 5-min standing, an additional 5-min sonication, withdrawal of an aliquot, and analysis.


Figure 1: Comparison between extraction methods. Squares represent low-moisture samples, and circles represent high-moisture samples. (IMAGE IS COURTESY OF PHILIPPE LAM)
Figure 1 summarizes the results for these experiments. For consistency with historical data, water content is reported as weight percent moisture of the lyophilized product cake. To calculate weight percent moisture values for Method A, the sample vials had to be weighed before extraction, emptied after extraction, cleaned, dried, and reweighed to obtain the actual freeze-dried cake's mass. These extra steps added complexity and lengthened the overall analysis time. Method B would also require following the same procedure.

Comparing results from the two methods, it is clear that Scheme II of Method A extracted more water than the recommended Scheme I, suggesting that Scheme I was not able to fully extract the water from the sample. Yet it is unclear whether Scheme II achieved complete extraction.


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