Variability of USP Lot P Prednisone Reference Standard Tablets

The United States Pharmacopeial Convention provides reference standard tablets for use in performance verification testing of dissolution Apparatus 1 and 2 (1–3). The performance verification test (PVT) is specified in United States Pharmacopeia (USP) General Chapter Dissolution ‹711› (4) with specific elements provided by USP in its web-based Dissolution Toolkit (5). In the PVT, USP sets acceptance criteria (tolerances) for each tablet tested based on a collaborative study conducted for each lot of Reference Standard (RS). For the assembly to pass, each tablet must pass. The overall approach speaks to a horizontal standard (i.e., the dissolution procedure) and the various ways in which such a procedure is verified based on sound metrologic principles (2, 6).

In recent years, acceptance criteria for prednisone tablets have been criticized as being too wide (6), and the basis for this width has been judged, erroneously, to be the USP Prednisone RS Tablets (7). In response, USP has conducted research on the quality of its Prednisone RS Tablets and on the dissolution procedure itself. The goals of this research have been to understand the extent to which the prednisone tablets contribute to the variability seen in dissolution performance verification testing and to identify critical parameters in the dissolution assembly and procedure that contribute to the variability in results (1–3, 8–14). Of the studies cited, five provided insight into the variability of dissolution results of the USP Prednisone RS Tablets. This article summarizes these findings, adding new data and expanding on some of the data from the original reports.

US Pharmacopeia

Methods

Reanalyses. This paper summarizes data from four of five published studies (2, 3, 12, and 13) and provides additional detail about a fifth study that was reported only briefly (15). To enhance comparability of results across the studies, data were reanalyzed in a consistent manner. As a consequence, some data reported in this article differ slightly from the data originally reported in References 3 and 14. The method of analysis was first to transform the data to the natural log scale because previous work has found that the normality assumption is better satisfied in this scale. Variances, S², are determined from the log-scaled data and then transformed back to the original scale by the log-normal formula,

where %CV equals percent coefficient of variation.

Because this paper focuses on the contribution of the prednisone tablets to variability, the authors are reporting the lowest-level variance available for each study. That is, we are reporting the variance calculated removing whatever other sources of variance (e.g., analyst and assembly) that can be removed depending on the design of the study. The goal is to obtain a variance that includes the variability due to the tablet but with as little else as possible. When the variance is determined by an analysis of variance (ANOVA), the lowest-level variance is that usually termed the residual variance. When the variance is determined directly from the log-scale data, the variance is first determined within each experiment of six tablets and then pooled over experiments in the study. The magnitude of each variance depends on sources of variability—such as the assay—other than the tablets. These sources differ by study and are described for each study.

Study 1. This study was conducted to compare the dissolution variability of USP Prednisone Lot P RS Tablets to that of four commercially available products (13). Two of the products dissolved so quickly that no meaningful comparison was possible. Reference 13 and the present article thus present data only for ranitidine hydrochloride and metformin tablets compared to the USP Prednisone Lot P Tablets.

Study 1 was a single-laboratory study. Each of the three products was tested by two six-tablet experiments on each of three assemblies configured as Apparatus 2 (paddles), each by two analysts, for a total of 12 experiments. Following log transformation, the variance for each set of six was calculated and pooled over the two experiments. The pooled variance was then converted to the intra-experiment %CV as described above. These intra-experiment %CVs include contributions from, at least, the tablets tested, variability among positions in each assembly, the assay, and the position of the tablet when dropped into the vessel. As conducted, position stands for the physical position as well as the particular combination of shaft, paddle, and vessel used at that position. The "combined" results were obtained by a random-effects ANOVA. The following were included as random effects: assembly, position nested within assembly, analyst, and experiment nested within analyst and assembly. The data were analyzed by SAS Proc Mixed [SAS Inc., Cary, NC]. The residual %CVs thus obtained include contributions from the same sources as for the assembly- and analyst-specific %CVs except for position within the assembly.

Study 2. This study was conducted to evaluate the reproducibility of PVT results following the relocation of equipment between different laboratory sites (16). Dissolution experiments with USP Lot P Prednisone RS Tablets were conducted at two sites (Site 1 and Site 2). Equipment from two manufacturers was used for the experiments. Three dissolution experiments (six tablets per experiment) were repeated by the same two analysts at each of the sites. Experiments were performed at Site 1. The testers were then moved to Site 2, a thorough mechanical calibration was performed, and PVT experiments were repeated. Finally, the equipment was moved back to Site 1, the mechanical calibration was repeated, and PVT experiments were repeated. Vessels, paddles, and shafts were serialized and retained the same position in the instrument throughout the experiments. The reported %CVs were pooled across the three experiments as described for Study 1. Contributions to these intra-experiment %CVs are also as described for Study 1.

Study 3. This study was conducted to evaluate the contributions of analyst and assembly to dissolution results using both Apparatus 1 and Apparatus 2 (2). The study focused on intermediate precision within a single laboratory. Five assemblies were used, each by at least four of six analysts. For each analyst–assembly combination and for each of Apparatus 1 and Apparatus 2, there were six experiments, and each experiment was performed using six to eight tablets, depending on the equipment.

The data were first analyzed by a random-effects ANOVA separately for each assembly. Random effects were position, analyst, and experiment nested within analyst. The %CVs reported are from the residual variance from the ANOVAs. These intra-experiment %CVs include contributions as for the Study 1 ANOVA results, namely from, at least, the tablets tested, the assay, and position of the tablet when dropped into the vessel.

Study 4. This study was conducted to evaluate the relationship between vessel dimensions and dissolution results (12). This was a single-laboratory study, and all work was done by one analyst on one assembly configured as Apparatus 2. Three sets of six vessels were evaluated, and each set was obtained from a different manufacturer. For each set, six dissolution experiments were conducted, and each consisted of six tablets of USP Lot P Prednisone RS Tablets. Following log transform, the variance for each set of six was calculated and pooled over the six experiments for a given vessel manufacturer. The pooled variance was then converted to the intra-experiment %CV as described above. As with Study 1, these intra-experiment %CVs included contributions from, at least, the tablets tested, variability among positions in each assembly, the assay, and the position of the tablet when dropped into the vessel.

Study 5. This study was a 28-laboratory collaborative study to determine the Apparatus 1 and Apparatus 2 PVT acceptance criteria for USP Lot P Prednisone RS Tablets (3). Participants were given the then-current Lot O tablets for assembly suitability testing and two sets of blinded tablets, one of which was Lot O and the other Lot P. Results reported here are for the two sets of blinded tablets. Results from this study, conducted in 2005, are compared with those obtained for Lots N and O in a collaborative study conducted in 2003.

Each laboratory was instructed to run two experiments of six tablets each. For each apparatus, two analysts were told to perform experiments using different equipment. After dropping experiments as outliers for failure to follow protocol or failure of assembly suitability (see Reference 3 for explanations), 38 experiments for Lot P using Apparatus 1 and 40 using Apparatus 2 were available for statistical analysis.

These data were analyzed in a random-effects ANOVA similar to that for Studies 1 and 2. Random effects were laboratory, analyst, and experiment within analyst. The %CVs reported are from the residual variance from the ANOVAs. These intra-experiment %CVs include contributions from, at least, the tablets tested, variability among positions within the assemblies, assay, and position of the tablet when dropped in the vessel. The data shown in Figure 1 are based on pooling the intra-experiment results for each laboratory, as done for Study 1.

Figure 1. Lot P intra-experiment %CVs from Study 5.

Results

Intra-experiment variability results from Study 1 are shown in Table I. For the USP Prednisone RS Tablets, the %CVs were consistently at or below 5% and had a combined value of 3.6%. The %CVs for the two commercially available tablets were much higher (ranitidine hydrochloride = 13.9% and metformin = 12.8%). Results from Study 2 are shown in Table II. Variability differs by assembly and shows fairly consistent results across site and analyst. Results from Study 3 are shown in Table III. In this study, again variability differed by assembly, even though assemblies alpha, gamma, and epsilon of Study 1 were the same as in Study 3. By switching the vessels used in assembly alpha with those in assembly gamma, analysts found that the high variability in assembly alpha was due to the vessels (2). Of note, the authors did see some low %CV for Apparatus 2 corresponding to those of Table I. Eight of 24 assembly–analyst combinations (before switching vessels) had a %CV <6% for Apparatus 2 (data not shown). Study IV's intra-experiment CVs for the vessels from the three manufacturers are shown in Table IV. Results vary considerably by vessel manufacturer.

Table I: Intra-experiment variability of USP Lot P Prednisone and commercial tablets (Apparatus 2, Study 1).

Table V shows the results from Study V for the intra-experiment %CV combined across all participating laboratories. Results remain fairly consistent across the three lots. Table V does not show the variability among laboratories in the %CV. This is shown in Figure 1 for Lot P. In Figure 1, the median values are 8.0% for Apparatus 1 and 6.5% for Apparatus 2. That is, half the laboratories had an intra-experiment %CV with Apparatus 2 of no more than 6.5%. The larger values of Table V are a result of the inclusion of the results from the laboratories with the larger values at the top of Figure 1, many of which are greater than 10% and thus are considered highly variable (16).

Table II: Intra-experiment %CV (Apparatus 2, Study 2).

Discussion

The intent of the USP PVT for nonsolution orally administered drug products is to ensure continuing equivalence between the clinical trial material on which safety and efficacy conclusions were made and the manufactured article following approval. US regulatory approaches do not require postapproval reconsideration of bioequivalence for drug products, barring postapproval change. Thus, reliance on the USP performance test becomes a key, if not the sole, means of ensuring consistency in drug product performance both within and among manufacturers over many years of drug product manufacture.

Table III: Intra-experiment %CV (Apparatus 2, Study 3, using six analysts)*.

The dissolution procedure described in Dissolution ‹711› requires an assembly that allows a kinetic measure of drug release over time. Combining effects from the analyst and analytic procedure, the experimental study of drug product dissolution is performed at batch release and also as part of stability studies. The assembly is a complicated mechanical device with many factors that can influence results. In part, the complexity results from the need to provide some sense of surrogacy for the in vivo condition. For this reason, USP emphasizes the importance of a periodic PVT together with careful mechanical calibration to ensure that the combined experimental study yields consistent results. General Chapter ‹711› has been revised, and the term Apparatus Suitability Test has been replaced with the term Performance Verification Test (1). Although USP reference standard tablets have been referred to as calibrators, this is a misnomer. The tablets are not, and cannot, be used for calibration.

Table IV: Intra-experiment %CVs, assembly epsilon (Apparatus 2, Study 4, using two analysts)

The recent FDA draft guidance for industry attributes the current wide acceptance criteria for USP's PVT to the quality of the USP Prednisone RS Tablet (6). The studies summarized here addressed this issue by determining the contribution to variability that may be attributed to the tablets. Across the studies, one notes that the intra-experiment %CV for USP Lot P Prednisone RS Tablets is consistently below 6–9%, sometimes much below (see Tables I and II). Because the tablets remain relatively constant across the studies and laboratories of Study 5, variability in the %CVs probably is due to factors other than the tablets. If the USP laboratory can achieve 4–5% CV with these prednisone tablets, these numbers reflect the variability contribution from the tablets together with the assay because that cannot be separated out.

Table V: Intra-experiment %CVs from collaborative studies for USP prednisone tablets (Study 5).

The original Lot M of the 10-mg USP prednisone tablet was designed to reproduce FDA's National Center for Drug Analysis #2 10-mg prednisone tablets used for dissolution testing in FDA laboratories. It is of some interest then, that FDA reports for its prednisone tablets a residual variability of a magnitude similar to that seen in the USP laboratory for USP prednisone Lot P, namely 4–5% (17).

In summary, USP Lot P Prednisone RS Tablets are not a major source of variability in dissolution performance verification testing. As is most evident in Figure 1 and Reference 3, results vary considerably between laboratories. We also see results that vary by assembly (see Tables II and III) and by vessels (see Table IV). In part, this is the reason there is a PVT for dissolution. USP continues its efforts to identify sources of variability in dissolution and will update its Dissolution Toolkit to help all dissolution laboratories improve results (5).

Walter W. Hauck, PhD,* is a senior scientific fellow, Gang Deng, PhD, is group leader, Maria J. Glasgow, PhD, is a scientist IV, Mark R. Lidell, PhD, is a chemist III, Pallavi Nithyanandan, PhD, is a chemist III, and Roger L. Williams, MD, is executive vice-president and chief executive officer, at United States Pharmacopeia (USP), 12601 Twinbrook Parkway, Rockville, MD 20852-1790, tel. 301.816.8390, wh@usp.org Dr. Williams is also a member of Pharmaceutical Technology's Editorial Advisory Board.

*To whom all correspondence should be addressed.

References

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4. USP 31–NF 26, Dissolution ‹711›, US Pharmacopeial Convention, Rockville, MD, 2008, pp. 267–274.

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