Multiple testing time points

As a general rule, in vivo assays are more challenging than in vitro assays because the matrices for the samples are more complex. The most common use for in vivo assays is to measure the concentration of NCE dosed into a laboratory animal by collecting multiple sample time points, one can use the analytical results to plot the PK profile of the NCE and also obtain various PK parameters that help determine a test compound s PK properties. Preclinical PK parameters of a test compound are then used to predict its human PK parameters. Another use of in vivo assays is combining the results with pharmacodynamic (PD) observations to perform PK/PD modeling.77 82 PK/PD modeling is an important aspect of new drug discovery because it can be used to predict the exposures and durations required to determine clinical efficacy of a NCE. 

Dissolution indicates the rate-limiting step for compound absorption when drugs are administered orally. The solubility of a pharmaceutical compound represents its maximum concentration in an aqueous buffer. Additional compound will not dissolve above this concentration. The solubility value is often heavily dependent upon pH and temperature and is typically measured at physiologically important pH levels and body temperature. The standards for dissolution testing are determined by the United States Pharmacopoeia (USP). Testing typically requires sampling of a solution at 15, 30, 45, and 60 min for immediate-release products. /./Pl.C is ideally suited for use in conjunction with USP apparatus types I or II and can rapidly analyze multiple time points or replicate samples. 

Dissolution testing has become an important component of the assessment of the quality of solid oral dosage forms and oral suspensions. The basic procedures for these oral dosage forms have been extended to transdermal delivery systems as well. The release rate for modified-release oral dosage forms adds a level of sophistication to the concept of dissolution testing, setting acceptance criteria at multiple time points. 

USP Dissolution Apparatus 1 (basket) and 2 (paddle) are commonly used for immediate-release formulations. USP Apparatus 3 (reciprocating cylinders) is the system of choice for testing extended-release products or a dosage form that requires release profiling at multiple pH levels and time points. Low-dose products may require the use of flow-through analysis or other low-volume test techniques (noncompendial 100- or 200-mL dissolution vessels). Once the apparatus is selected and has been shown to be suitable during method development, no further evaluation of another apparatus is required during validation. 

If observations are made at two points in space, it is possible to check the assumption g(z) = constant by comparing the initial distributions obtained from the two measured values. In general, if observations are made at n different space (or time) points, and an initial distribution g(z) is assumed with m arbitrary parameters, these parameters may be determined from consistency relations among the various observations, and there will be an additional n-m consistency relation to test the validity of the assumptions. Note that each test is actually multiplied by the number of particle size divisions, so that the initial distribution function q n be determined quite accurately if multiple observations are available. 

For each cluster, all combinations are calculated as described above. However, this is done for each experiment (time point) separately. Once all combinations are calculated, a Wilcoxon test is done to distinguish differences between the two different states (in our case control vs. drug). For each cluster, this is done for every calculated combination. Once all Wilcoxon tests for all clusters and all combinations are done, the P values are corrected for multiple testing. The clusters are then ranked according to the lowest P value that was achieved for the respective clusters. 

HPLC methods are preferred if excipients would interfere, if nonspecific detection techniques (mainly in UV) would be used, or when multiple APIs (combination product) are present in a drug product. Since dissolution sample set analysis can be very long due to six samples per bath as well as multiple time points for prohle testing, fast run times are preferred to quickly determine the results. If a fast HPLC method for CU is available, then the identical HPLC method can be utilized for dissolution analysis. 

One packaging (HDPE bottle with induction seal) Multiple packaging Multiple packaging (multiple count bottles, and multiple material blisters) Consumption test (for bottles) Light test (1200 K lux hours) Micro-testing at annual time points (to test the entire packaging operation and to track the changes over time)... 

Multiple exposure studies may include obtaining information on possible enzymatic inhibition or induction effects as well as the possibility of bioaccumulation. In such studies, animals are dosed by gavage (or other appropriate route) for 14 days with one dose, usually the highest anticipated dose sufficient animals are used so that three data points are available at each blood sampling time. Blood samples are taken at multiple time points after dosing and analyzed for test chemical or metabolite. These results are compared to the results of the single- exposure definitive study to determine possible enzymatic inhibition or induction effects and bioaccumulation of the test chemical. 

The assay itself is somewhat longer however, no radioactivity is required in this method and other end points such as immunophenotyping may be included by incorporating into a rat toxicity study at multiple time points. This test also allows the assessment of numbers of viable cells and avoids the complication of spontaneous leakage of detection reagent from target cells. 

An important problem when using statistical tests (Monte Carlo filtering) is to set the P value under which the effect studied is supposed to be statistically significant. Moreover, when a test is performed many times, the multiple test problem arises. In the case of model 1 (135 random input parameters), it is quite likely that, a non-negligible number of input parameters could be identified as important (under classic P values thresholds such as 0.05 or 0.01). In this case, some input parameters are know to be non-influential with no uncertainty, i.e. the solubility limit of the Sn in the near field when the output considered is the dose due to A criterion that has worked from a practical point of view is to take as such a limit the lowest P value associated to anon-influential input parameter. In average, for that appUcation such limiting P value was around 10 . ... 

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