Assay bioavailability, relative

A very important advantage of the relative bioavailability assay is that experimental parameters may be selected which are easily quantitated. Thus, Tso et al. (11) determined the relationship between dietary calcium concentration (X, g/kg) and... 

Although in vitro GR 175737 is less potent (pKi = 8.2) than clobenpropit (pK = 9.8), in vivo GR 175737 shows a higher activity (ED3o 1.4 mg/kg) in an ex vivo binding assay than clobenpropit (ED50 = 10.3 mg/kg) [20], Apparently the bioavailability of GR 174737 is much better than clobenpropit and the authors postulated that this observed difference might be caused by the relative ease of blood-brain-barrier passage. 

Even though substrate quality (i.e., chemical composition) is widely believed to be an important factor influencing microbial utilization of DOM, there are relatively few studies relating the composition and bioavailability of DOM. Sensitive assays for the measurement of the relative activities of various extracellular enzymes can provide an indication of the chemical composition of the bioreactive components of DOM (Sinsabaugh and Findlay, 1995 Findlay et al., 1998). The enzymatic potential of bacterial populations appears to respond fairly rapidly to seasonal changes in DOM composition in the Hudson River system. These observations clearly indicate that the chemical composition of DOM influences the microbial processing of DOM. 

Precision is a fundamental measure of assay performance and it should be assessed whenever possible [55]. This will allow a minimum understanding of how much confidence can be placed in the analytical data. In fully characterized assays designed to support definitive nonclinical and clinical studies, between-run and within-run assay precision of 15% or less (20% at the quantitation limit) is acceptable. Because of the shortened time scale for assay development and characterization in discovery these guidelines are excessively restrictive. Precision values of 20 to 30% are acceptable, as these are still less than intersubject variability associated with many nonclinical experiments. With this level of imprecision, data quality is sufficient to answer fundamental scientific questions such as relative levels of drug absorption, relative values of absolute bioavailability, and relative degree of drug clearance. 

If pharmacokinetics are dependent on dose or time, or a slow-release formulation is being studied, it is necessary to examine bioequivalence at steady state. For controlled-release formulations which are intended to produce relatively flat concentration-time profiles, an index of fluctuation is required, for example - Cn,jj])/C. A study at steady state may also be needed if the assay is not sensitive enough to quantify plasma concentrations of drug up to four half-lives after a single dose. Sometimes it is not technically feasible to assay a drug in plasma and it may then the justifiable to compare bioavailability by the total amount of drug excreted in urine, or pharmacodynamic data may be used, but these cases are exceptions. 

The method requires a compound in the low milligrams and works best for very insoluble compounds, which also are the most difficult to quantitate by other methods. The method is relatively slow, since for best results the titration rate is greatly slowed as the precipitation point is reached. The compound is solubilized at the beginning of the assay, either by pH dissolution of solid or by dissolution of a concentrated DMSO stock solution in aqueous media of appropriate pH. As a result there is no control as to which polymorphic form of the compound corresponds to the solubility product. The method is likely to be particularly useful in the solubility classification of compounds according to the FDA bioavailability waiver system because of the extensive validation of the method, its reproducibility, and its particular applicabifity to compounds with the poorest solubility. 

The chemical structure of crizotinib contains both pyridine and piperidine moieties which have measured p/fa values of 5.4 and 8.9, respectively. The compound thus exhibits pH-dependent solubility 0.034 mg/mL in water, 41 mg/mL in simulated gastric fluid (pH = 1.6), and 0.19 mg/mL in simulated intestinal fluid (pH = 6.5). Crizotinib displayed moderate human hepatocyte clearance and low-to-moderate permeability in Caco-2 cell assays. The compound also displayed good and consistent bioavailability in preclinical species, such as rat, dog, and monkey [%F 63 (rat), 65 (dog), 42 (monkey)] along with a relatively large volume of distribution (Vss about 13 L/kg) and moderate clearance values which translated to a long ti/2 (5.5-17 h). ... 

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