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ADME PK

Chemical stability assay

Use our chemical stability assay to measure degradation of your compound in aqueous buffer.

The chemical stability assay is one of Cyprotex's in vitro ADME screening services. Cyprotex deliver consistent, high quality data with cost-efficiency that comes from a highly automated approach.

Assessment of chemical stability

  • A compound is chemically unstable when it is degraded by non-enzymatic processes. Degradation may be caused by several mechanisms, the most common being hydrolysis, oxidation, or light-catalyzed degradation.
  • Compounds that are highly unstable may not be suitable as drug candidates since it may be difficult to maintain a therapeutically effective formulation.
  • Compounds designed for oral administration must be chemically stable at the low pH values observed in the stomach in order for this to be an acceptable route. A range of different pH values are available.
  • Cyprotex's Chemical Stability assay assesses degradation of the test article in buffer. The chemical stability assay can be performed using a range of different media including SGF (simulated gastric fluid), SIF (simulated intestinal fluid) or buffers at different pH values
At the drug discovery stage, when drug candidates are screened against biological targets, compounds need to have sufficient stability in the assay buffers for enzyme, receptor, or cell-based assays to reliably measure biological activity.

1Di L, Kerns E.H, Chen H, and Petusky SL. (2006) J Biomol Screen 11(1); 40-47

Protocol

Chemical stability assay protocol

Test Compound Concentration 5 μM (different concentrations available)
Time Points 0, 15, 30, 60, 120 minutes
Number of Replicates 2
Compound Requirements 50 µL of 10 mM solution
Analysis Method LC-MS/MS
Data Delivery % Parent compound remaining at each time point
Half life

Data

Data from Cyprotex's Chemical Stability assay

 
Figure 1
Degradation of omeprazole over 120 min in simulated gastric fluid.

Q&A

Questions and answers on chemical stability

Please provide an overview of Cyprotex's Chemical Stability assay.

The chemical stability screen measures the degradation of a compound by non-enzymatic processes. This may be caused by several mechanisms, the most common being hydrolysis, oxidation, or light-catalyzed degradation.

The test compound (final incubation concentration = 5µM) is incubated with the appropriate buffer. Samples are removed at 5 different time points into ice-cold methanol. The concentration of test compound is quantified by LC-MS/MS. The percentage of test compound remaining (relative to the 0 min time point) at the individual time points is then reported.

Why is the chemical stability of the compound important?

Oral administration is the route of choice for new drugs. Compounds must be chemically stable at the low pH values observed in the stomach in order for this to be an acceptable route. Therefore, knowing if a compound is stable at pH 2 is useful information in predicting if oral administration is likely to pose a problem.

Many in vitro screens are performed at physiological pH to mimic the in vivo situation. If the compound is unstable at pH 7.4, samples generated in the in vitro screens may decay rapidly prior to analysis. Situations such as these may be time consuming and expensive to resolve.

There are two main areas of concern. Firstly, if a compound degrades to a toxic degradation product, and secondly, maintaining a therapeutically effective formulation.

The formulation, storage and packaging of drugs may become important for chemically unstable compounds. Altering the pH of the formulation, removing moisture from the formulation, storing at lower temperatures, or protecting the drug from light may all be used as measures to reduce chemical degradation.

Can the chemical stability be investigated at different pH values?

The chemical stability assay can be performed at any pH. Commonly used pH’s include pH 2, pH 6 and pH 10. We can also investigate stability in a range of buffers such as simulated intestinal fluid (SIF) and simulated gastric fluid (SGF) and also at either room temperature or 37ºC.

How do I interpret the data from the chemical stability assay?

Obviously, a very highly unstable compound is problematic and may preclude development. The decision of whether to proceed with a compound may be based on the intended route of administration, the extent of decay, the reason for the instability and whether procedures can be put into place to reduce the decay. Typically, a compound is classed as unstable if the % compound remaining at the 120 minute time point is less than 85% of the 0 minute time point.

References

1 Di L et al. (2006) J Biomol Screen 11(1); 40-47

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