Plasma Stability Assay

Creative Bioarray's plasma stability assay can measure the stability of compounds in plasma, helping customers find unstable compound structures and screen prodrugs.

Plasma Stability Assay

Why do we need the plasma stability assay?

  • Plasma stability plays an essential role in drug discovery and development. In addition to hepatic metabolism, compounds will also be subject to degradation and modification by enzymes in plasma, particularly hydrolases and esterases. Unstable compounds tend to have rapid clearance and short half-life, which leads to poor in vivo performance.
  • Certain types of compounds are potentially unstable to the plasma enzymes. For instance, structures containing amide groups are susceptible to hydrolysis by esterase and other plasma enzymes. Poor plasma stability can result in fast clearance and short half-lives.
  • Pharmacokinetic studies are especially challenging for unstable compounds because they continue to degrade even after blood is sampled from the study animal, which results in ambiguous PK data.
  • Besides, some prodrugs become physiological active on account of the same enzyme reactions.
  • In addition, plasma enzymes can significantly change the bioavailability of active compounds, which should be assessed at the early stages of the drug discovery process.

Brief Protocol

  • The test compound was incubated with plasma at five different time points at 0, 15, 30, 60, and 120 minutes. The reaction was terminated by methanol containing internal standard.
  • After centrifugation, the concentration of the test compound in the supernatant was quantified by LC-MS/MS. The percentage of test compound remaining at each time point relative to the 0-minute sample is then reported.
  • Control compounds known to be metabolized by plasma esterases were also incubated with each batch of test compounds.
Test systemPlasma
Test Compound Concentration 1 µM (or custom)
Microsome Concentration 0.5 mg/mL
Time Points 0, 15, 30, 60, 120 minutes
Number of Replicates 3
Negative Control Pepstatin, Leupeptin
Positive Controls Propantheline, Benfluorex
Compound Requirements 10 µL of 10 mM DMSO solution
Analysis Method LC-MS/MS
Data Delivery Intrinsic clearance; Half-life

Case Study

  • Plasma

Pooled and heparinised plasma is available in a range of species.
e.g. Human, dog, rat, mouse and monkey.

  • Methods and Results

Plasma and test compounds are added to the individual wells of a 96-well microtiter plate. Compounds are incubated at 37°C. Five time points over 120 minutes are analyzed (0, 15, 30, 60 and 120 min). All tests are performed in duplicates. The samples are analyzed by HPLC-MS.

Percent Remaining (%)T1/2
0 min15 min30 min60 min120 min
Sample #1100.

% Remaining = 100 x (PAR at appointed incubation time / PAR at T0 time)

where PAR is the peak area ratio of analyte versus internal standard (IS).

The in vitro plasma half life (T1/2) was calculated using the expression: T1/2=0.693/K,

where K is the slope found in the linear fit of the natural logarithm of the fraction remaining of the parent compound vs. incubation time (Roula, 2008).

Quotation and Ordering

If you have any special needs or questions regarding our services, please feel free to contact us. We look forward to cooperating with you in the future.


  1. Konsoula, Roula; Mira Jung. "In vitro plasma stability, permeability and solubility of mercaptoacetamide histone deacetylase inhibitors." International journal of pharmaceutics 361.1-2 (2008): 19-25.

For research use only. Not for any other purpose.