Blood Plasma Partitioning

The pharmacodynamic and pharmacokinetic characteristics of drugs are conspicuously affected its blood partitioning. As with plasma proteins, binding to red blood cells will affect the distribution of compounds between blood and plasma, thereby reducing the free plasma concentration of the drug. In addition, accumulation of drugs in red blood cells can lead to cytotoxicity. Therefore, it is important to determine blood distribution early in the drug discovery process. Blood plasma partitioning (BPP) assays from BOC Sciences provide standard and advanced analytical methods to measure plasma stability, red blood cell distribution, and blood-to-plasma ratios across species using fresh blood from humans and multiple species.

Typically, the concentration of a test compound in biological analysis refers to the concentration of the compound in plasma. For most compounds, plasma concentrations were not significantly different from those in whole blood. However, since the compound may selectively bind to proteins in blood cells, plasma concentrations may differ from whole blood concentrations. The whole blood/plasma distribution ratio of compounds helps to understand whether the compound enters and binds to proteins in blood cells. If the whole blood/plasma distribution ratio is greater than 1, it means that the compound enters the blood cells. Meanwhile, the PK parameters calculated by serum concentration will not reflect the PK properties of the compound well. Therefore, knowing the blood plasma partitioning ratio of compounds can reasonably select suitable biological fluids (whole blood, plasma or serum) for bioanalysis of in vivo PK samples, or correct the in vitro clearance data to in vivo data.

Brief Protocol

• Spike test compounds into fresh heparinized whole blood, reference red blood cells, and reference plasma
• Incubate at around 37°C for a period of time (e.g. 60 minutes) and take plasma and whole blood samples midway through the analysis
• Centrifugation of whole blood samples to produce plasma
• Two fractions of whole blood (plasma and red blood cells) were analyzed by LC-MS/MS along with the reference sample
• The following equation calculates the ratio of blood to plasma:

Where

KRBC/PL: the partition coefficient of a given drug in red blood cells;

H: hematocrit;

IREFPL and IPL: the LC-MS/MS response (peak area ratio to an internal standard) for the reference plasma and the plasma fraction.

Species Available

At BOC Sciences, we can evaluate blood from any of the following species in this assay:

• Mouse, rat, rabbit, dog, monkey, hamster, guinea pig, mini pig and human

Deliverables

An LC-MS/MS method will be developed for the test samples to analyze whole blood and separated plasma and estimate the amount of unchanged test substance in the sample based on the analyte/internal standard peak area ratio. The deliverables of this testing service include:

• Stability (%)
• Distribution to red blood cells (%)
• Blood to plasma ratio

In the blood plasma partitioning assay, we can determine the ability of the test compound to preferentially sequester into red blood cells by measuring the amount of the test compound in whole blood and separated plasma, and determine the whole blood/plasma ratio (r) of various animals or humans . If you have any special needs or questions about our services, please feel free to contact us.