Plasma protein binding (PPB) of drugs is an important factor to consider when evaluating a drug. Drugs are bound to plasma proteins to varying degrees in the plasma, and the degree of binding can affect the absorption, distribution, metabolism and excretion of the drug in the body, which in turn affects the pharmacodynamic behaviour of the drug. BOC Sciences offers plasma protein binding assays to help clients accurately detect the binding of compounds to plasma proteins and predict the distribution of compounds in blood and tissues.
Plasma protein binding of a drug means that the drug enters the circulation and first becomes bound to plasma proteins, while the unbound drug is known as the free drug. The binding of drugs to plasma proteins is reversible, the pharmacological activity of the conjugated drugs disappears temporarily, and the conjugates cannot be temporarily "stored" in the blood because the molecules become larger and cannot pass through the capillary wall. The specificity of drug binding to plasma proteins is low, and as plasma protein binding sites are limited, two drugs may compete for binding to the same protein, resulting in displacement. Therefore, PPB is an important factor affecting the pharmacokinetic and pharmacodynamic properties of drugs. In addition, PPB information is also useful for estimating metabolism-related drug-drug interactions (DDI) if two drugs are used simultaneously.
Equilibrium dialysis measures plasma protein binding rate by using a dialysis membrane to separate the protein solution from the buffer, creating an equilibrium between the two in which only small molecular weight drug molecules can pass. Equilibrium dialysis directly measures the number of small molecules bound to protein, which is the key to analysing the binding of protein to small molecules, thus allowing the number of binding sites and binding constants to be derived.
Ultrafiltration requires extremely small amounts of compounds and is suitable for rapid screening. Similar to the equilibrium dialysis method, the assay is also performed using a two-compartment device separated by a semi-permeable membrane. The drug-containing protein solution is added to the upper compartment and the free drug is rapidly moved to the lower buffer side under external pressure or centrifugal conditions, and the PPB is calculated by measuring the concentration of the compound on both sides.
The most important feature of this method is the use of very high centrifugal forces (625,000g) to separate free and bound drugs. After centrifugation, the bound drug and plasma proteins are distributed in the bottom layer, the middle layer is the free drug distribution layer, and the top layer is the very low-density lipoprotein and chylomicron floats. PPB can be calculated by measuring the initial drug concentration in the intermediate layer and before centrifugation.
Fig. 1 Comparison of assay methods for the determination of plasma protein binding.
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As a leading global life sciences group, BOC Sciences has extensive experience in in vitro pharmacokinetic studies such as plasma protein binding assays. Our specialist engineers can develop different protocols to suit the different needs of our customers. If you need one, please contact us.