Drugs in the bloodstream are available in both free and bound forms in which the free drug is responsible for pharmacological activities. Since protein binding determines the amount of free concentration of the drug in the blood, determining the protein binding in the early stages of drug discovery and development is of great importance. Besides, it is most beneficial to measure the free concentration of a drug in personalized medicine and therapeutic drug monitoring. For this reason, the need for sensitive, selective, and fast analytical methods to measure the free concentration of drugs and their protein binding has increased. This review aims to summarize recent advancements in analytical approaches used for the determination of free drug concentration and plasma protein binding and will focus on the most important approaches used to determine plasma protein binding. Furthermore, the concepts of each method will be described and discussed, along with their inherent advantages and disadvantages.

血液中的药物有游离形式和结合形式，其中游离药物负责药理活性。由于蛋白质结合决定了血液中药物自由浓度的量，因此在药物发现和开发的早期阶段确定蛋白质结合非常重要。此外，在个性化药物和治疗药物监测中，测量药物的自由浓度是最有益的。由于这个原因，对灵敏，选择性和快速分析方法来测量药物及其蛋白质结合的自由浓度的需求增加了。这篇综述旨在总结用于测定游离药物浓度和血浆蛋白结合的分析方法的最新进展，并将侧重于用于测定血浆蛋白结合的最重要方法。此外，将描述和讨论每种方法的概念，以及它们固有的优点和缺点。