Human endogenous biomarkers can often serve as substrates for drug metabolizing enzymes or transporters, and changes in their concentrations can indirectly reflect functional variations in these enzymes or transporters. An emerging strategy in drug-drug interaction (DDI) risk assessment involves evaluating changes in plasma or urinary concentrations of endogenous biomarkers to determine the extent of DDI. By eliminating the need to introduce exogenous probes, endogenous biomarkers significantly simplify the DDI evaluation process. This approach not only avoids potential safety risks and tolerability concerns associated with exogenous probes but also enables simultaneous and dynamic monitoring of multiple metabolic enzymes and transporters. Furthermore, they offer notable advantages for evaluating special populations such as patients with hepatic or renal impairment, elderly individuals, and children, And endogenous biomarkers can be effectively combined with quantitative pharmacological models or artificial intelligence approaches, thus providing a novel paradigm to improve the efficiency of DDI assessment. This review summarizes recent studies on endogenous biomarkers associated with CYP enzymes as well as hepatic and renal transporters, providing reference information for future DDI research and evaluation.