Mycotoxins, toxic secondary metabolites produced by fungi, often contaminate agricultural products, posing risks to human and animal health. Among these, enniatins (ENNs) and beauvericin (BEA) have gained attention due to their widespread occurrence in food and feed products and their potential toxic effects. This review comprehensively examines bioanalytical methods for detecting ENNs and BEA in biological matrices, focusing on liquid chromatography-mass spectrometry (LC-MS/MS), solid-phase extraction (SPE), and advanced sample preparation techniques. It emphasizes the significance of assessing bioaccessibility, determining the proportion of mycotoxins absorbed during digestion and their associated exposure risk. The review also explores the toxicokinetic profiles of ENNs and BEA, providing insights into their absorption, distribution, metabolism, and excretion (ADME) in humans and animals. The review also explores the biotransformation of these mycotoxins, shedding light on their metabolism into various products. Biomonitoring studies are discussed, focusing on detecting the metabolites of ENNs and BEA in biological fluids such as urine, plasma, and tissues to assess exposure. Overall, this review advocates for more research on toxicokinetics and biotransformation to fully understand the health risks of ENNs and BEA, and calls for enhanced bioanalytical detection and biomonitoring methods to improve food safety and regulatory measures.