Zearalenone-14-glucoside (ZEN-14-G), a major modified mycotoxin, has attracted considerable attention due to its potential to convert back into free zearalenone (ZEN), thereby posing toxicological risks to animals and humans. Given the limited toxicokinetic data available, this study examined the absorption, distribution, and elimination of ZEN-14-G in Arbor Acres broiler chicks following oral gavage at a dose of 0.5 mg/kg body weight to assess its potential health risks. Blood and tissue samples were collected at 0, 0.5, 1, 6, and 12 h after administration. Plasma concentrations of ZEN-14-G and its primary metabolite, ZEN, were quantified using a validated UHPLC-Q/TOF-MS method. Pharmacokinetic parameters were derived via non-compartmental analysis. The results demonstrated rapid in vivo hydrolysis of ZEN-14-G to ZEN. After oral administration, ZEN-14-G reached peak plasma concentration (Cmax) rapidly. It exhibited a short elimination half-life (T½), a limited apparent volume of distribution (Vd/F), and low total plasma clearance (CL/F). Notably, ZEN was detected in various tissues, with the highest concentrations observed in the liver, glandular stomach, and pectoral muscle. In summary, these findings underscore the important role of biotransformation in ZEN-14-G metabolism and provide key insights for assessing the health risks associated with broiler exposure to glucoside-modified mycotoxins.