Objective: In vitro biosensors are limited in capturing biologically relevant molecular interactions. We aimed to develop genetically encoded, live-cell biosensors for function-based screening of bioactive compounds from natural extracts. Method: Sensor cells were engineered using split intein-mediated protein trans-splicing and trans-cleavage, activating fluorophore-tagged signal peptides upon target recognition. Cortisol-, estrogen-, and EGF-detecting sensor cells were constructed and tested with known agonists, antagonists, and natural products, including pharmacopuncture materials. Result: GR sensor cells identified cortisol, dexamethasone, and novel agonists from medicinal plant extracts, including decursin as a selective glucocorticoid receptor agonist (SEGRA). Screening of pharmacopuncture materials revealed additional GR effectors. ER sensor cells enabled functional discovery of estrogenic compounds from natural sources, while EGF sensor cells discriminated agonists and antagonists and monitored downstream Ca²⁺ signaling. The system eliminated false positives associated with conventional split reporter assays and provided rapid, specific, and sensitive detection. Conclusion: Intein-mediated live-cell biosensors represent a versatile platform for function-guided screening of complex natural extracts. By enabling the discovery of selective receptor agonists and monitoring of signaling cascades, these biosensors hold strong potential for drug discovery and chemical biology applications.