This research was planned to build a Pharmacokinetic/Pharmacodynamic (PK/PD) model of 5-hydroxytryptophan (5-HTP) challenge study including a circadian rhythm component of cortisol and to predict serum cortisol based on saliva cortisol. Data from three 5-HTP challenge studies in healthy volunteers were collected. Serum 5-HTP, saliva, and serum cortisol were sampled as PK and PD marker. The population PK/PD modeling approach was applied. A baseline model of serum cortisol was built to assess the circadian rhythm before a pharmacodynamic model was used to evaluate the drug effect of the 5-HTP on cortisol. Finally, linear and power function relationships were tested to predict serum cortisol based on saliva cortisol. The PK of 5-HTP could be described using a one-compartment model with a transit compartment. The typical value for clearance was 20.40 L h and showed inter-study variability. A cosine function was chosen and properly described the circadian rhythm of serum cortisol. A linear approximation model was applied to fit the 5-HTP PD effect on cortisol data with a slope of 4.16 ng mL h. A power function provided a better description than a linear function to relate the saliva and serum cortisol. In conclusion, a circadian rhythm component was built in the PK/PD model of the 5-HTP challenge test which could better improve the understanding of the stimulating effect on HPA with cortisol change. After the 5-HTP challenge, saliva cortisol correlated well with serum cortisol and was predictable by a population PK-PD model.