AIM: To study the in vivo material basis that is involved in the rapid antidepressant effects of saffron glycoside-I, so as to provide evidences to facilitate the interpreting  the inconsistency of PK-PD, and the exploring the underlying mechanism. METHODS: The antidepressant efficacy of saffron glycoside-I and saffron aglycone was evaluated by investigating depressive-like behaviors such as Sucrose Preference Test (SPT), Social Interaction Test (SIT), Tail Suspension test (TST), and the Forced Swim Test (FST) in mice induced by Chronic Unpredictable Stress (CUMS) and Chronic Social Defeat Stress (CSDS). The LPS-induced model of inflammation was used to investigate the regulatory effect of saffron glycoside-I on primary inflammatory factors. The regulation of saffron glycoside-I and saffron aglycone on small molecules and neurotransmitters in blood and intestine were further studied by non-targeting metabolomics and targeting metabolites of neural transmitters based on HPLC-qTOF/MS and LC/MS-MS techniques. RESULTS: Saffron glycoside-I and its aglycone showed rapid and efficient antidepressant effects, and they significantly improved the performance of CSDS mice on SIT, TST, and FST. Saffron glycoside-I did not show obvious effect on reducing LPS-induced inflammatory factors of IL-6 and TNF-α. On the contrary, typical anti-inflammatory drug components of paeoniflorin, silybin and magnesium isoglycyrrhizinate significantly reversed the elevation of IL-6 and TNF-α induced by LPS, but they could not rescue the depressant behaviors of CSDS mice. Metabolomic study revealed perturbation of metabolic phenotype, small molecules and neural transmitters in plasma and gut contents of both CUMS and CSDS mice. To a large content, and Saffron glycoside-I and saffron aglycone modulated metabolic phenotype, similar to the normal controls. Saffron aglycone successfully regulated a variety of metabolites, metabolites associated in purine pathway, and transmitters, such as 5-HT, γ-GABA, glutamic acid and norepinephrine that were perturbed in plasma and gut contents in CSDS mice. CONCLUSION: Saffron aglycone shows a rapid antidepressant effect similar to saffron  glycoside-I, the antidepressant effect of saffron glycoside-I is closely associated with its primary metabolite saffron aglycone in vivo. The antidepressant effect of saffron aglycone is involved in its regulation effects on endogenous small molecules and neurotransmitters in the circulatory system and intestinal tract of depression model mice, instead of that on inhibition on inflammatory factors.