Exposure to natural environments, including soil and microorganisms, is known to enhance emotional well-being and mitigate psychiatric disorders such as anxiety and depression. This study investigated the effects of short-term exposure to Streptomyces rimosus-inoculated soil, on mice with chronic restraint stress (CRS)-induced anxiety and the associated metabolic changes. Behavioral tests, including the light/dark box, open field, and novelty-suppressed feeding tests, revealed that exposure significantly alleviated anxiety-like behaviors, stress-induced neuroinflammation and hypothalamic-pituitary-adrenal (HPA) axis dysregulation of CRS mice. Untargeted metabolomics revealed alterations in amino acid metabolic pathways in the hippocampus and plasma. Potential biomarkers were highlighted, including glutamate and sphingosine in the hippocampus, and 5-hydroxytryptophan, valine, and methionine in the plasma, which showed improved metabolite levels and significant correlations with behavioral improvements, neuroinflammation, and HPA axis factors. This study provides evidence that short-term exposure to S. rimosus-inoculated soil may alleviate anxiety-like behaviors in a CRS mouse model by modulating amino acid metabolism and restoring potential anxiety-related biomarkers in hippocampus and plasma. These findings highlight a potential link between soil microbial exposure and stress-related physiological and metabolic responses.