Objective To investigate the effects of walnut paste on kidney-yang deficiency and the gut microbiota.Methods Using hydrocortisone [25 mg/(kg·d)] to establish a kidney-yang deficiency mouse model, the experiment is randomly divided into a normal group, a model group, a positive control group, and low-, medium-, and high-dose walnut paste groups. The mice are continuously administered walnut paste via gavage for 14 days. Changes in body weight, autonomous activity, and organ indices are analyzed. Serum levels of cyclic adenosine monophosphate (cAMP), cyclic guanosine monophosphate (cGMP), corticosterone (CORT), testosterone (T), and estradiol (E2) are measured. Pathological conditions of the kidney, testis, and spleen are examined using HE staining, and the composition of the gut microbiota is analyzed.Results Compared to the model group, walnut paste restores the body weight of mice to near-normal levels, mitigates changes in organ indices, and increases autonomous activity. It significantly elevates serum levels of cAMP, T, and CORT in kidney-yang deficiency mice (P<0.05) and markedly reduces serum levels of cGMP and E2 (P<0.05). Additionally, walnut paste alleviates pathological damage in the kidney, testis, and spleen tissues. 16S rDNA sequencing reveals that walnut paste counteracts the reduction in gut microbiota richness and diversity in kidney-yang deficiency mice. It significantly increases the relative abundance of Bacteroidetes, Lachnospira, and Lactobacillus (P<0.05) and significantly decreases that of Firmicutes, Actinobacteria, and Staphylococcus (P<0.05).Conclusion Walnut paste has beneficial effects on kidney-yang deficiency and on the diversity and composition of the gut microbiota.