Background.The microbiome-derived trimethylamine-N-oxide (TMAO) and the intestinal permeability marker zonulin are considered to be linked with depression. Moreover, carbohydrate malabsorption (CMA) was shown to be associated with signs of depression. This study is aimed at investigating possible sex-specific associations between TMAO and zonulin and the presence of depressive signs in individuals with and without CMA.Methods.Serum concentrations of TMAO and zonulin were determined in 115 and 136 individuals with the presence or absence of CMA. All 251 study participants underwent lactase gene C/T_-13910polymorphism genotyping and fructose H₂/CH₄breath testing. Additionally, they filled in the Beck Depression Inventory (BDI-II) questionnaire.Results.The median TMAO and zonulin serum concentrations were 2.66 (1.93–4.14)μmol/L and 40.83 (34.73–47.48) ng/mL. Serum TMAO levels were positively correlated with depressive symptoms (P=0.011,ρ=0.160). The strongest correlations were observed in 87 females (P=0.010,ρ=0.274) and 49 males (P=0.027,ρ=0.315) without CMA, whereas 115 patients with CMA showed no significant correlations. Zonulin tended to be negatively correlated with the BDI-II score in 49 males without CMA (P=0.062,ρ=−0.269).Conclusion.This study demonstrates a positive correlationship between the serum TMAO concentrations and the severity of depressive symptoms in females and males without CMA. Serum zonulin levels were negatively correlated with signs of depression in males without CMA. These findings suggest a gender-specific relationship between the serum TMAO and zonulin concentrations, depression, and CMA.