We examined how CBD extract influences the activity of the immune system in the hemolymph of honey bees in the hive test. The bees were divided into 3 groups: (CSy) bees fed with CBD in sugar syrup with glycerin; (CSt) cotton strip with CBD placed in hive bees fed pure sugar syrup, (C) control bees fed sugar syrup with glycerin. CBD extract increased the total protein concentrations, proteases and their inhibitor activities in each age (the except for acidic protease activities in the 21st and 28th day and alkaline protease inhibitor activities in the 28th day in CSt group) in comparison with group C. In the groups with the extract there was also an increase in the enzymatic marker activities: ALP, AST (decrease on day 28 for CSt), ALT; and non-enzymatic marker concentrations: glucose; triglycerides; cholesterol and creatinine. The urea acid and albumin concentrations were lower in CSy and CSt groups compared to the C group (higher concentration of albumin was displayed by control bees). Higher activities/concentrations of most of biochemical parameters were obtained in the CSy compared to the CSt and C. CBD supplementation can positively influence workers’ immune system.