The experiment was conducted to determine the effect of the mixed herbal medicine for the substitution of antibiotics on the performance of laying hens. Day old hyline 1,500 layer chicks were randomly assigned to 4 treatments. Control were 600 and three each treatment was 300. Eggs were collected at every weeks for measuring egg production and week 26, 27, 28 for chemical analyses. Chemical analyses were done for fatty acid profile of egg yolks, amino acid, antibiotics residue in eggs(collect eggs after supplying OTC 21ppm/bird/day to control for four day at 26, 27, 28) during the laying period($19{sim}77wk$). On $0{sim}4$ week, T-3 showed higher feed intake than those fed the other diets and on $5{sim}13$ week, T-2 showed highest feed intake among treatments. After birds were moved ($14{sim}18$ wk) CT-3 showed highest, but for overall rearing period, there was not significantly different among treatments. In conclusion the mixed herbal medicine did not tended to influence palatability to rearing birds. Egg quality(egg yolk color, haugh unit, eggshell breaking strength) showed no difference among treatments. Ratio of unsaturated fatty acid (mg/100g)/saturated fatty acid (mg/100g) was not difference among treatments, but linolenic acid and docosahexaenoic acid(DHA) of the mixed herbal medicine treatments showed higher then control. The antibiotics residue of CT, T treatments egg was not detected and control was not detected or below allowance. In conclusion the mixed herbal medicine can be possible to feed laying hen without antibiotics.