Objective To evaluate the effect of orally administrated self-developed chlortetracycline microcapsule on microflora quantity in sheep rumen.Method Eighteen sheep of (29.0±2.3) kg of 6-month-old with good health condition were randomly assigned to six groups with three replicates in eash grcup, including one control group and five medicated groups. The sheep in the control group were only fed with basal diet. The treatments in five medicated groups were as follows. Group I: Basal diet supplemented with normal chlortetracycline premix granule; Group II: Basal diet supplemented with hydrogenated oil prescription chlortetracycline microcapsule; Group III: Basal diet supplemented with PEG4000/glycerol stearate (mass ratio of 5︰5) prescription chlortetracycline microcapsule; Group IV: Basal diet supplemented with hydrogenated oil/glycerol stearate (mass ratio of 9︰1) prescription chlortetracycline microcapsule; Group V: Basal diet supplemented with hydrogenated oil/glycerol stearate (mass ratio of 8︰2) prescription chlortetracycline microcapsule. The chlortetracycline administration dosages of sheep in the five medicated groups were all 25 mg/kg. The sheep were administrated once a day before morning feeding for five days. The rumen contents were collected by rumen fluid collector and quickly filtered through four layers of gauze after continuous administration. The filtrate were collected in the cryopreserved tube and quickly frozened by liquid nitrogen then stored at ?80 ℃. The rumen microorganisms of liquid phase were quantitatively detected by real-time fluorescence quantitative PCR (RT-PCR). Result Compared with the normal chlortetracycline premix granule, hydrogenated oil/glycerol stearate (mass ratio of 9︰1) prescription chlortetracycline microcapsule had better effect, only significantly increased the quantity of Fibrobacter succinogens, and had no significant influence on the quantities of other microbial flora in sheep rumen. Compared with the control group, hydrogenated oil/glycerol stearate (mass ratio of 9︰1) prescription chlortetracycline microcapsule and PEG4000/glycerol stearate (mass ratio of 5︰5) prescription chlortetracycline microcapsule had relatively better effects, significantly decreased the quantities of Streptococcus bovis and F. succinogens respectively, and had no significant influence on the quantities of other microbial flora in sheep rumen.Conclusion The hydrogenated oil/glycerol stearate (mass ratio of 9︰1) prescription chlortetracycline microcapsule has less inhibition effect on rumen microflora quantity and meets clinical application requirements.