The invention provides a pharmaceutical composition for chickens' internal lesion caused by overexertion and a preparation method of the pharmaceutical composition. The pharmaceutical composition comprises angelica sinensis, montane parsely, rehmannia glutinosa libosch, bulb of fritillary, rehmannia glutinosa, cuscuta chinensis, asparagus cochinchinensis, thalictrum simplex L. var. brevipes Hara, ophiopogon japonicus, Chinese wolfberries, limonium bicolor, poria cocos, cornus officinalis, arthraxon hispidus, Chinese yam, medicago archiducis-nicolai, radix codonopsis, tokay gecko, Chinese magnoliavine fruit, spinach beet seeds, Chinese arborvitae seeds, achyranthes bidentata, cinnamon, eucommia ulmoides, cyperus malaccensis Lam.var.brevifolius Bocklr, melastoma dodecandrum Lour, gymnadenia conopsea, fructus mori alba, pileostegia viburnoides Hook. f. et Thoms., polygonatum odoratum, arisaema cum bile, asterina pectinifera and lotus seeds. The pharmaceutical composition has the advantages that the pharmaceutical composition is capable of tonifying Qi, replenishing blood, nourishing Yin and tonifying Yang, not only can cure chickens' internal lesion caused by overexertion, but also can improve immunity of chickens and is remarkable in curative effect, low in price and free of toxic and side effects; additionally, fertilization rate of hens and hatching rate of hatching eggs can be well guaranteed.