Abstract: In order to study the effect of different amounts of L-malic acid on rumen fermentation parameters in vitro, the influence of four different amounts of L-malic acid（group A was not added, 5 mmol/L in group B,10 mmol/L in group C,15 mmol/L in group D） were selected for the in vitro fermentation for 8, 16, 24 and 48 h, then the dry matter disappearance rate, pH value, ammonia nitrogen, microbial protein and volatile fatty acid were determined. The results showed that the gas production increased with the increase of the amount of L-malic acid addition at each fermentation time, and the dry matter disappearance rates in groups B, C and D（except for 16 h of fermentation time in group B） were significantly higher than that in group A（P＜0.05）. After 48 h of fermentation, the concentrations of ammonia nitrogen and microbial protein in group C and D were significantly higher than those in group A and B（P＜0.05）, and the difference between C and D groups was not significant（P＞0.05）. At each fermentation time, the concentration of volatile fatty acids increased with the increase of the amount of L-malic acid, the concentration of volatile fatty acids in group D reached the maximum value, and the difference between C and D groups was not significant（P＞0.05）. The results showed that the addition of L-malic acid could improve the substrate fermentation level, the concentration of ammonia nitrogen, microbial protein and volatile fatty acid in the fermentation broth, and improve the rumen fermentation condition. Considering the cost, the comprehensive effect of adding 10 mmol/L was the best.