Abstract: [Objective] The aims of the present study were to assess the optimal ratio of corn straw and spent Flammulina velutipes substrate as roughage in preparation of fermented total mixed ration （FTMR） for beef cattle, and to provide technical support for the high-value utilization of by-products such as mushroom bran. [Method] For the preparation of roughage, a total of 7 treatment groups with different mixing ratios of corn straw and spent Flammulina velutipes substrate were set up. Total mixed ration （TMR） were made from even mixing of corn straw and spent Flammulina velutipes substrate at the ratios of 8∶2 （T1 group）, 7∶3 （T2 group）, 6∶4 （T3 group）, 5∶5 （T4 group）, 4∶6 （T5 group）, 3∶7 （T6 group） and 2∶8 （T7 group） with addition of beef cattle concentrate supplement at a concentrate-roughage ratio of 4∶6 （fresh weight）. TMR samples from different treatment groups were placed into silage bags, and then compacted, sealed, vacuumized and stored in a dark room to naturally ferment for 60 days to prepare FTMR. The nutritional component and mycotoxin content of the prepared TMR and FTMR samples were measured and statistically compared. Furthermore, the fermentation parameters of FTMR were assessed to determine the optimal ratio of corn straw and spent Flammulina velutipes substrate. [Result]①As the ratios of corn straw to spent Flammulina velutipes substrate decreased, the DM content of TMR gradually reduced, the CP content gradually increased, the NDF and ADF contents generally elevated, and the EE content fluctuated. ②As the ratios of corn straw to spent Flammulina velutipes substrate decreased, the DM content of FTMR gradually reduced as well. Compared with the corresponding TMR, the FTMR in T4 group had the lowest DM loss rate. The CP content of FTMR in T3 and T4 groups were significantly （P<0.05） higher than that in the other groups. Compared with the corresponding TMR, the CP content of FTMR in T3 and T4 groups increased. No significant （P>0.05） differences in EE content of FTMR among different groups were observed. Compared with the corresponding TMR, the EE content of FTMR decreased in T1-T4 groups, but increased in T5-T7 groups. The NDF content of FTMR in T1-T5 groups were significantly （P<0.05） lower than that in T6 and T7 groups, with the lowest NDF content observed in T3 group. Compared with the corresponding TMR, the NDF content of FTMR in T3 and T5 groups decreased. The ADF content of FTMR in T1-T5 groups were significantly （P<0.05） lower than that in T6 and T7 groups. Compared with the corresponding TMR, the ADF content of FTMR in T4 and T5 groups decreased. ③The FTMR samples from all the treatment groups had acidic aroma and good fermentation quality. The pH value of FTMR in T3 group was the lowest, which was significantly （P<0.05） lower than that in the other groups. The pH value of FTMR in T4 group was lower as well. As the ratios of corn straw to spent Flammulina velutipes substrate decreased, the LA content of FTMR showed a trend of first increasing and then decreasing, with T4 group having the highest content, followed by T3 group, both of which were significantly （P<0.05） higher than the other groups. As the ratios of corn straw to spent Flammulina velutipes substrate decreased, the AA content of FTMR exhibited a trend of first increasing and then decreasing, with T5 group having the highest content, followed by T4 and T3 groups. Trace level of PA were detected in FTMR samples from all the treatment groups, and there were no significant differences （P>0.05） among the different groups. Trace level of BA were detected in FTMR in T6 and T7 groups, while the other groups were not detected. The AN/TN ratio of FTMR in T3 group was the lowest. FTMR in T6 and T7 groups had the AN/TN ratio exceeding 10%, while the other groups was less than 10%. The differences in AFB₁, ZEN and OTA contents of TMR and FTMR samples among different groups were not significant （P>0.05）. According to the Feed Hygiene Standard （GB 13078-2017）, the mycotoxin content of the TMR and FTMR samples did not exceed the threshold. ④The main effects analysis showed that the fermentation treatment significantly （P<0.05） reduced the ABF₁ and ZEN contents of TMR, while the treatment of the mixing ratios of corn straw and spent Flammulina velutipes substrate, as well as the combination effect of fermentation and mixing ratios, had no significant （P>0.05） effects on the ABF₁ and ZEN contents of the TMR and FTMR samples. In addition, the fermentation treatment, the treatment of the mixing ratios of corn straw and spent Flammulina velutipes substrate, and the combination effect of fermentation and mixing ratios had no significant （P>0.05） effects on the OTA content of the TMR and FTMR samples. [Conclusion] Based on the comprehensive analysis on nutritional component, mycotoxin content and fermentation quality, the preparation of FTMR with corn straw and spent Flammulina velutipes substrate as roughage was an efficient way to preserve and utilize the by-products such as straw and mushroom bran. Corn straw and spent Flammulina velutipes substrate mixed at the ratios of 6∶4 （T3 group） or 5∶5 （T4 group） had the better performance.

Key words: corn straw, spent Flammulina velutipes substrate, fermented total mixed ration, nutritional component, fermentation quality, mycotoxin