Abstract:This study aimed to explore the effects of supplementary feeding on milk yield, blood biochemical indexes, milk components, and fecal microflora diversity of Yili horses under grazing condition.18 grazing Yili mare horses with physiological condition consistent were randomly assigned to three groups with six mare horses each. The control group normal grazing, whereas the test group I and II were provided with supplemented with 1 kg/d of concentrate and 1 kg/d of concentrate + 0.4 kg/d coated fatty acids, respectively. After 3 days of pre-feeding, begin the experiment.Horse milk was collected every 7 days, and 10% of the milk yield of each horse was mixed in 100 mL milk sample bottles and stored in -20℃ refrigerator.Which blood samples were collected from the jugular vein before morning feeding (0 h) and at 1,2,3,4,6,9 and 12 h after morning feeding on day 40, and the plasma were centrifuged at 4000 rpm for 15 min and conservation at-20℃. Excrement was collected within 8 h on days 45 and 46 of the test, mixed with 5g of fresh fecal samples in a 5 mL frozen storage tubes and stored in liquid nitrogen. Compared with the control group, the lactation volume of Yili horses in test groups I and II was significantly higher throughout the trial period (P <0.05), and the late test group II was significantly higher than test group I (P <0.05). The yield of milk fat, lactose, and milk protein in test group II was significantly higher than that in test group I and control group (P <0.05); the urea nitrogen content in test group I was significantly lower than the control group (P <0.01). The lauric acid, tetradecanoic acid, cis-11,14,17-eicosotrienic acid, total saturated fatty acid, and total saturated fatty acid/total unsaturated fatty acid in test group II were significantly higher than those in the control group and test group I (P <0.05); The plasma glucose concentration of mares in test group II was significantly higher than that of control group and test group I at 1, 2 and 9 h after feeding (P <0.05), and significantly higher at 4 h after feeding (P <0.05); At 3 h and 6 h after feeding, test group II mares had significantly higher plasma free fatty acids and triglycerides concentrations than the control and test group I (P <0.05), at 4 h after feeding, the plasma triglyceride concentrations were significantly higher than the control and test I groups (P <0.05). Plasma low density lipoprotein and very low density lipoprotein concentrations were significantly higher in mares at 3 h and 9 h after feeding than in control and group I (P <0.05). The total number of OTUs was 3483, and the number of OTUs was 4984,5487, and 5158 in the control and test I and II, respectively. At the phylum level, the total abundance of Bacteroidetes and Firmicutes was more than 75%, and the test group Verrucomicrobia was significantly higher than the control group (P <0.05), and the test group II was significantly higher than the test group I (P <0.05). The Treponema_saccharophilum was significant difference in the feces of the control group (P <0.05); WCHB 1_41, kiritimatiellae, and Verrucomicrobia were significant difference of the test group II (P <0.05) and the fecal flora was mainly related to lipid synthesis, metabolism, and energy metabolism (P <0.05). Supplemental feeding can improve lactation volume of grazing Yili horses, improve the absorption and utilization of fatty acids, and improve the composition and diversity of horse milk and fecal flora to a certain extent.