There remains a significant gap in production performance and disease resistance between silkworms reared on artificial diets and those reared on mulberry leaves. This study aims to identify key differential proteins through proteomic analysis of the midgut of silkworms fed artificial diets compared to those fed mulberry leaves. Utilizing molecular docking technology, three anti-nutritional factors that consistently bind to the UGT40B4, UGT340C2, and UGT40A1 proteins were selected, and the differential expression of these UGT genes in response to various anti-nutritional factors was examined. The findings indicate that variations in feed significantly influence the expression of digestive, metabolic, and immune-related proteins within the silkworm midgut. Notably, the expression levels of the UGT40B4, UGT340C2, and UGT40A1 genes vary across different silkworm organs and developmental stages, reflecting their respective physiological roles. Furthermore, the effects of soybean isoflavone, tannic acid, and arabinoxylan on silkworm growth and cocoon quality were found to differ when these substances were incorporated into semi-synthetic feed. This research is anticipated to provide valuable insights for future studies on the role of UGT genes in the silkworm midgut and the formulation of artificial diets for silkworms.