Alhar Baishan, Dilihuma Dilimulati, Alifeiye Aikebaier, Yipaerguli Paerhati, Xiaoxiao Qiu, Nazhakaiti Yusufujiang, Yilixiati Wusiman, Adili Abudoureheman, Wenting Zhou
Background: Heart failure (HF) has a complex pathogenesis involving oxidative stress, inflammation, and energy metabolism disorders, and requires multi-target agents. Berberis kaschgarica Rupr. (BKR) is used in Uyghur folk medicine to improve cardiovascular health, but its cardioprotective mechanisms against HF remain unclear. Methods: UPLC-MS/MS was used to identify BKRE components; DPPH/ABTS assays evaluated antioxidant activity. The MTC of BKRE was determined in zebrafish, and its effects on ISO-induced HF zebrafish were assessed via cardiac function, apoptosis, oxidative stress, and inflammation indicators. Network pharmacology, molecular docking, transcriptomics, and qRT-PCR clarified targets and pathways. Results: BKRE contained 14 bioactive flavonoids/alkaloids with favorable drug-likeness, showing concentration-dependent DPPH and ABTS scavenging. In HF zebrafish, BKRE (5/10/20 μg/mL) dose-dependently improved cardiac function, inhibited apoptosis, reduced ROS and TNF-α/IL-6, restored GSH/T-SOD, activated the AMPK-PPARα-PGC-1α pathway by binding ALOX5/NQO1, etc. Conclusions: BKRE exerts multi-mechanistic cardioprotective effects, validating BKR’s ethnopharmacological value and highlighting it as a promising HF agent/functional food.