Optimization of the Flavonoid Extraction Process from the Stem and Leaves of Epimedium Brevicornum and Its Effects on Cyclophosphamide-Induced Renal Injury.

Cyclophosphamide (CTX) is a broad-spectrum alkylated antitumor drug. It is clinically used in the treatment of a variety of cancers, and renal toxicity is one of the adverse reactions after long-term or repeated use, which not only limits the therapeutic effect of CTX, but also increases the probability of kidney lesions. The total flavonoids of Epimedium stem and leaf (EBF) and Icariin (ICA) are the main medicinal components of Epimedium, and ICA is one of the main active substances in EBF. Modern pharmacological studies have shown that EBF has a variety of biological activities such as improving osteoporosis, promoting cell proliferation, antioxidant and anti-inflammatory properties, etc. However, few studies have been conducted on the nephrotoxicity caused by optimized CTX extraction, and protein-ligand binding has not been involved. This research, through the response surface optimization extraction of EBF, obtained the best extraction conditions: ethanol concentration was 60%, solid-liquid ratio of 25:1, ultrasonic time was about 25 min. Combined with mass spectrometry (MS) analysis, EBF contained ICA, ichopidin A, ichopidin B, ichopidin C, and other components. In this study, we adopted a computational chemistry method called molecular docking, and the results show that Icariin was well bound to the antioxidant target proteins KEAP1 and NRF2, and the anti-inflammatory target proteins COX-2 and NF-κB, with free binding energies of -9.8 kcal/mol, -11.0 kcal/mol, -10.0 kcal/mol, and -8.1 kcal/mol, respectively. To study the protective effect of EBF on the nephrotoxicity of CTX, 40 male Kunming mice (weight 18 ± 22) were injected with CTX (80 mg/kg) for 7 days to establish the nephrotoxicity model and were treated with EBF (50 mg/kg, 100 mg/kg) for 8 days by gavage. After CTX administration, MDA, BUN, Cre, and IL-6 levels in serum increased, MDA increased in kidney, GPT/ALT and IL-6 increased in liver, and IL-6 increased in spleen and was significant ((