Isolation and Anticancer Effect of Brucine in Human Colon Adenocarcinoma Cells HT‑29

Objectives: The objective of the study is to isolate and examine the effect of brucine on human colon adenocarcinoma cell line HT‑29. Materials and Methods: Crude brucine was acquired by the extraction of Nux vomica with 80% EtOH. Diatomite chromatography and semipreparative high‑performance liquid chromatography were used to obtain brucine in pure form. HT‑29 cells were treated with brucine (125, 250, and 500 μM) for 24–72 h. 3‑(4,5‑Dimethylthiazol‑2‑yl)‑2,5‑diphenyltetrazolium bromide assay was used to assess the cell proliferation. Annexin V‑FITC/propidium iodide (PI) staining was used to identify the activity of apoptotic. Flow cytometry was used to scrutinize the effect of brucine on cell cycle progression and mitochondrial membrane potential (MMP). Bcl‑2, p53, caspase-3, PARP, and caspase-9 were spotted by Western blotting test. Results: Brucine reduced cell viability with an IC50 value of 0.368, 0.226, and 0.168 μmol/L at 24, 48, and 72 h, respectively. The apoptosis of HT‑29 was persuaded by 33.06%, 44.47%, and 71.96% at 125, 250, and 1000 μmol/L of brucine, respectively. Brucine at 250 μmol/L led to cell cycle arrest in the G1/S/G2 phase and inhibited the HT‑29 cells in the G1 phase. H1‑UL/H1‑UR was determined to be 1.79, 1.26, and 0.54 at 125, 250, and 1000 μmol/L, respectively. Brucine at 125, 250, and 1000 μmol/L downregulated the expression of Bcl‑2 but augmented the expression of p53, caspase-3, PARP, and caspase-9. Conclusion: The outcomes displayed that brucine could prevent cell proliferation, arrest the cell cycle, and increase the loss of MMP in the HT‑29 cell line. Furthermore, brucine could also persuade cell apoptosis through the expression of proapoptotic and apoptotic proteins comprising p53, caspase-3, caspase-9, and PARP. To sum up, our preclinical data designated that brucine was a probable therapeutic agent for colon cancer.