Abstract
Natural products have gained significant interest in cancer therapy. Thymoquinone (TQ), a bioactive compound mainly derived from Nigella sativa seeds, and bee venom (BV), a complex mixture of bioactive components secreted by honeybees (Apis mellifera), are notable examples. This study aimed to evaluate the synergistic anticancer effects of TQ-BV combination on selected cancer cell lines—HeLa, MCF-7, HCT—and normal human skin fibroblasts (HSF). BV was collected using a novel remote-controlled extraction device designed to maintain venom purity by isolating it from environmental contaminants and minimizing light exposure. HPLC was used to quantify the main components of the venom, thereby detecting the sample’s purity. MTT assays assessed cell viability. Apoptotic activity was analyzed through Annexin V-FITC/PI staining. Flow cytometry was used to evaluate the cell cycle distribution, focusing on the Sub-G1, G1, and S phases in HeLa cells. Results from the MTT assay showed that the TQ-BV combination showed markedly increased potency against HeLa cells with an IC(50) value of (1.495 ± 0.198 µg/mL). Combination Index (CI) analysis confirmed a synergistic effect in all cell lines. The apoptosis assay revealed an increase in both early and late apoptotic cells with the combination treatment in HeLa cells, which exhibited a notable rise in late-stage apoptosis, indicating enhanced apoptotic activity. Cell cycle analysis revealed that the combination appeared to induce arrest at the G1 and S phases, which may have contributed to reduced proliferation. Overall, the TQ-BV combination exhibited strong anticancer potential by inducing cell cycle arrest and promoting apoptosis. The high purity of BV, achieved through an optimized extraction method, may have enhanced its efficacy. Further in vivo studies are needed to confirm these findings and explore potential clinical applications. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1038/s41598-025-28733-9.