*4.2. Cell Culture*

Colorectal cancer cell lines HCT-116 (Accession number: CRL-3504), HT-29 (Accession number: HTB-38), and SW620 (Accession number: CVCL\_0547) were obtained from ATCC (Houston, TX, USA). Cells were maintained in RPMI-1640 and DMEM supplemented with 100 μg/mL streptomycin, 100 units/mL penicillin, and 10% heat-inactivated fetal bovine serum in a humidified, 5% (*v/v*) CO2 atmosphere at 37 ◦C [67]. All cell lines and cell line materials were confirmed to be mycoplasma free.

## *4.3. Cytotoxicity Assay*

The cytotoxicity of GCB and Terr was tested against HCT-116, HT-29, and SW620 cells using the SRB assay as previously described. Exponentially growing cells were harvested using 0.25% Trypsin-EDTA and plated in 96-well plates, at concentrations of 1000–2000 cells/well. Cells were exposed to GCB, Terr, and GCB + Terr for 72 h (normoxia and hypoxia) and subsequently fixed with TCA (10%) for 1 h at 4 ◦C. After the plates were washed several times, the cells were exposed to a 0.4% SRB solution for 10 min in the dark and subsequently washed with 1% glacial acetic acid. After leaving the plates to dry overnight, Tris-HCl was used to dissolve the SRB-stained cells, and their color intensity was measured at 540 nm using a microplate reader [68].
