*3.2. Cell Proliferation Assays*

The synthesised arctigenin derivatives were measured for anti-proliferative activity against colorectal cancer HCT-116 and triple negative breast cancer MDA-MB-231 cell lines using 3H-thymidine incorporation assays. Cell lines were purchased from the American Type Culture Collection (ATCC). The anti-proliferative assays were conducted according to our previously reported methods. [36–38]. In short, cells were seeded in 96 well plates with 3000 cells per well and incubated with 10 μM of arctigenin-derived compounds for three days. An amount of 0.04 μCi of 3H-thymidine was added per well and incubated for 5 h before cells were harvested and counted. All experiments were performed in duplicate wells on separate plates with three repeats. The percentage of cells which showed incorporation of 3H-thymidine into the DNA relative to the control samples directly measured the cell proliferation. Two known previously active compounds were used as positive controls [39], alongside a negative control, with no compounded added.
