Background: Plants from the Asteraceae family were commonly used to treat various diseases. The metabolomic profile of this family consisted of bioactive flavonoids and other phenolics. Chamomile is a member of the Asteraceae family. Jordanian and European chamomile are two varieties ofMatricaria chamomilla(German chamomile), which were grown under different environmental conditions, were studied. Many examples of plant varieties with significant distinction in the secondary metabolite they afford have been described in the literature. Multivariate statistical analysis was employed to measure the depth of this variation in two chamomile varieties. Methods: From both types, crude extracts were prepared using solvents of different polarities and tested for their biological activity. The semipolar fraction of the European variety showed anticancer and antioxidant activity. Meanwhile, the semipolar fraction of the Jordanian type exhibited only antioxidant activity. Both extracts were fractionated, and then the biological activity was again assayed. Results: European and Jordanian chamomile fractions produced dicaffeoylquinic acid isomers exhibiting antioxidant capability. Additionally,Z-glucoferulic acid was produced from the European chamomile, demonstrating antioxidant activity. The European samples afforded two major compounds, chrysosplenetin and apigenin, that displayed anticancer activity. Conclusions: Different environmental conditions between Jordanian and European chamomile affected the type of isolated compounds. Structure elucidation was performed with HPLC-MS coupled with dereplication techniques and 2D NMR experiments.