A botanical mixture spiked with the known compounds berberine, magnolol, cryptotanshinone, and alpha-mangostin was fractionated three times and separated into equal sample sets containing 3, 5, or 10 final pools. The resulting pools were suspended at 0.1 or 0.01 mg mL−1 (reported as mass of dry extract per volume solvent) in methanol for UPLC-MS analysis. Each data subset was analyzed using hierarchical cluster analysis (HCA) before and after filtering to remove chemical interferents.