1,2,6,8-Tetrahydroxyxanthone (SR1) and 2,8-dihydroxy-1,6-dimethoxyxanthone (SR2) have been qualitatively and quantitatively determined in leaves and rhizomes of Swertia paniculata. The methanol extracts were analyzed on a C18 column using MeOH and H20 as mobile phase at a flow rate of 1 mL/min. Identification was achieved using an UV variable lambda detector set at 260 nm. Variation in the bioactive compounds with respect to season and altitude was determined, along with the examination of the antidiabetic activity of the plant. The maximum concentration of SR1 and SR2 was found in leaves collected from Tungnath (3600 m), Uttarakhand, India, in October. The blood sugar level in alloxan induced rats was decreased by 51.0% by the methanol extract of the leaves. Overall, results showed that the concentration of bioactive compounds increased with altitude. October was the period of prime potency and should be the time of harvest for S. paniculata.