Picrorrhiza kurroa Benth is an important plant of Indian System of Medicine for its well-known pharmacological activities. In the present paper, we report our work on isolation and characterization of two important irridoid glycosides, namely, Picroside I and Kutkoside with a modified method as reported earlier for the plant. Further, these two marker compounds were quantified by thin layer chromatography densitometric methods using high performance thin layer chromatography. To the best of our knowledge, this is the first report of simultaneous quantification of these two compounds using HPTLC from this plant. The thin layer chromatography densitometric methods were found to be precise with RSD for intra-day in the range of 2.69-4.16 and 0.45-2.3 and for inter-day in the range of 1.96-4.25 and 0.75-3.09 for different concentrations of Picroside I and Kutkoside, respectively. Instrumental precision was 3.19 and 3.05 (% RSD) for Picroside I and Kutkoside, respectively. Accuracy of the method was checked by conducting recovery studies at three different levels for both the compounds, and the average percentage recoveries obtained were 99.9 and 99.92%, respectively, for Picroside I and Kutkoside. Picrorrhiza kurroa sample was found to contain 3.66% and 4.44% w/w of Picroside I and Kutkoside, respectively. [ABSTRACT FROM AUTHOR]