Abstract: Fern fronds (about 500gm) of Cheilanthes dalhousiae Hook. Vouch. Sp. No. 21 was collected from Pindari glacier routes (2200-2800m above sea level) of Almora district of Uttarakhand state. It was extracted with acetone-water (1:1, V/V) and extract was concentrated under reduced pressure until H2O layer (up to 50ml) remained. The H2O layer was partitioned with CH2Cl2, EtOAc and BuOH Successively. The CH2Cl2 fraction gave antibacterial tests against Bacillus subtilis, Pseudomonas aeruginosa, Staphylococcus epidermidis and Escherichia coli by the standard method of disc-diffusion using DMSO-d6 solution of CH2Cl2 residue impregnated on Whatman No. 3, paper disc (6 nm) and base plates containing 10ml MH agar. Antibacterial activity was expressed as the ratio of the inhibition zone produced by CH2Cl2 extract and the inhibition zone caused by the reference, neomycin (2ìg). No antibacterial activity was observed in ethyl-acetate and n-butanol fractions. EtOAc fraction was evaporated to dryness and residue obtained was dissolved in MeOH. The MeOH soluble of EtOAc fraction was fractionated on Whatman No. 3 chromatographic papers using BAW (n- BuOHAcOH- H2O, 4:1:5, V/V, upper layer) as an eluent. Two blue UV fluorescent flavone-5-O-glycosides: Quercetin- 3-methyl ether-5-O-glycoside and Kaempferol-5-O-(6”-O-malonyl)-glycoside were isolated by RPPC from EtOAc fraction of acetone-H2O (1:1) extract of fern fronds of Cheilanthes dalhousiae. The structural elucidation of the compounds was carried out by UV, 1HNMR and MS spectral studies. [Nature and Science 2010;8(5):139- 143]. (ISSN: 1545-0740).