Bemisia tabaci is one of the most notorious agricultural pests in the world. A vicious circle among insect resistance, dose increased, environment and human body impaired as the overuse of synthetic pesticides are becoming increasingly evident. Entomopathogenic Beauveria sp. is known as an effective natural enemy to control B. tabaci. Therefore, this study aimed to purify and identify the biological compounds from Beauveria sp. LY2 via extensive chromatographic techniques, NMR and MS and evaluated for their insecticidal activities against B. tabaci via contact and feeding assay. The outcome identified that one new cerebroside, cerebroside F (1), nine known compounds, cerebroside B (2), bassiatin (3), methyl 1,4-dihydro-4-oxo-2-quinolinecarboxylate (4), cerevisterol (5), 9-hydroxycerevisterol (6), 6-dehydrocerevisterol (7), (22E,24R)-ergosta-8(14),22-diene-3β,5α,6β,7α-tetrol (8), melithasterol B (9) and ergosterol peroxide (10) were isolated. Among the known compounds, methyl 1,4-dihydro-4-oxo- 2-quinolinecarboxylate (4) was isolated from natural origin for the first time. It is evident from the results that compounds 3, 4 and 7 strongly featured insecticidal activities against B. tabaci, being the LC50 value as 10.59, 19.05, 26.59 μg/mL respectively in contact as well as 11.42, 5.66, 5.65 μg/mL respectively in feeding experiment. Moreover, no adverse effect on plant growth/height or phytotoxicity was observed on pepper, cucumber, tomato and cotton. The data from the current study has provided the foundation for the use of newly purified compounds against Bemisia tabaci as an alternative to synthetic chemical compounds.