Background: Aerva lanata (Linn.) Juss. ex Schult.(AL), belonging to family Amaranthaceae, is a popular medicinal herb known for its immense antiurolithic properties. Several researchers have reported the antilithiatic efficaciousness of the extract. However, there does not exist any evidence regarding the bioassay-guided fractionation of the same as a route to identification of potential antiurolithic phytoconstituents. Objective: The present study aims to evaluate the antiurolithic competence of different extract based phytoconstituent enriched fractions (Fr) of the aerial parts and roots of AL under in vitro conditions to screen out the Fr possessing the best activity. Materials and Methods: Different Fr from the extracts of aerial parts and roots were prepared by solvent-solvent partitioning and were subjected to preliminary phytochemical screening to affirm the presence of intended phytometabolites. The Fr were analyzed for their antilithiatic efficacy by in vitro aggregation and growth assay to assess their ability to counteract calcium oxalate (CaOx) aggregation and growth. The phytochemical evaluation was also performed by high-performance thin layer chromatography (HPTLC). Results: Preliminary phytochemical screening ascertained the presence of expected phytochemicals in the Fr. Antiurolithic assays demonstrated that the flavonoid and phenolic enriched Fr III derived from the extract of aerial parts exhibited the maximum hindrance to CaOx aggregation and growth at 1000 μg/ml with a percentage inhibition of 67.14% ± 1.84% and 66.66% ± 1.65%, respectively. HPTLC analysis revealed the presence of flavonoids quercetin, kaempferol, and myricetin in the same. Conclusion: The antiurolithic ability of AL on the overall process of stone formation might be attributed to the presence of potent phenolic and flavonoid compounds in the aerial parts which may act as a source for isolation of powerful antiurolithic leads from the same.