The present study intended to investigate the variation of the physico-chemical status, zooplankton and macrophytes diversity of three semi-intensive ponds (each having 0.40 acre of water area) under Khano village of Galsi-II Block of Purba Bardhaman district, West Bengal, India. One was marked with control (C: fed only with locally available fish feed) and two others ponds (P1 and P2: fed with farm-made prebiotics plus locally available fish feed) were considered as treatment. The ‘farm-made prebiotics’ (made of fish-meal, ground-nut oil cake, bar-yeast, jaggery: fermented) was applied directly into the pond water fortnightly at the rate of 15 kg per acre fortnightly up to three months. The ponds were reared with Catla (Catla catla): Rahu (Labeo rohita): mrigal (Cirrhinus mrigala) at the ratio of 3:4:3. The water samples were drawn on day15, day30, day45, day60, day75 and day 90 at six corners of each pond. The average value was considered while computing the results of physico-chemical parameters, viz., transparency, temperature, pH, electrical conductivity (EC), carbon dioxide (CO2), dissolved oxygen (DO), total hardness (TH), total alkalinity (TA), phosphate (PO43-), chloride (Cl-), ammoniacal-nitrogen (NH4+-N), nitrate-nitrogen (NO3--N), sodium (Na+) and potassium (K+ ). The variations in the diversity of zooplankton and macrophytes in the study area were also measured. A marked difference in the case of physico-chemical parameters as well as the zooplankton and macrophytes diversity were observed under prebiotic exposure which may finally help in better growth of the fish species under prebiotic exposure. Finally, the fish farmers will be highly benefited from the application of such farm-made prebiotics.