Reverse-Phase High-performance Liquid Chromatography (RP-HPLC) based technique is one of the most sensitive techniques to detect the antibiotics present in honey. In the southern part of Tamilnadu, India, majority of the farmlands are occupied by plantations such as coconut, banana and rubber. A variety of antimicrobial compounds and antibiotics, which have been reported in pollen, nectar and other floral parts of the plant, gets accumulated in honey through honeybees (Apis mellifera). We have collected the nectar samples from banana (Musa paridasiaca) and rubber (Ficus elastica) flowers and the honey from honey hives of banana and rubber cultivated areas. The extracted nectar and honey samples are subjected to RP-HPLC analysis with authentic antibiotic standards. Nectar and honey samples showed 4-17, 11-29 ${mu}g/kg$ of streptomycin, 2-29, 3-44 ${mu}g/kg$ of ampicillin and 17-34, 26-48 ${mu}g/kg$ of kanamycin respectively.