Rhodamine derivatives have been persistently used as fluorescent/solvatochromic dyes in various areas. For example, they are used as a labeling agent in biotechnology and a chemosensor. Also, rhodamine derivatives are used for cell imaging experiment. Herein, we report on the synthesis of fluorescent derivatives based on rhodamine B (RHB1, RHB2, and RHB3). We confirmed the structures and purities of the synthesized probes i.e., RHB1, RHB2, and RHB3 by using analytical technique's viz. 1 H NMR, 13 C NMR, Mass and FT-IR followed by exploring spectroscopic properties by using fluorescence/UV absorption spectroscopy. The effect of solvents on the fluorescence and absorption properties of the synthesized compounds has been comprehensively studied using a series of solvent. In addition, synthesized RHB1, RHB2, and RHB3 were found to be non-toxic against the cancer cell (MDA-MB-231). It is clear from the result of the conventional fluorescence microscopy that probe RHB2 exhibits significant optical properties as compared with those of RHB1 and RHB3.