Cancer refers a disease that develops when a few cells grow-out and spread into other organs- internal. Cancer can manifest almost anywhere in body. The present research was based on the synthesis and evaluation of anti-cancer potential of novel derivatives of 1,4-dioxane. 1,4-Dioxane was first identified in 1863. It started to be used commercially in the 1930s as a solvent in the production of plastics and cellulose acetate. Starting in the 1950s, 1,4-dioxane was utilized as a stabilizer that were chlorinated. Since 1,4-Dioxane is a contaminant, there are further worries regarding the amount of this substance in shampoos, detergents, and cosmetics. To novel derivatives based on physicochemical properties i.e., melting point, Rf value, FTIR, NMR and Mass spectroscopy techniques and evaluated for in-vitro anticancer potential of synthesized novel derivatives on following cell lines i.e., PC3, MCF-7. For the 1,4-dioxane derivatives, the boiling point was determined as 128ºC ºC, 146ºC, 164ºC, 144ºC, 158ºC and 172ºC for compounds C1, to C6, respectively. Significant findings were also made regarding molecular weight in the produced 1,4-dioxane derivatives. Compounds C1 to C6 were determined to have molecular weights of 155.5, 247, 135.1, 137.1, 200, and 151.1, respectively. At concentration (100µg/ml), 1,4-dioxane derivatives demonstrated the % cytotoxicity as 73.53%, 74.23%, 72.24%, 75.48%, 74.62%, and 76.16%, in the C1, C2, C3, C4, C5 and C6., respectively. Thus, all the derivatives were shown the significant anti-cancer effect when observed in the PC3 cells. Thus, it might be said that effects of 1,4-dioxane derivatives were almost similar in both the models i.e., PC3 cell and MCF-7 cells. It concluded that the novel derivatives of 1,4-dioxane have a potential anti-cancer activity when observed in-vitro. All the derivatives exhibited anti-cancer response in the both the models i.e., PC3 cells and MCF-7 cells with % cytotoxicity of 70-80%, approximately at the concentration of 100µg/ml.