The present study was carried out to enhance the genetic variability in Kinnow mandarin (Citrus nobilis Loureiro Physiological and biochemical responses of Kinnow mandarin (Citrus nobilis × Citrus deliciosa) to EMS induced mutagenesis Citrus deliciosa Tenora) using ethyl methanesulfonate (EMS) during 2016-17 and 2017-18. Plants created with different doses of EMS concentration, viz. 0.05%, 0.1%, 0.2% and 0.5% were examined for the various physiological and biochemical alterations in putative mutants aged six years and compared with the wild type (WT). A dose-dependent decrease in leaf area was recorded in the mutants with increasing doses of EMS concentration and as compared to WT, maximum reduction of 135.52 and 104.92% was witnessed in the mutants E-19 and E-17 developed from 0.5% EMS, whereas the leaves were more succulent in E-16. Chlorophyll a content was higher than chlorophyll b in the mutated population. As compared to WT, maximum decrease in chlorophyll a, total chlorophyll was noticed in the mutants E-19 generated from 0.5% EMS. Contrary to the physiological alterations, the total phenol, proline and total protein content in the mutants were upregulated by nearly two fold in the mutants developed beyond 0.1% EMS. Significant increase was also noticed in the antioxidant enzymes SOD, CAT, POX and GR activity in the mutants E-16 to E-20 induced from 0.5% EMS. The variation in traits of interest observed in the present study such as better leaf succulency and upregulated antioxidant enzyme activity in the mutant E-16 (0.5% EMS) suggests that the developed mutant could be utilised in Kinnow improvement for abiotic stress tolerance.