The research work titled “Biochemical and Metabolic Studies of Wheat (Triticum aestivum L.) in Response to Salinity Under Laboratory and Field Conditions” aimed to screen 10 wheat genotypes Kharchia-65, KRL-3-4, FLW-16, PBW-757, DBW-252, FLW-2, FLW-1, DBW-11, HD-2851, and FLW-5 for salinity stress tolerance. Salinity stress treatments were administered at 25mM, 50mM, 75mM, and 100mM, along with a control at the Instructional Farm of Acharya Narendra Deva University of Agriculture & Technology, Kumarganj, Ayodhya. Genetic variation was observed in germination percentage, with a drastic reduction in genotypes FLW-5 and FLW-16 under stress, while Kharchia-65 and KRL 3-4 showed better performance. Seedling shoot length significantly decreased in FLW-1 and FLW-2, and root length in DBW-11 under stress, whereas Kharchia-65 and DBW-252 performed better. Plant height saw a drastic reduction in HD-2851 and FLW-5, while Kharchia-65 and KRL 3-4 maintained better heights. Tiller number significantly reduced in FLW-16, FLW-2, and FLW-1, with Kharchia-65 and KRL 3-4 less affected. Spikelet length drastically reduced in DBW-252, FLW-5, and FLW-2, with Kharchia-65 and KRL 3-4 performing better. Salinity stress at the reproductive stage severely reduced yield, especially in FLW-1, while HD-2851, Kharchia-65, and KRL 3-4 were less affected. Catalase, peroxidase activities and SDS-PAGE increased under stress, with Kharchia showing the highest increments.Proline levels increased significantly, especially in Kharchia-65 and KRL 3-4, while protein levels decreased, with HD-2851 and FLW-5 showing the highest reductions. These findings show the differential response of wheat genotypes to salinity stress, with Kharchia-65 and KRL 3-4 consistently showing better performance across various parameters, providing insights for selecting and breeding wheat varieties with enhanced salinity tolerance.