Production of sourdough bread is a natural biotechnological process. For fulfilling the increasing consumer demand for healthier and natural foods, sourdough bread was produced by using the multigrain flour. During this study, different wheat varieties were analyzed to assess their suitability for the production of sourdough bread. On the basis of physico-chemical, rheological and sensory characteristics of the sourdough bread, wheat variety Millat-2011 was found the most suitable for the production of sourdough bread. Consequently, this variety was milled to get the straight grade flour for further use in multigrain sourdough bread. The anti-nutritional factors in Indian vetch were determined after steeping and fermentation. Both these processes significantly reduced the level of anti-nutritional factors. Sourdough multigrain bread was prepared by using the barley, Indian vetch and wheat. The levels of barley and Indian vetch in composite flour were optimized by using response surface methodology and their effects on the textural and sensory properties were studied. The optimized levels of barley (16.48%) and Indian vetch (12.35%) in multigrain bread formulation generated through RSM resulted in sensory acceptable multigrain bread. After this, the processing parameters (time and temperature) were optimized by using the response surface methodology. The increase in fermentation time and temperature significantly affected the textural and sensory properties of bread and the best quality attributes were obtained at a temperature and time combination of 29ºC and 19 h, respectively. Sourdoughs were analyzed for pH, acidity, organic acids and volatile compounds. Optimized multigrain breads were analyzed for physic-chemical parameters such as texture, proximate composition, organic acids, volatile compounds, in-vitro protein and starch digestibilities, glycemic index, minerals and amino acid profiles. All these parameter except mineral contents were significantly affected by the time and temperature of sourdough fermentation. The in-vitro starch digestibility and glycemic index of the body were found to reduce and in-vitro protein digestibility was improved. The multigrain sourdough bread prepared at 29ºC for 19h was found best regarding the sensory attributes than the yeast leavened bread.