In as much as spontaneous fermentation of cowpea leaves enhances product diversification, the process is rather slow with poor product quality. Limited work has been undertaken to provide input toward standardization of the process and enhancing of product quality. The current study sought to evaluate the in‐process fermentative bacteria profile changes due to the effect of optimization of fermentation process of cowpea leaves. Lactic acid bacteria (LAB) isolates from spontaneous and optimized fermentation were characterized using biochemical tests, whereby optimization was done using the Response Surface Methodology model of the central composite design in the Design Expert Software. The RSM models accounted for 89% and 60% variability in the response variables of pH and titratable acidity, respectively (p < .001). Increasing the sugar concentration and period of fermentation significantly (p < .05) increased the titratable acidity, while reducing the pH. The optimal fermentation parameters were established as sugar and salt concentrations of 5% and 2%, respectively, 16 days of fermentation, pH of 3.8 and titratable acidity of 1.22% with a desirability of 0.859. Of the 13 identified LAB isolates, Lactobacillus brevis and Lactococcus lactis dominated the onset stage of spontaneous fermentation whereas only Lactobacillus brevis dominated the onset stage of optimized fermentation. Additionally, the final stage with the dominant isolates of L. plantarum was longer in the spontaneous fermentation process than in the optimized process. Evidently, optimizing the fermentation process resulted in increasing dominance by heterofermenters in the production of soured cowpea leaves, with the yielded product having enhanced acidity.