This work investigates the application of a "Process Analytical Technology" (PAT) analyser to control the substrate concentration over traditional sequential batch control for an industrial scale fed-batch penicillin fermentation. A simulation that utilises the historical data from four batches, where a sequential batch control strategy was implemented, was used as the benchmark reference for this comparison. The simulation accurately predicts the main outputs variables of biomass and penicillin, given the inputs from the historical data set. The simulation includes a PAT analyser, used to build a calibration model with the available off-line substrate concentration from one of the batches. The prediction from this calibration model was used as the controlled variable within a proportional integral (PI) controller to manipulate the substrate feed rate for the three remaining batches. Performance of each control strategy was analysed by comparing the final penicillin yield of each batch. An increase of 35, 20 and 9% was observed for the three batches controlled using the PI controller compared with the sequential batch control strategy.