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Poly-γ-glutamic acid ( γ -PGA) is a versatile biopolymer with widespread applications in the food, pharmaceutical, and medical industries. One of the main challenges in expanding  γ -PGA industrial applications is the high cost of production. Developing an efficient and low-cost fermentation process such as bacterial cultivation with pulsed feeding can significantly reduce production costs. Thus, initially, a new pulsed-feeding strategy of citrate and glutamate was developed for  γ -PGA production enhancement in the fed-batch culture of  Bacillus licheniformis  ATCC 9945 a . Then, the effects of pulse number, feeding amount, feeding times, the addition time of calcium and manganese solutions, the pH of the added citrate solution, and the concentration of feed stock solutions of pulse-feeds on  γ -PGA production were investigated. Under optimal conditions: feeding two pulses at 8 and 24 hours of culture, 20 g citrate and glutamate per liter of culture medium per pulse (about 52 mL of each of citrate and glutamate feeding solutions prepared with a concentration of 384 g/L by adding distilled water) about 88 ± 4 g/L of  γ -PGA was obtained. It is one of the highest values ever reported for  γ -PGA production with  Bacillus licheniformis  ATCC 9945 a , of course with a much simpler process than the other fed-batch processes.

Enhanced production of poly-γ-glutamic acid by Bacillus licheniformis ATCC 9945a using simultaneous pulse-feedings of citrate and glutamate