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Lactobacillus paracasei HD1.7 immobilization methods and Optimization

Author: ChenJianWei
Tutor: LiXiuLiang
School: Heilongjiang University
Course: Microbiology
Keywords: Lactobacillus paracasei HD1.7 immobilization bacteriocin optimization
CLC: TS252.1
Type: Master's thesis
Year: 2009
Downloads: 111
Quote: 1
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Abstract

The Lactobacillus paracasei HD1.7 which was obtained by our lab could produce bacterocin, which had antimicrobial activity. Two immobilized methods were seperately employed to optimize immobilized and fermentation conditions in order to improve bacteriocin production, decurtate fermentation period and save cost. It would establish foundation for the further study of bacteriocin and finally extraction.Two better immobilized methods-calcium alginate and PVA-Alginate gel were selected by considering their property and cost of immobilization.In the method of calcium alginate gel, the concentration of sodium alginate and CaCl₂, immobilized time, inoculation volume , culture time and the density of entrapped cells were determined. A 26?2 two-level fractional factorial design was employed first where 6 variables were studied for their influence on bacteriocin production. The results showed that the concentration of sodium alginate, inoculation volume and culture time were the most significant variables to improve bacteriocin production. The greatest response area was estimated by the design of steepest ascent approach. A three-level Box–Behnken factorial design was employed for maximizing the bacteriocin production and the number of cell cycle. The optimal conditions for bacteriocin production and the number of cell cycle were with sodium alginate 2.8%, CaCl₂ 5%, immobilization time 4h, the density of entrapped cells 1/20, culture time 63h and inoculation volume 8.2%. On the basis of immobilization, the repeated batch fermentation using immobilized cells was studied, the first batch fermentation was 63h, after that, the period of the fermentation was changed to 24h, bacteriocin production was about 410AU/mL and was 91% of the production of free-cell fermentation. Under the optimal conditon of batch fermentation, the total production of bacteriocin was increased by 1.6 times during the period of 375hIn the method of PVA-Alginate gel, the concentrations of PVA, sodium alginate, boracic acid and CaCl₂, immobilized time, inoculation volume, culture time and the density of entrapped cells were determined. A Plackett-Burman design was employed first where 8 variables were studied for their influence on bacteriocin production. The results show that inoculation volume, culture time, the concentration of boric acid and PVA were the most significant variables to improve bacteriocin production. The greatest response area was estimated by the design of steepest ascent approach. A four-level Box–Behnken factorial design was employed for maximizing the bacteriocin production and the number of cell cycle. The optimal condition for bacteriocin production and the number of cell cycle was with inoculation volume13.8%, culture time 70.5h, the concentration of boric acid 4.8%, the concentration of PVA 9.45%. On the basis of immobilization, the repeated batch fermentation using immobilized cells was studied, the first batch fermentation was 70h, after that, the period of the fermentation was changed to 27h, bacteriocin production was 67% of the production of free-cell fermentation. Under the optimal condition of batch fermentation, the total production of bacteriocin was increased by 1.15 times during the period of 610h.Compared with cost, bacteriocin production and fermentation period of calcium alginate and PVA-Alginate gel, the method of calcium alginate gel was proved to be better.

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