Effects of pH and feeding strategy on metabolite profiling of beta-lactamase producing Bacillus licheniformis

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Tezin Türü: Yüksek Lisans

Tezin Yürütüldüğü Kurum: Orta Doğu Teknik Üniversitesi, Mühendislik Fakültesi, Kimya Mühendisliği Bölümü, Türkiye

Tezin Onay Tarihi: 2005

Öğrenci: NAZAR İLERİ

Danışman: PINAR ÇALIK

Özet:

In this study, the effects of pH and different feeding modes on beta-lactamase production and cell metabolism were investigated with Bacillus licheniformis (ATCC 25972). For this purpose, first, the effects of pH on beta-lactamase activity, cell formation, substrate consumption, as well as intracellular sodium, potassium, ammonium ion, amino acid and organic acid concentrations were investigated in V= 3.0 dm3 batch bioreactors consisting of temperature, pH, foam, stirring rate and dissolved oxygen controls. Among the investigated uncontrolled pH operation with pH0=7.5 and controlled pH operations, pHc=6.75 yielded the highest cell concentration and beta-lactamase activity as Cx=0.60 kg/m3 and A= 54 U/cm3, respectively. Next, the production medium was redesigned in terms of initial glucose and phosphate ion concentrations in order to increase the enzyme activity and cell growth rate, and to determine the feeding strategy in laboratory scale batch-bioreactors using shake bioreactors having V=33 ml working volumes. The medium containing (kg/m3), glucose 2.5 (7.0); Na2HPO4, 1.0; K2HPO4 1.0; (NH4)2HPO4, 7.1 and salt solution at pHc=6.75 was accepted as optimized medium for fed-batch (batch) processes. Using this optimized medium the feeding strategy was investigated for linear and exponential feeding profiles and compared with batch operation. Throughout the fermentation, cell, substrate and intracellular and extracellular by-product, sodium, potassium, ammonium ion concentrations, beta-lactamase activity, yield coefficients, specific rates, oxygen uptake rates and liquid phase mass transfer coefficient values were determined. The highest beta-lactamase activity was obtained at fed-batch operation with exponential feeding (FB1) condition as A= 108 U/cm3, which is ca. 1.7-fold higher than that of the batch operation with optimized medium. Finally, to invesitigate the