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Isfahan University of Medical Sciences
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| Document Type
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Latin Dissertation
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| Language of Document
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English
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| Record Number
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102537
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| Doc. No
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T9037
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QV,354,A138O,2004
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| Main Entry
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Abedi, Daryoush
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| Title & Author
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Optimization of ampicilin production using an immobilized penicillin- G acylase enzyme/ Daryoush Abedi
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| College
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Schools, Parmacy
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| Date
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, 2004
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| Degree
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Pharmaceutical, Ph.D
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| Page No
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153 p.: fig, tab
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| Note
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This thesis is also research project with project ID 80013
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داریوش عابدی
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| Abstract
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Introduction: Chemical methods used to be the only ones for production of semi-synthetic antibiotics, in which very low temperatures (-30 ░C), non-aqueous conditions and using toxic compounds (like pyridine and dimethylaniline) were necessary. But, nowadays, by using penicillin acylase, there is no need for these conditions. Penicillin acylases catalyze two-way reaction of attachment and removal of acyl branch to and from 6aminopenicillanic acid (6-APA). In order to use this enzyme in industrial scale, we need to immobilize it.The main objective of this project was to optimize penicillin acylase (extracted from E. coli TA1) application as a biocatalyst in the synthesis of ampicillin from the substrates 6aminopenicillanic acid (6-APA) and phenylglycine methyl ester (PGME), in order to contribute to the science in further optimization of this biosynthesis, and introduce new approaches to our antibiotic production industries inside our country with more details than that can be found from the relevant text books and papers.Methods: The cells were disrupted using three different methods: sonication, homogenization and osmotic shock and the cell extracts were concentrated using ammonium sulfate.The free enzyme was immobilized by a new method called CLEA. E. coli TA1 cells were also immobilized by entrapment in 2 agar. Synthetic and hydrolytic activity of the enzyme in these two immobilization methods were compared with these activities in the free enzyme and the commercially available immobilized and free enzymes.The enzyme activity was optimized in all forms of the immobilized and free enzyme considering temperature, pH, biocatalyst and substrates concentrations.Results and Discussion: The optimum pH was 5.5-6 for synthetic activity and 7.5-8 for hydrolytic activity. The optimum temperature was 25░C for synthetic activity and 40-45░C for hydrolytic activity. The optimum biocatalyst concentration was 6 U/ml. The best ratio of the substrates (6-APA:PGME) for production of ampicillin was 1:3.In all different conditions of pH, temperature and concentration, the immobilized enzyme by CLEA method showed better activities in different forms of the biocatalyst, except the free enzyme. The maximum conversion percent for the optimum condition of the free enzyme, CLEA, PGA-450 and the immobilized cells were 88 ,84 ,81 and 78, respectively.Conclusion: Immobilization of penicillin acylase by CLEA method was managed successfully and it was shown that the product acts even better than the commercial one. Comparison of the different forms of the enzyme and optimization of the activities, especially ampicillin production are the other important outcomes of this research, which can be used in further researches in this field..
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| Descriptor
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Osmotic Pressure
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Immunoenzyme Techniques
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Cells
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Ampicillin
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Penicillins
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Hydrogen
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Chromatography, High Pressure liquid
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Antibiotics
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| Added Entry
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Fazeil , Mohammad Reza , استاد راهنما
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Jaffarian Dehkordi, Abbas , استاد راهنما
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http://elib.mui.ac.ir/site/catalogue/102537
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