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Large-scale Enzyme Production. Tina C. Lung Syracuse University Dept. of Chemical Engineering and Material Science CEN 551-Biochemical Engineering January 22, 2004. Examples of Enzymes. Protease (subtilisin, rennet) Hydrolases (pectinase, lipase, lactase) Isomerases (glucose isomerase)

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Large scale enzyme production l.jpg

Large-scale Enzyme Production

Tina C. Lung

Syracuse University

Dept. of Chemical Engineering and Material Science

CEN 551-Biochemical Engineering

January 22, 2004

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Examples of Enzymes

  • Protease (subtilisin, rennet)

  • Hydrolases (pectinase, lipase, lactase)

  • Isomerases (glucose isomerase)

  • Oxidases (glucose oxidase)

  • Produced using overproducing strains of certain organisms.

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Separation and Purification

  • Disruption of cells

  • Removal of debris and nucleic acids

  • Precipitation of proteins

  • Ultrafiltration of the desired enzyme

  • Chromatographic separation

  • Crystallization

  • Drying

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Separation and Purification

  • The process depends on whether or not the enzyme is intra or extracellular.

  • Sometimes inactive (dead or resting) cells are used with desired enzyme activity in immobilized form.

    • No separation and/or purification steps

    • Reduces costs

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Growing Enzymes

  • (1) Cultivate the organisms producing the desired enzymes.

    • Production can be regulated

    • Fermentation conditions ca be optimized for overproduction.

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Enzymes and Sources

  • Proteases

    • Overproducing strains of Bacillus, Aspergillus, Rhizopus, and Mucor.

  • Pectinases

    • Aspergillus niger.

  • Lactases

    • Yeast and Aspergillus.

  • Lipases

    • Certain strains of yeast and fungi.

  • Glucose isomerase

    • Flavobecterium arborescens or Bacillus coagulans

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Growing Enzymes (2)

  • (2) Cell separated from the media usually by filtration or something by centrfugation.

    • Depending on intra/extracellular nature of the enzyme, the cell or fermentation broth is further processed.

    • Recovery of intracellular enzymes is more complicated and involves the disruption of cells and removal of debris and nucleic acids.

      • Increasing permeability of cell membrane (CaCl2 (salt) or dimethylsulfoxide (DMSO) or change in pH

      • Last resort is cell disruption.

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Medical and Industrial Utilization of Enzymes

  • Pharmaceutical Uses

    • Wants chirally pure compounds (IMPORTANT!).

    • Often a particular enantiomer maybe useful while others cause side effects or no effect at all.

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Medical and Industrial Utilization of Enzymes

  • Technological advances

    • Increasing in wider ranges of process conditions.

    • Enzymes grow in unusual environments (eg: deep ocean, salt lakes, and hot springs).

    • New enzymes and better control allow the use of enzymes in more extreme environments.

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  • Number of enzymes made at high volume for industrial purposes evolves more slowly.

  • 1996: sales of industrial enzymes--$372 million.

  • 2006: expected to reach $686 million.

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Table 3.5

Table 3.6

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Industrial Protease

  • Hydrolyze proteins into small peptide groups.

  • Obtained from:

    • Bacteria (Bacillus)

    • Molds (Aspergillus, Rhizopus, and Mucor)

    • Animal pancreas

    • Plants

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Major Uses

  • Cheese making (rennet)

  • Baking

  • Meat tenderization (papain, trypsin)

  • Brewing (trypsin, pepsin)

  • Detergents (subtilisin Carlsberg)

  • Tanning products

  • Medical treatments of wounds

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Hebei Shenzhou Animal Medicine Co., Ltd

  • Specialized manufacturer of zinc bacitracin in China.

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Genencor International

  • Second largest developer and manufacturer of industrial enzymes.

  • Introduced the first industrial scale, recombinant enzyme in 1988.

  • Leader in the areas of protein engineering, expression/secretion technology and enzyme-substrate interaction.

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  • Largest producers of enzymes.

  • World Headquarter: Denmark (1941).

  • More than 500 enzymes in over 130 countries.

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Words of Wisdom

  • “I imagine a future where our biological solutions create the necessary balance between better business, cleaner environment, and better lives.”

    ~Steen Riisgaard, President and CEO