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Enzyme sources for urinary oxalate measurements

Rahim, Rukhsana (2007) Enzyme sources for urinary oxalate measurements. Masters thesis, University of Huddersfield.

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    Abstract

    An alternative source of oxalate oxidase (Oxox) was to be purified for use in a biosensor
    system to detect oxalate in patients suffering from primary hyperoxaluria (PH). Oxox has
    been isolated from a number of sources, however there is significant variation in the
    information available. Prior to the purification of Oxox, the properties of commercially
    available Sigma Aldrich Ltd. Oxox were researched.
    An optimum enzymatic assay, substrate specificity, optimum buffer and optimum pH for
    Sigma Oxox were investigated prior to its purification. The purification of the enzyme led
    to the finding that Sigma Oxox was unstable in a number of salt buffers, and insoluble. To
    determine the ability of this enzyme to detect oxalate, amperometric analysis was
    undertaken, which demonstrated the Sigma Oxox based sensors’ reusability and
    reproducibility.
    Oxox from barley roots was successfully purified using a five step purification protocol,
    the findings obtained were in accordance with those published. The barley root extracts,
    after each purification procedure, were analysed amperometrically and highlighted the
    requirement for purification to optimise the analytical signal.
    Over 140 sources were screened from a number of plant families, to detect the presence of
    Oxox activity. Three possible sources of Oxox were identified by level of Oxox activity
    and chosen for further purification: cabbage, carrot and mint leaves. The presence of
    Oxox in different varieties of these sources varied, illustrating the dissimilarities in Oxox,
    confirming the diversity of the data available on Oxox.
    Oxox from cabbage, carrot and mint leaves were purified using the standardised protocol
    employed during the purification of Sigma and barley root Oxox. The studies undertaken
    led to the finding that the components of interest possessed dissimilar properties to
    documented Oxox. Cabbage, carrot and mint leaves oxalate oxidative components were
    found to be smaller than 700 kDa, extremely thermally stable, did not possess strong
    positively or negatively charged groups.
    The identity of the oxalate oxidising components being Oxox was disproved. The
    possibility of the oxidising components being cofactors, or novel oxalate oxidising
    components were addressed, however, no distinct conclusion could be made.

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    Item Type: Thesis (Masters)
    Additional Information: © The Author 2007
    Uncontrolled Keywords: enzyme sources urinary oxalate measurements
    Subjects: Q Science > Q Science (General)
    Q Science > QD Chemistry
    Q Science > QR Microbiology
    Schools: School of Applied Sciences
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    Depositing User: Sara Taylor
    Date Deposited: 20 Dec 2007
    Last Modified: 28 Jul 2010 19:21
    URI: http://eprints.hud.ac.uk/id/eprint/353

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