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    Reverse phase HPLC method for detection and quantification of lupin seed ?-conglutin

    Access Status
    Fulltext not available
    Authors
    Mane, S.
    Bringans, S.
    Johnson, Stuart
    Pareek, Vishnu
    Utikar, Ranjeet
    Date
    2017
    Type
    Journal Article
    
    Metadata
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    Citation
    Mane, S. and Bringans, S. and Johnson, S. and Pareek, V. and Utikar, R. 2017. Reverse phase HPLC method for detection and quantification of lupin seed ?-conglutin. Journal of Chromatography B. 1063: pp. 123-129.
    Source Title
    Journal of Chromatography B
    DOI
    10.1016/j.jchromb.2017.08.025
    ISSN
    1570-0232
    School
    School of Public Health
    URI
    http://hdl.handle.net/20.500.11937/58052
    Collection
    • Curtin Research Publications
    Abstract

    © 2017 Elsevier B.V. A simple, selective and accurate reverse phase HPLC method was developed for detection and quantitation of ?-conglutin from lupin seed extract. A linear gradient of water and acetonitrile containing trifluoroacetic acid (TFA) on a reverse phase column (Agilent Zorbax 300SB C-18), with a flow rate of 0.8 ml/min was able to produce a sharp and symmetric peak of ?-conglutin with a retention time at 29.16 min. The identity of ?-conglutin in the peak was confirmed by mass spectrometry (MS/MS identification) and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis. The data obtained from MS/MS analysis was matched against the specified database to obtain the exact match for the protein of interest. The proposed method was validated in terms of specificity, linearity, sensitivity, precision, recovery and accuracy. The analytical parameters revealed that the validated method was capable of selectively performing a good chromatographic separation of ?-conglutin from the lupin seed extract with no interference of the matrix. The detection and quantitation limit of ?-conglutin were found to be 2.68 µg/ml and 8.12 µg/ml respectively. The accuracy (precision and recovery) analysis of the method was conducted under repeatable conditions on different days. Intra-day and inter-day precision values less than 0.5% and recovery greater than 97% indicated high precision and accuracy of the method for analysis of ?-conglutin. The method validation findings were reproducible and can be successfully applied for routine analysis of ?-conglutin from lupin seed extract.

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