A Bioreactor System for the Determination of Cholesterol Distribution in Separated Human Serum Lipoprotein Classes

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  • MAKINO Keisuke
    Department of Polymer Science and Engineering, Faculty of Textile Science, Kyoto Institute of Technology
  • SASAKI Ichiro
    Department of Polymer Science and Engineering, Faculty of Textile Science, Kyoto Institute of Technology
  • UENISHI Tohru
    Department of Polymer Science and Engineering, Faculty of Textile Science, Kyoto Institute of Technology
  • TAKEUCHI Tamio
    Department of Polymer Science and Engineering, Faculty of Textile Science, Kyoto Institute of Technology
  • HARA Ichiro
    Scientific Instrument Division, Toyo Soda Manufacturing Co. Ltd
  • UMINO Masuo
    Scientific Instrument Division, Toyo Soda Manufacturing Co. Ltd

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Other Title
  • ヒト血中リポタンパクサブフラクションのコレステロール分布測定用バイオリアクターシステム
  • ヒト ケッチュウ リポ タンパク サブフラクション ノ コレステロール ブンプ

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Abstract

In order to determine cholesterol distribution in human serum lipoprotein classes, which has been known to be directly related to heart-attack deseases, a bioreactor-type detection system consisting of cholesterol ester hydrorase (CEH) and cholesterol oxidase (COD) immobilized on size-exclusion chromatographic gel (TSKgel G 6000 PW) was combined with size-exclusion chromatographic separation of the lipoproteins. Glass separation columns packed with G 5000 PW and G 3000 SW were found to be suitable for the separation. CEH and COD were coimmobilized on the gel activated with glutaraldehyde and packed into a glass column, which was connected to the exit of the separation column array. To the inlet of the enzyme column, introduced was staining fluid composed of peroxidase, 4-aminoantipyrine, N-ethyl-N-(2-hydroxy-3-sulfopropy1)-m-toluidine, and Triton X-100. In this system, cholesterol ester in the lipoproteins could be converted into cholestenone and subsequently thus released H202 could be stained in a Teflon reaction coil attached to the exit of the column. Upon using a polymer-coated pump, precisely controlled by the computer, and polymer parts to avoid stainless steel surface in the system, the peaks due to low density and high-density lipoproteins appeared in the chromatogram. The elution profile of the peaks was in good agreement with that obtained by the method reported previously in which a large amount of CEH and COD were consumed as staining reagents. From the elution volumes, the height equivalent to a theoretical plate number, and the peak area of the peaks obtained, types of hyperlipidemia could be recognized.

Journal

  • NIPPON KAGAKU KAISHI

    NIPPON KAGAKU KAISHI 1987 (3), 530-530, 1987-03-10

    The Chemical Society of Japan

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