1996 Fiscal Year Final Research Report Summary
A study on a sensitive assay of AKBR for rapid evaluation of pathophysiological state of liver
| Project/Area Number |
08557070
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 試験 |
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| Research Field |
General surgery
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| Research Institution | Shiga University of Medical Science |
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Principal Investigator |
OZAWA Kazue Shiga University of Medical Science, Vice-president, 医学部, 副学長 (00026858)
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| Co-Investigator(Kenkyū-buntansha) |
UNO Shizuo Sanwa Kagaku Kenkyusho CO., LTD.Diagnostics Section of Mie Research Laboratory S, 製薬研究所, 試薬グループ長
TERADA Yasuji Tenri Yorozu soudannsho Hospital, Doctor, 医員 (60243010)
TABATA Rhoko Shiga University of Medical Science, School of Medicine, Associate Professor, 医学部, 助教授 (20155229)
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| Project Period (FY) |
1996
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| Keywords | acetoacetate / beta-hydroxybutyrate / arterial blood ketone body ratio (AKBR) / NADH oxidase |
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| Research Abstract |
A new rapid method for the measurement of arterial blood ketone body concentration and determination of arterial blood ratio (AKBR) was developed last year. This method involves the generation of NADH from NAD^+ in proportion to the concentration of beta-hydroxybutyrate in the presence of beta-hydroxybutyrate dehydrogenase. NADH is reached with oxygen in the presence of NADH oxidase to produce hydrogen peroxide. The oxidative coupling reaction of 3-methyl-2-benzothiazolinone-hydrazone-6-sulfate (MBTH-S) and N-ethyl-N-sulfopropyl-3,5-dimenthylaniline (MAPS) produces a red-purple product in the presence of peroxidase. The concentration of beta-hydroxybutyrate is determined by measuring the increase in intensity of the color. The acetoacetate concentration is determined by completely converting it to beta-hydroxybutyrate, in the presence of beta-hydroxybutyrate dehydrogenase, and then assaying the total beta-hydroxybutyrate concentration. The acetoacetate concentration is obtained by subtraction of beta-hydroxybutyrate concentration from the total ketone body concentration. We have been studying reaction conditions between MBTH-S (or MBTH analog) as a coupler and Trinder's reagent to get a higher sensitivity. MBTH analog ; 2- [(methylsulfonyl) hydrazono] -2,3-dihydro-3-methyl-6-benzothiazole-sulfonic acid (A) 2-hydrazone-2,3-dihydro-3-methyl-6-benzo-thiazolesulfonyl aminoacetic acid (B)
Results Sensitivity using MBTH-S and MAPS in this buffer (pH 8.0) became twofold as high as the one in the current condition, though the increase of the reagent blank was larger, which had a bad influence on the reproducibility. We are studying the method to improve it. Neither A nor B brought higher sensitivity or stability than MBTH-S.
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