| Project/Area Number |
60460221
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
Physical pharmacy
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| Research Institution | School of Pharmaceutical Sciences, Showa University |
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Principal Investigator |
TSUJI Akio Professor, 薬学部, 教授 (80053784)
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| Co-Investigator(Kenkyū-buntansha) |
TAKEDA Motoko Assistant, 薬学部, 助手 (70053973)
MAEDA Masako Associate Professor, 薬学部, 助教授 (00053869)
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| Project Period (FY) |
1985 – 1987
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| Project Status |
Completed (Fiscal Year 1987)
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| Budget Amount *help |
¥3,900,000 (Direct Cost: ¥3,900,000)
Fiscal Year 1987: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1986: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1985: ¥2,500,000 (Direct Cost: ¥2,500,000)
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| Keywords | Fluorescence high-performance liquid chromatography / chemiluminescence high-performance liquid chromatography / congenital hypothyroidism / congenital adrenal hyperplasia / inborn error / 3 -hydroxysteroid dehydrogenase / corticosteroid / bile acid / fatty acid / lucigenin / 化学発光HPLC |
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| Research Abstract |
In this project, we have developed highly sensitive HPLC methods for biological substances related to inborn errors; congenital hypothyroidism and congenital adrenal hyperplasia etc.. The results are summarized as follows :
1. Fluorescence HPLC method using an immobilized 3 -hydroxysteroid dehydrogenase column as a post-column enzyme reactor has been developed for the determination of corticosteroids and their metabolites in the urine of subjects with congenital adrenal hyperplasia. Each steroid was measured at the 2.5 ug/dl at the highest sensitivity of the detector. The mean recoveries and reproducibilities were 91.5 - 108.2% with 0.9 - 6.5% (CV%).
2. Chemiluminescence HPLC method based on the chemiluminescence reaction of lucigenin has been developed for the determination of corticosteroids and their metabolites, and phenacyl esters of bile acids. The detection limits of steroids were about 50 pmol - 1 nmol. The CV% was 4.0 - 9.7%.
3. Chemiluminescence HPLC method using N-(4-aminobutyl)-N-ethylisolumionl as a pre-column reagent has been developed for the determination of bile acids. The chemiluminescent derivative of cholic acid was 20 fmol. This value was higher than that of chemiluminescence HPLC using 3 -hydroxysteroid dehydrogenase reactor.
4. Chemiluminescence determination of NADH using flow injection system and its application to HPLC detection system have been developed. Bile acids and related 3 -hydroxysteroids could be assayed by this method.
5. The HPLC method for the determination of free fatty acids in plasma by using phenacyl bromide as a pre-column reagent has also developed.
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