| Budget Amount *help |
¥4,810,000 (Direct Cost: ¥3,700,000、Indirect Cost: ¥1,110,000)
Fiscal Year 2011: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000)
Fiscal Year 2010: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000)
Fiscal Year 2009: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
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| Research Abstract |
(1) We developed a simultaneous HPLC method for practical and rapid determination of coenzyme A, dephospho-coenzyme A, and acetyl-coenzyme A in tissues. These coenzymes are biosynthesized from a vitamin pantothenic acid, which are involved in the metabolism of fatty acids, amino acid catabolism and other many nutrients. The method employs a Tosoh TSK-GEL ODS-100V(250 mm×4. 6 mm I. D., particle size 5μm) column eluted with 100 mmol/L NaH_2PO_4 and 75 mmol/L CH_3COONa(pH was adjusted to 4. 6 by addition of concentrated H_3PO_4)-acetonitrile(94 : 6, v/v) at a flow-rate of 1. 0 mL/min. The UV detector was set at 259 nm. The analysis time of one sample was 25 min. The quantification limits for coenzyme A, dephospho-coenzyme A, and acetyl-coenzyme A were all 10 pmol.
(2) We challenged to detect in vitro the activity of a hypothetical enzyme pantethine adenylyltransferase, but we could not detect the enzyme activity.
(3) The technique was applied to the analyses of several tissues of rats fed normal and pantothenic acid-free diets. The present results show that the technique was suitable for the simultaneous determination of coenzyme A, dephospho-coenzyme A, and acetyl-coenzyme A in liver, heart, kidney, spleen, testis, large colon, and muscle, but not for small intestine.
(4) The rats made by feeding a pantothenic acid-free diet for 47 days were fed either the diet containing pantothenic acid or pantethine to compare which compounds are suitable for recovery from the pantothenic acid-deficiency. The indices for various pantothenic acid nutritional status revealed that pantothenic acid and pantethine had exactly the same activity as precursors of CoA.
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