Poster Presenter

Highly Sensitive Assay of Metallothinein And Hepcidin By HPLC-NAM Fluorometry
Tsuneo Kamata, Masato Yamaguchi And Hiroshi Meguro
Japan

We have been studying a micro assay of SH compounds such as GSH, carnitine, and coenzyme A at nano-mole to pico-mole level by N-(9-acridinyl) maleimide (NAM), a detective reagent for the sulfhydryl group (SH). And we have assayed the enzyme activity including glutathione peroxidase, glutathione reductase, carnitine acethyl- transferase and cholinesterase Also dehydrogenazes could be assayed by coupling with glutathione reductase. The sensitivity of this method is 250 times higher than ordinary colorimetry or UV method.

NAM has many analytical significances; the reagent has no fluorescence, reacts with SH compounds in an aqueous solution above pH 5.5, and forms an SH-NAM complex that emits strong fluorescence. The FI of the complex is directly proportional to the number of SH. NAM is so bulky that it does not react with most proteins, NAM is so bulky that it does not react with most proteins, but forms fluorescent complexes with lower molecular proteins such as metallothionein and hepcidim. These are usually detected by HPLC with atomic absorption. We are planning more sensitive micro assay of these proteins by HPLC with NAM fluorometry in order to diagnose the poisoning of heavy metals, chemicals or stresses.