A time-resolved assay of dopamine β-hydroxylase activity utilizing high-pressure liquid chromatography

✍ Scribed by Natalie B. Feilchenfeld; Helen W. Richter; Walter H. Waddell

• Publisher: Elsevier Science
• Year: 1982
• Tongue: English
• Weight: 429 KB
• Volume: 122
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(82)90260-3

✦ Synopsis

A time-resolved assay of dopamine /3-hydroxylase (EC 1.14.17.1) activity utilizing highpressure liquid chromatography is described. The conversion of tyramine to octopamine by the enzyme was used as a standard reaction. The analytical separation of the assay substrate and product employed a reversed-phase ion-pair chromatographic system, with ultraviolet absorbance detection of eluents at 280 nm. Aliquots of the assay solution were injected directly onto the high-pressure liquid chromatography column and were separated in 6 min total elapsed time, thus permitting time-resolved determination of the product. Quantities of octopamine as small as 20 pmol could be measured. This facile method is more straightforward, convenient, and sensitive than previously published physical and spectroscopic methods of determining dopamine p-hydroxylase activity