Title: [18] Highly sensitive methods for assaying the enzymes of vitamin B6 metabolism
Abstract: This chapter describes highly sensitive methods for assaying the enzymes of vitamin B6 metabolism. The B6 vitamers absorbed by mammals from the diet are pyridoxine, pyridoxamine, and pyridoxal. All three are phosphorylated by a single kinase, and pyridoxine and pyridoxamine 5′-phosphates are converted to pyridoxal 5′-phosphate (PLP) by a flavoprotein oxidase. As the liver is a major site of vitamin B6 metabolism and contributes most of the PLP found in plasma, therefore it has been used for most studies of these enzymes. The assays are optimized for small samples of human liver, but can be applied to other tissues and animals. All assay procedures involving pyridoxyl compounds are conducted in dim light to avoid photodegradation. The most sensitive assay for pyridoxal kinase is to measure the formation of [3H]pyridoxine 5′-phosphate, which can be easily separated from [3H]pyridoxine using DEAE-cellulose columns or paper disks. It is also possible to use pyridoxal as the substrate and quantitate the PLP enzymatically or by preparation of fluorescent derivatives.
Publication Year: 1986
Publication Date: 1986-01-01
Language: en
Type: article
Indexed In: ['crossref', 'pubmed']
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Cited By Count: 6
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