This chapter discusses the separation of cobyrinic acid and its biosynthetic precursors by ion-exchange paper chromatography. Cobyrinic acid and its biosynthetic precursors, δ-aminolevulinic acid, uroporphyrinogen III, heptacarboxylic urophorphyrinogen III, and S-adenosylmethionine can be rapidly separated by ion-exchange paper chromatography. This method can serve as a relatively fast assay compared to the thin-layer chromatography of cobyrinic acid heptamethyl ester and phenol extraction of the free acid. The last two methods also suffer from incomplete recovery of cobyrinic acid. This method discussed in the chapter is based on the fact that all the precursors of cobyrinic acid contain nitrogen atoms that can be protonated and retained by a weak, cation-exchange paper. Cobyrinic acid that has a positive charge on the cobalt atom exists as the diaquo form in acidic pH. However, the positive charge is prevented from interacting with the functional groups on the paper by the water ligands, resulting in a rapid elution of cobyrinic acid. This method can be adapted to assay of cobyrinic acid formation in cell-free systems of Propionibacterium shermanii. Incubation products have to be adjusted to pH 3–4 before application to the paper. Series: Methods in Enzymology (Book 67) Hardcover: 579 pages Publisher: Academic Press; Language: English ISBN-10: 0121819671 ISBN-13: 978-0121819675 Product Dimensions: 6 x 9 inches Link Download http://nitroflare.com/view/22F998DAA1DE75Fhttps://drive.google.com/drive/folders/1yLBzZ1rSQoNjmWeJTZ3WGQHg04L1
