title: Pyrroloquinoline quinone, a method for its isolation and identification by mass spectrometry.
abstract: Procedures for the unambiguous detection and for the isolation and mass spectrometric identification of pyrroloquinoline quinone (PQQ) are presented. The procedure involved acid hydrolysis of protein in the presence of phenylhydrazine and successive isolation and identification of the formed adduct using mass spectrometry. In HPLC the phenylhydrazone of PQQ gave many methylated products, of which the predominant compound was the pentamethylated derivative. After reaction of the phenylhydrazone derivative of PQQ (PHPQQ) with ammonia, a product was obtained which did not contain phenylhydrazine and which formed a pentamethylated derivative as the main methylation product. The HPLC profiles of the methylated products of PHPQQ and of its ammonia derivative were very characteristic and could be used for identification in addition to mass spectrometry. However, prolonged treatment of proteins with phenylhydrazine during hydrolysis can result in the formation of a material that resembles PQQ in some aspects of its behaviour. Thus, analysis by MS is essential for unambiguous identification. This analytical procedure was applied to pig plasma benzylamine oxidase, pig aorta lysyl oxidase, pig kidney diamine oxidase and bovine serum albumin with negative results. However, samples of pronase contained variable quantities of non-covalently bound PQQ: this can lead to erroneous identification of PQQ in enzyme after pronase digestion.
authors.affiliation: Department of Pharmacology, University of Florence, Italy.
authors.forename: F
authors.initials: F
authors.lastname: Buffoni
authors.forename: S
authors.initials: S
authors.lastname: Cambi
authors.forename: G
authors.initials: G
authors.lastname: Moneti
mesh_terms: D000818:Animals; D002851:Chromatography, High Pressure Liquid; D006868:Hydrolysis; D013058:Mass Spectrometry / Q000379:methods*; D045542:PQQ Cofactor; D010659:Phenylhydrazines; D011506:Proteins / Q000737:chemistry; D015363:Quinolones / Q000032:analysis / Q000302:isolation & purification*; D013552:Swine
chemical_list: D010659:Phenylhydrazines; D011506:Proteins; D015363:Quinolones; C030299:phenylhydrazine; D045542:PQQ Cofactor