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Quantitative 1H Nuclear Magnetic Resonance (qNMR) of Aromatic Amino Acids for Protein Quantification

  • Hydrolysis of protein samples into amino acids facilitates the use of NMR spectroscopy for protein and peptide quantification. Different conditions have been tested for quantifying aromatic amino acids and proteins. The pH-dependent signal shifts in the aromatic region of amino acid samples were examined. A pH of 12 was found to minimize signal overlap of the four aromatic amino acids. Several aromatic compounds, such as terephthalic acid, sulfoisophthalic acid, and benzene tricarboxylic acid, were applied as internal standards. The quantification of amino acids from an amino acid standard was performed. Using the first two suggested internal standards, recovery was ~97% for histidine, phenylalanine, and tyrosine at a concentration of approximately 1 mM in solution. Acidic hydrolysis of a certified reference material (CRM) of bovine serum albumin (BSA) and subsequent quantification of Phe and Tyr yielded recoveries of 98% ± 2% and 88% ± 4%, respectively, at a protein concentration ofHydrolysis of protein samples into amino acids facilitates the use of NMR spectroscopy for protein and peptide quantification. Different conditions have been tested for quantifying aromatic amino acids and proteins. The pH-dependent signal shifts in the aromatic region of amino acid samples were examined. A pH of 12 was found to minimize signal overlap of the four aromatic amino acids. Several aromatic compounds, such as terephthalic acid, sulfoisophthalic acid, and benzene tricarboxylic acid, were applied as internal standards. The quantification of amino acids from an amino acid standard was performed. Using the first two suggested internal standards, recovery was ~97% for histidine, phenylalanine, and tyrosine at a concentration of approximately 1 mM in solution. Acidic hydrolysis of a certified reference material (CRM) of bovine serum albumin (BSA) and subsequent quantification of Phe and Tyr yielded recoveries of 98% ± 2% and 88% ± 4%, respectively, at a protein concentration of 16 g/L or 250 µM.zeige mehrzeige weniger

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Metadaten
Autor*innen:Teodor Tchipilov, Klas MeyerORCiD, Michael G. WellerORCiD
Dokumenttyp:Zeitschriftenartikel
Veröffentlichungsform:Verlagsliteratur
Sprache:Englisch
Titel des übergeordneten Werkes (Englisch):Methods and Protocols
Jahr der Erstveröffentlichung:2023
Organisationseinheit der BAM:1 Analytische Chemie; Referenzmaterialien
1 Analytische Chemie; Referenzmaterialien / 1.4 Prozessanalytik
1 Analytische Chemie; Referenzmaterialien / 1.5 Proteinanalytik
Veröffentlichende Institution:Bundesanstalt für Materialforschung und -prüfung (BAM)
Verlag:MDPI
Verlagsort:Basel, Schweiz
Jahrgang/Band:6
Ausgabe/Heft:1
Aufsatznummer:11
Erste Seite:1
Letzte Seite:13
DDC-Klassifikation:Naturwissenschaften und Mathematik / Chemie / Analytische Chemie
Freie Schlagwörter:AAA; AAAA; Amino acid analysis; Aromatic amino acid analysis; Benzene-1,3,5-tricarboxylic acid; Bovine serum albumin (BSA); Calibration; Compound-independent calibration; Histidine; Hydrochloric acid; Internal standard; Metrology; NIST; Phenylalanine; Phenylketonuria; Protein hydrolysis; Quantitative protein analysis; Reference materials; Terephthalic acid; Traceability; Tryptophan; Tyrosine
Themenfelder/Aktivitätsfelder der BAM:Chemie und Prozesstechnik
DOI:10.3390/mps6010011
URN:urn:nbn:de:kobv:b43-570943
Verfügbarkeit des Dokuments:Datei für die Öffentlichkeit verfügbar ("Open Access")
Lizenz (Deutsch):License LogoCreative Commons - CC BY - Namensnennung 4.0 International
Datum der Freischaltung:09.03.2023
Referierte Publikation:Ja
Datum der Eintragung als referierte Publikation:09.03.2023
Schriftenreihen ohne Nummerierung:Wissenschaftliche Artikel der BAM
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