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Determination of the protein content of complex samples by aromatic amino acid analysis, liquid chromatography-UV absorbance, and colorimetry

  • Fast and accurate determination of the protein content of a sample is an important and non-trivial task of many biochemical, biomedical, food chemical, pharmaceutical, and environmental research activities. Different methods of total protein determination are used for a wide range of proteins with highly variable properties in complex matrices. These methods usually work reasonably well for proteins under controlled conditions, but the results for non-standard and complex samples are often questionable. Here, we compare new and well-established methods, including traditional amino acid analysis (AAA), aromatic amino acid analysis (AAAA) based on the amino acids phenylalanine and tyrosine, reversed-phase liquid chromatography of intact proteins with UV absorbance measurements at 220 and 280 nm (LC-220, LC-280), and colorimetric assays like Coomassie Blue G-250 dye-binding assay (Bradford) and bicinchoninic acid (BCA) assay. We investigated different samples, including proteins withFast and accurate determination of the protein content of a sample is an important and non-trivial task of many biochemical, biomedical, food chemical, pharmaceutical, and environmental research activities. Different methods of total protein determination are used for a wide range of proteins with highly variable properties in complex matrices. These methods usually work reasonably well for proteins under controlled conditions, but the results for non-standard and complex samples are often questionable. Here, we compare new and well-established methods, including traditional amino acid analysis (AAA), aromatic amino acid analysis (AAAA) based on the amino acids phenylalanine and tyrosine, reversed-phase liquid chromatography of intact proteins with UV absorbance measurements at 220 and 280 nm (LC-220, LC-280), and colorimetric assays like Coomassie Blue G-250 dye-binding assay (Bradford) and bicinchoninic acid (BCA) assay. We investigated different samples, including proteins with challenging properties, chemical modifications, mixtures, and complex matrices like air particulate matter and pollen extracts. All methods yielded accurate and precise results for the protein and matrix used for calibration. AAA, AAAA with fluorescence detection, and the LC-220 method yielded robust results even under more challenging conditions (variable analytes and matrices). These methods turned out to be well-suited for reliable determination of the protein content in a wide range of samples, such as air particulate matter and pollen.zeige mehrzeige weniger

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Autor*innen:K. Reinmuth-Selzle, Teodor Tchipilov, A. T. Backes, Georg TscheuschnerORCiD, K. Tang, K. Ziegler, K. Lucas, U. Pöschl, J. Fröhlich-Nowoisky, Michael G. WellerORCiD
Dokumenttyp:Zeitschriftenartikel
Veröffentlichungsform:Verlagsliteratur
Sprache:Englisch
Titel des übergeordneten Werkes (Englisch):Analytical and Bioanalytical Chemistry
Jahr der Erstveröffentlichung:2022
Organisationseinheit der BAM:1 Analytische Chemie; Referenzmaterialien
1 Analytische Chemie; Referenzmaterialien / 1.5 Proteinanalytik
Veröffentlichende Institution:Bundesanstalt für Materialforschung und -prüfung (BAM)
Verlag:Springer Nature Limited
Verlagsort:New York, Heidelberg
Erste Seite:1
Letzte Seite:14
DDC-Klassifikation:Naturwissenschaften und Mathematik / Chemie / Analytische Chemie
Freie Schlagwörter:150th anniversary of BAM; 280 nm; Air filter samples; Air particulate matter; Aromatic amino acid analysis; Atmospheric aerosol; BCA test; Bradford; Chemical protein modification; Chromatography; Derivatization; Fluorescence; HPLC; Hydrolysis; Kjeldahl; LC-UV absorbance; Nitration; Nitrotyrosine; Phenylalanine; Pollen extract; Protein content; Protein quantification; Protein test; Topical collection: Analytical Methods and Applications in the Materials and Life Sciences; Tyrosine
Themenfelder/Aktivitätsfelder der BAM:Chemie und Prozesstechnik
Chemie und Prozesstechnik / Chemische Charakterisierung und Spurenanalytik
DOI:10.1007/s00216-022-03910-1
URN:urn:nbn:de:kobv:b43-545924
URL:https://pubmed.ncbi.nlm.nih.gov/35320366/
Verfügbarkeit des Dokuments:Datei für die Öffentlichkeit verfügbar ("Open Access")
Lizenz (Deutsch):License LogoCreative Commons - CC BY-NC - Namensnennung - Nicht kommerziell 4.0 International
Datum der Freischaltung:04.04.2022
Referierte Publikation:Ja
Datum der Eintragung als referierte Publikation:04.04.2022
Schriftenreihen ohne Nummerierung:Wissenschaftliche Artikel der BAM
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