Simultaneous determination of free biliverdin and free bilirubin in serum : a comprehensive LC-MS approach

Albreht, Alen (avtor), Martelanc, Mitja (avtor), Žiberna, Lovro (avtor)

Background: Prognosis, diagnosis, and treatment of several diseases strongly rely on the sensitive, selective, and accurate determination of specific biomarkers in relevant biological samples. Free biliverdin and free bilirubin represent important new biomarkers of oxidative stress, however, the lack of suitable analytical methods for their determination has hindered progress in biomedical and clinical research. Results: Here, we introduce a first comprehensive approach for robust and simultaneous determination of these bilins in serum using liquid chromatography – mass spectrometry (LC-MS). The developed analytical method exhibits linearity for both analytes within the concentration range of 0.5–100 nM, with limits of detection and quantitation determined at 0.1 nM and 0.5 nM, respectively. Moreover, several analytical pitfalls related to the intrinsic molecular structures of free bilirubin and free biliverdin and their trace concentration levels in biological samples are discussed here in detail for the first time. We have shown that the solubility, chemical stability, and affinity of these bilins to various materials strongly depend on the solvent, pH, and addition of stabilizing and chelating agents. Finally, the validated LC-MS method was successfully applied to the analysis of both bilins in fetus bovine serums, yielding higher free bilirubin/biliverdin ratios compared with previously reported values for human serum. Significance: Failure to recognize and address the challenges presented here often leads to substantial analytical errors and consequently biased interpretation of the obtained results. This pertains not only to LC-MS, but also to many other analytical platforms due to the compound-derived sources of error.

• Jezik: Angleški jezik
• Ključne besede: biomedical research, sensitivity, robustness, serum, stability, matrix effects
• Vrsta gradiva: Članek v reviji
• Tipologija: 1.01 - Izvirni znanstveni članek
• Organizacija: MF - Medicinska fakulteta ; FFA - Fakulteta za farmacijo
• Status publikacije: Objavljeno
• Različica publikacije: Objavljena publikacija
• Leto izida: 2024
• Št. strani: 8 str.
• Številčenje: Vol. 1287, art. 342073
• PID: 20.500.12556/RUL-152925
• UDK: 543
• ISSN pri članku: 1873-4324
• DOI: 10.1016/j.aca.2023.342073
• COBISS.SI-ID: 176790787
• Datum objave v RUL: 12.12.2023

Gradivo je del revije

• Naslov: Analytica chimica acta
• Založnik: Elsevier
• ISSN: 1873-4324
• COBISS.SI-ID: 23064325

Licence

• Licenca: CC BY-NC-ND 4.0, Creative Commons Priznanje avtorstva-Nekomercialno-Brez predelav 4.0 Mednarodna
• Povezava: http://creativecommons.org/licenses/by-nc-nd/4.0/deed.sl
• Opis: Najbolj omejujoča licenca Creative Commons. Uporabniki lahko prenesejo in delijo delo v nekomercialne namene in ga ne smejo uporabiti za nobene druge namene.

Sekundarni jezik

• Jezik: Slovenski jezik
• Ključne besede: analizna kemija, bolezni, zdravljenje, biomarkerji, serum