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Title:Application of high-performance liquid chromatography combined with ultra-sensitive thermal lens spectrometric detection for simultaneous biliverdin and bilirubin assessment at trace levels in human serum
Authors:ID Martelanc, Mitja, Univerza v Novi Gorici (Author)
ID Žiberna, Lovro, Univerza v Trstu (Author)
ID Passamonti, Sabina, Univerza v Trstu (Author)
ID Franko, Mladen, Univerza v Novi Gorici (Author)
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Language:English
Work type:Not categorized
Typology:1.01 - Original Scientific Article
Organization:UNG - University of Nova Gorica
Abstract:We present the applicability of a new ultra sensitive analytical method for the simultaneous determination of biliverdin and bilirubin in human serum. The method comprises isocratic reversed-phase(RP) C18 high-performance liquid chromatography (HPLC) and thermal lens spectrometric detection (TLS)based on excitation by a krypton laser emission line at 407 nm. This method enables the separation of IX-α biliverdin and IX-α bilirubin in 11 min. with limit of detection (LOD) and limit of quantitation (LOQ) for biliverdin of 1.2 nM and 3 nM, and 1nM and 2.8 nM for bilirubin,respectively.In addition, a step-gradient elution was set up, by changing the mobile phase composition, in order to further enhance the sensitivity for bilirubin determination with LOD and LOQ of 0.5 nM and 1.5 nM, respectively. In parallel, an isocraticHPLC-DAD method was developed for benchmarking against HPLC-TLS methods. The LOD and LOQ forbiliverdin were 6 nM and 18 nM, and 2.5 nM and 8nM for bilirubin,respectively. Additionally, both isocratic methods were applied for measuring biliverdin and free bilirubin in human serum samples (from 2 male and 2 female healthy donors). Combining isocratic HPLC method with TLS detector was crucial for first ever biliverdin determination in serum together with simultaneous free bilirubin determination. We showed for the first time the concentration ratio of free bilirubin versus unbound biliverdin in human serum samples.
Keywords:Bilirubin, biliverdin, serum, HPLC, TLS
Publication version:Author Accepted Manuscript
Year of publishing:2016
Number of pages:7
Numbering:154
PID:20.500.12556/RUNG-2220 New window
COBISS.SI-ID:4178683 New window
DOI:10.1016/j.talanta.2016.03.053 New window
NUK URN:URN:SI:UNG:REP:9HIVYATN
Publication date in RUNG:29.03.2016
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Record is a part of a journal

Title:Talanta
Shortened title:Talanta
Publisher:Elsevier
Year of publishing:2016
ISSN:0039-9140

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