18 February / March 2020
Figure 4: Representative LC/MS chromatogram of thyroids in human serum with RPA on-line cartridge.
Comparing the two types of on-line cartridges, RPA appears to deliver greater signals (peak height and area) for all three thyroid analytes compared to the C8 cartridge. The RPA phase has amide groups that are known to form hydrogen bonds and can provide better retention for analytes through hydrogen bonding interactions. Although the retention times and peak widths for thyroid hormones on the analytical column is very similar whether using the RPA or the C8 SPE cartridges, there appears to be a slight advantage of using RPA SPE with regards to the analyte’s retention from the direct injection of protein precipitated serum samples.
Summary
An on-line SPE-LC/MS method has been developed for the rapid detection of thyroid hormones in human serum with minimal
Table 1: Ruggedness of the system with C8 Cartridge.
hands-on effort and time-consuming steps. Both C8 and RP-Amide on-line cartridges were shown to be capable of capturing trace amounts of thyroids from protein precipitated human serum samples. All three thyroid analytes, T3, rT3 and T4 were resolved on a Biphenyl column, with sharp and symmetrical peak shapes. In addition, reproducibility (RSD%) of the retention time of the thyroids from 120 consecutive injections is between 0.1% and 0.2%, with either C8 or RPA on-line cartridges, while the peak area reproducibility (RSD%) is between 5.1% and 7.7%. These RSD’s indicate great ruggedness of the on- line SPE-LC/MS system.
References
1. Offie P. Soldin, Stevem J. Soldin, thyroid hormone Testing by Tandem Mass Spectrometry. Clinical Biochemistry, 2011; 44: 89-94.
2. Kahric-Janicic N, Soldin SJ, Soldin OP, West T, Gu J, Jonklaas J, Tandem mass spectrometry improves the accuracy of free thyroxine measurements during pregnancy. Thyroid. 2007;17(4): 303-11.
3. Dongli Wang and Heather M. Stapleton, Analysis of thyroid hormones in serum by liquid chromatography-tandem mass spectrometry. Anal Bioanal Chem. 2010; 397(5): 1831–1839.
4. Susan S-C. Taia, Lorna T. Sniegoski and Michael J. Welch, Candidate Reference Method for Total Thyroxine in Human Serum Use of Isotope-Dilution Liquid Chromatography-Mass Spectrometry with Electrospray Ionization. Clinical Chemistry, 2002; 48(4): 637-642.
Table 2: Ruggedness of the system with RPA Cartridge.
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