A team of researchers from the University of Münster shows that the mercury compound thiomersal added as an adjuvant to influenza vaccine is reacting with the proteins in the vaccine.
Background:Thiomersal (THI), an ethylmercury releasing compound, is added to some vaccines as an adjuvant, mainly to hinder bacterial growth in multidose vials. Ethylmercury (EtHg+) is a potent neurotoxin, with a toxicity resembling that of methylmercury in some aspects but with different toxicokinetics. Introduced in 1931, THI never went through a safety evaluation, but can be used in vaccines because of its introduction before the establishment of such regulation. Banned in the EU since 2001, THI-containing vaccines (TCV) are still in use in the USA and in developing countries.
Mercury species show a high affinity to sulphur and tend to form adducts with biomolecules having thiol groups, such as glutathione (GSH) or cysteine-containing proteins. So far, adduct formation of organic mercury species was observed with GSH, human serum albumin (HSA), hemoglobin and carbonic anhydrase. Previous studies have also shown that adducts of mercury and xenobiotic proteins can lead to a formation of mercury-specific antibodies. This observation leads to the conclusion that adducts of EtHg+ and antigens are not only important to investigate for understanding toxicokinetics, but may also induce a mercury-specific immune response. Anyhow, such unwanted reaction by the use of TCV has not been reported before.
The new study:The researchers from the University of Münster studied the interaction of THI with the seasonal tetravalent influenza vaccine "Vaxigrip tetra" (Sanofi, Paris) containing inactivated split virions of two influenza A and two influenza B strains (each 30 μg mL−1 hemagglutinin (HA), propagated in chicken eggs). After adding different amounts of the adjuvant THI to the vaccine, the vaccine was stored for 20 days in a refrigerator to allow incubation under storage conditions of vaccines.
The incubated samples were analysed by LC-ICP-MS after desalting via ultrafiltration. Separations were performed by size-exclusion chromatography. The corresponding chromatographs are shown in figure 1.
Figure 1: SEC-ICP-MS chromatograms (m/z 202) for influenza vaccine, which was incubated with different concentrations of THI in a refrigerator for 20 d.
It can be seen that adduct formation increases with higher concentrations of THI and with the highest concentration used, adduct formation is still on the rise. Although binding positions on the protein seem not to be saturated, most of the EtHg+ occurs freely in solution. Mercury binds on a protein, that is larger than 133 kDa, indicating adduct formation with HA. External calibration using THI was possible using meso-2,3-Dimercaptosuccinic acid (DMSA) as complexing agent and a mercury concentration of 141 μg Hg L−1 (SD: 6 μg Hg L−1) was found to bind to the high-molecular fraction of the vaccine. These results clearly show that thiomersal is not only a preservative but reacts with the proteins of the vaccine forming adducts.
The original study:
Used techniques and instrumentation:
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last time modified: September 22, 2024
