Using L-cysteine to enhance calibration range and prevent a memory effect in mercury analysis of complex samples via ICP-OES

Background

Mercury (Hg) is a persistent pollutant occurring in the environment able to transition between different species. It can therefore be found in air, soil and water reservoirs becoming a present concern for the general population but also sensitive populations like pregnant women. Therefore, investigating organ-specific transfer mechanisms of Hg is mandatory for Hg toxicity testing. For this, an in vitro system using microporous inserts to monitor the transfer across an in vitro placental barrier has been used. However, due to the cytotoxicity of Hg only low concentrations (1.26 ×10⁻⁴ – 1.36 ×10⁻² µg/µL Hg) can be applied, making Hg determination in cell culture medium using inductively coupled plasma-optical emission spectrometry challenging, especially when these trace amounts should be determined alongside other trace elements which are naturally occurring in cells and cell culture medium like the essential metals manganese (Mn), iron (Fe), copper (Cu), and zinc (Zn). Additionally, Hg analysis on an ICP system holds also a number of challenges like a persistent memory effect and instability of Hg standard solutions.

Methods

The development of a rapid and sensitive ICP-OES method to determine Hg in different matrices like cell culture medium and cells has been performed on an Avio 220 Max ICP-OES (Perkin-Elmer) equipped with a cyclonic spray chamber and MicroMist® nebulizer. Cell lysates and cell culture medium were diluted in a mixture of 0.2 % L-cysteine, 2 % HNO₃ and 0.1 % HCl and directly introduced into the ICP-OES system. Further method development included the suitability of the analysis of multiple elements like Mn, Fe, Cu, and Zn as well as the determination of the limit of detection and limit of quantification.

Results

The combination of 0.2 % L-cysteine, 2 % HNO₃ and 0.1 % HCl is able to bind and stabilize Hg ions in standard solutions and in biological matrices over a wide dynamic concentration range (1 – 500 µg/L) also alongside other metals like Mn, Fe, Cu and Zn without losses of sensitivity. A short run time of 3 min enables high throughput analysis. Additionally, the high salt and carbon concentrations in the culture medium do not affect Hg sensitivity using the ICP-OES.

Conclusion

This method is a useful tool for the quantification of Hg in a variety of complex matrices including cells and cell culture media (high salt and carbon-rich (∼1 % each)) with high sensitivity and minimal sample preparation allowing high throughput. Furthermore, not only Hg can be determined in biological matrices, but even multiple elemental analysis can be carried out to address the effect of Hg on other metals homeostasis.