Nguyen Cuc Thi

The aim of this work was to use the gel electrophoresis technique coupled with a laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP MS) to study selenoproteins which may be present in plants samples. Sodium dodecyl sulfate polyacrylamide gel electrophoresis separation technique (SDS-PAGE) was employed to separate proteins. LA-ICP MS was applied for screening of Se incooperated into proteins. The LA conditions were optimized to obtain the highest sensitivity. Three different kinds of material (none-treated gel, lyophilized gel, and membranes) were also tested in LA-ICP MS analysis. Chives plant (Allium schoenoprasum) was grown in soil enriched with selenite and selenate. Different extraction methods were applied to obtain different soluble proteins fractions. The extraction solvents were as follows: water, NaCl 0.5M, NaOH 0.1M, Tris-buffer 0.5mM (pH=7.5). The sequential extraction was also performed in the order of water-salt-alkaline extraction. The total Se and protein content in each extract was determined. As expected, chives enriched with Se(VI) have accumulated a magnitude higher amount of selenium than chives fortified with Se (IV) in both parts (leaves, roots). In the sequential extraction, most Se was released in water fraction, but the obtained amount of protein was highest in alkaline fraction. For the separated extractions, the total Se content was similar in cases of water, NaOH, and Tris-buffer extract. However, a significant greater amount of protein was observed in the case of alkaline extraction. Unfortunately, SDS-PAGE was failed in separation proteins in NaOH extracts. For other extracts, LA-ICP MS was carried out. The obtained electropherograms showed a failure of  the Se-detection in proteins. For better understanding Se distribution in chives samples, chromatography separation (HPLC) coupled with ICP MS was also applied. The results were untypical for Allium plants: Se(VI) was the dominating form of Se(IV)-enriched samples extracts, and in Se(VI)-enrichments cases, besides Se(VI), SeMetSeCys was also the major form.