• Bulletin of the Korean Chemical Society
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• v.34 no.12
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• pp.3817-3824
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• 2013

Various separation modes in HPLC, such as anion exchange (AE), reversed-phase (RP), and affinity (AF) chromatography were examined for the separation of selenium species in human blood serum and urine. While RP- and AE-HPLC were mainly used for the separation of small molecular selenium species, double column AF-HPLC achieved the separation of selenoproteins in blood serum efficiently. Further, the effluent of AF-HPLC was enzymatically hydrolyzed and then analyzed with RP HPLC for selenoamino acid study. The versatility of the hybrid technique makes the in-depth study of selenium species possible. For quantification, post column isotope dilution (ID) with $^{78}Se$ spike was performed. ORC ICP/MS (octapole reaction cell inductively coupled plasma/mass spectrometry) was used with 4 mL $min^{-1}$ Hydrogen as reaction gas. In urine sample, inorganic selenium and SeCys were identified. In blood serum, selenoproteins GPx, SelP and SeAlb were detected and quantified. The concentration for GPx, SelP and SeAlb was $22.8{\pm}3.4\;ng\;g^{-1}$, $45.2{\pm}1.7\;ng\;g^{-1}$, and $16.1{\pm}2.2\;ng\;g^{-1}$, respectively when $^{80}Se/^{78}Se$ was used. The sum of these selenoproteins ($84.1{\pm}4.4\;ng\;g^{-1}$) agrees well with the total selenium concentration measured with the ID method of $87.0{\pm}3.0\;ng\;g^{-1}$. Enzymatic hydrolysis of each selenium proteins revealed that SeCys is the major amino acid for all three proteins and SeMet is contained in SeAlb only.

• Analytical Science and Technology
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• v.27 no.2
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• pp.92-99
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• 2014

Interferences were removed using anion exchange solid phase extraction (AE SPE) in quantification of selenoproteins in Korean human blood serum with affinity high performance liquid chromatography (AF HPLC)-inductively coupled plasma/mass spectrometry (ICP/MS). The average selenium level obtained for healthy Koreans was $94.3{\pm}2.3ngg^{-1}$ using isotope dilution method. AE SPE was coupled to AF column to separate 3 selenoproteins, glutathione peroxidase GPx, selenoprotein SelP, and selenoalbumin SeAlb. Post column isotope dilution was employed to quantify the proteins. The certified reference material of human blood serum BCR-637 was analyzed to give total selenoprotein concentration of $85.4{\pm}3.4ngg^{-1}$, which agreed well with the reference value of $81{\pm}7ngg^{-1}$. The pooled concentration of GPx, SelP, and SeAlb from healthy Koreans (n=20) was $12.1{\pm}1.4ngg^{-1}$, $57.2{\pm}2.0ngg^{-1}$, and $20.0{\pm}1.9ngg^{-1}$, respectively. The sum of selenoproteins is $89.3ngg^{-1}$, which is about the same as the total selenium concentration of $94.3ngg^{-1}$. The fact suggests that selenium in blood serum is mostly consisted of selenoproteins. After the removal of interference, GPx showed a significant decrease (more than 50%) from $25.0ngg^{-1}$ to $12.1ngg^{-1}$. It was identified that the interference in blood serum was mostly from GPx and the use of AE SPE was proven to be efficient in eliminating Cl and Br that cause interference to GPx.

• Analytical Science and Technology
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• v.28 no.3
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• pp.196-203
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• 2015

Selenium species (inorganic selenium, selenoaminoacids, and selenoproteins) were analyzed using anion exchange and affinity chromatography, which were connected to ICP/MS for the blood serum of sows fed by seleniumfortified feed. The Anion Exchange PRP X-100 column was used for the analysis of inorganic selenium (Se⁴⁺ and Se⁶⁺) and selenoaminoacids. The HEP column was used to separate SelP from GPx+SeAlb in selenoproteins. A quantitative analysis was performed using the post-column isotope dilution technique. The lactating sows were divided into three groups and fed by selenium fortified feed (organic 0.3 mg/kg, 0.6 mg/kg and inorganic 0.6 mg/kg) for four weeks. The test groups showed increases in selenoaminoacids compared with the control group, except the inorganic feed group. There was no significant difference between the organic feed groups. All test groups showed increases in selenoproteins. In particular, SelP showed a large increase that was 1.5 times higher than the other proteins.