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KMID : 0366919970090010046
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Sungkyun Pharmceutical Journal
1997 Volume.9 No. 1 p.46 ~ p.52
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Reversed-phase high-performance liquid chromatography of radioiodinated salmon calcitonins
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Lee, Kang Choon
Kang, Tae Suk/Woo, Byung Ho/Lee, Jung Tae/Lee, Hye Suk/DeLuca, Patrick P.
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Abstract
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Reversed-phase HPLC conditions for simultaneous separation of salmon calcitonin, mono- and di-radioiodinated salmon calcitonins and their tryptic digested fragments have been developed. Salmon calcitonin was radioiodinated with NA^125 I by the iodo-beads method. After solid-phase exraction from the reaction mixtures using C_18 Bond Elut cartridges, mono- and di-radioiodinated salmon calcitonins were separated from each other, as well as from unlabeled salmon calcitonin, on a Bondclone 10 C_18 column (300×7.8 mn I.D.) by isocratic elution with 0.1% trifluoroacetic acid in 34% aqueous acetonitrile. The characteristics of eithe iodinated peptides or unlabeled salmon calcitonin were evaluated on the basis of UV absorbance (215 and 280 nm), fluorescence (λ_ex = 282 nm, λ_em = 310 nm) and measurement of specific radioactivity by means of a flow-through radio-isotope detector. HPLC separation of a tryptic digest of iodinated salmon calcitonin fraction on a W-porex 5C_18 300Å column (250×4.6 mm I.D.) and subsequent amino acid analysis, led to the conclusion that radioiodination took place at the Tyr residue and not at the His moiety.
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