Paired-ion reversed-phase high-performance liquid chromatography of human and rat calcitonin

P. W. Lambert, B. A. Roos

Research output: Chapter in Book/Report/Conference proceedingChapter

7 Citations (Scopus)

Abstract

Improved methods for isolation, characterization, and quantitation of immunochemically heterogeneous forms of calcitonin (CT) in tissue and plasma must be developed before the biological origins and clinical importance of CT moieties can be elucidated. We are now proposing reversed-phase high-performance liquid chromatography (RP-HPLC) as one possible means of achieving high recovery, high resolution of CT moieties. In this paper we report RP-HPLC analyses of trace amounts of radiolabeled and unlabeled synthetic human and rat CT. we have systematically evaluated our application of RP-HPLC by employing several elution modes, including isocratic and gradient elution, as well as several elution reagents. We determined that high recovery and high resolution were best achieved with alkyl ion-pairing reagents, such as tetrabutylammonium phosphate, pH 7.5, or sodium sulfonyl hexane, pH 3.5. The most sensitive UV detection of trace amounts of CT was achieved with tetrabutylammonium phosphate buffer (TBAP). We recommend for RP-HPLC of CT a C18-bonded silica column and elution with a 20-min linear gradient of methanol-water (20:80 to 80:20, v/v) containing 0.005 M TBAP. Combined with appropriate extraction procedures, such as silica adsorption or immuno-adsorbant chromatography, this paired-ion RP-HPLC method can be an important aid in achieving more accurate and extensive information about CT moieties in biological samples. This method will also allow the rapid, optical detection and quantitation of CT moieties recovered from tissues, and perhaps from plasmas.

Original languageEnglish
Title of host publicationJournal of Chromatography
Pages293-299
Number of pages7
Volume198
Edition3
StatePublished - Dec 1 1980
Externally publishedYes

Fingerprint

Calcitonin
High performance liquid chromatography
Reverse-Phase Chromatography
Rats
High Pressure Liquid Chromatography
Ions
Phosphates
Silicon Dioxide
Buffers
Tissue
Plasmas
Recovery
Hexanes
Chromatography
Adsorption
Methanol
Sodium
Water

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Molecular Medicine

Cite this

Lambert, P. W., & Roos, B. A. (1980). Paired-ion reversed-phase high-performance liquid chromatography of human and rat calcitonin. In Journal of Chromatography (3 ed., Vol. 198, pp. 293-299)

Paired-ion reversed-phase high-performance liquid chromatography of human and rat calcitonin. / Lambert, P. W.; Roos, B. A.

Journal of Chromatography. Vol. 198 3. ed. 1980. p. 293-299.

Research output: Chapter in Book/Report/Conference proceedingChapter

Lambert, PW & Roos, BA 1980, Paired-ion reversed-phase high-performance liquid chromatography of human and rat calcitonin. in Journal of Chromatography. 3 edn, vol. 198, pp. 293-299.
Lambert PW, Roos BA. Paired-ion reversed-phase high-performance liquid chromatography of human and rat calcitonin. In Journal of Chromatography. 3 ed. Vol. 198. 1980. p. 293-299
Lambert, P. W. ; Roos, B. A. / Paired-ion reversed-phase high-performance liquid chromatography of human and rat calcitonin. Journal of Chromatography. Vol. 198 3. ed. 1980. pp. 293-299
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