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Molecular Biomethods Handbook pp 479–489Cite as

Reversed-Phase HPLC

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Abstract

Reversed-phase HPLC (RP-HPLC) is now well establrshed as a technique for rsolanon, analysts, and structural elucrdatron of peptrdes and proteins (1,2) Its use m protem rsolatron and purificatron may have reached a peak owing to recent developments m high efficiency ron-exchange and hydrophobic-mteraction supports, which are now capable of equivalent levels of resolution to RP-HPLC without concomrtant risk of denaturatron and loss of brologrcal actrvtty. Nevertheless, there are many appllcatrons in which denaturatron may be unimportant and high concentrattons of organic modifier can be tolerated (e.g., purity analyses, structural studies, and micropreparative purification prior to mrcrosequencing) It is a widely used tool m biotechnology for process momtoring, purity studies, and stability determmatrons

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References

  1. Hancock, W S and Harding, D R K (1984) Review of separation condltlons, in CRC Handbook of HPLC for the Separation of Ammo Acids, Peptldes and Proteins, vol 2 (Hancock, W S, ed), CRC, Boca Raton, FL, pp 303–312

    Google Scholar 

  2. Regmer, F E (1987) Peptlde mapping LUGC 5(5), 392–395

    Google Scholar 

  3. Dill, K A (1980) The mechanism of solute retention in reversed-phase liquid chromatography J Phys Chem 91, 1987–1992

    Google Scholar 

  4. Melander, W and Horvath, C S (1980) Reversed-phase chromatography, in HPLC—Advances and Perspectwes, vol 2 (Horvath, C S, ed), Academic, New York, p 114

    Google Scholar 

  5. Hearn, M T W (1980) HPLC of peptldes, in HPLC—Advances and Perspectives, vol 3 (Horvath, C S, ed), Academic, New York, p 99

    Google Scholar 

  6. Guo, D, Mant, C T, Parker, J M R, and Hodges, R S (1986) PredIctIon of peptlde retention times in reversed-phase HPLC I Determmatlon of retention coefficients of ammo acid residues of model synthetic peptldes J Chromatogr 359,499–517

    Article  CAS  Google Scholar 

  7. Meek, J L and Rossettl, Z L (1981) Factors affecting retention and resolution of peptides in HPLC J Chromatogr 211, 15–28

    Article  CAS  Google Scholar 

  8. Hemltz, M L, Flaingan, E, Orlowskl, R C, and Regnier, F E, (1988) Correlation of calcltonm structure with chromatographlc retention in HPLC J Chromutogr 443,229–245

    Article  Google Scholar 

  9. O’Hare, M J, Capp M W, Nice, E C, Cooke, N H, and Archer, B G (1982) Factors mfluencing chromatography of protems in short alkylsllane-bonded large pore-size slhcas Anal Biochem 126, 17–128

    Article  PubMed  Google Scholar 

  10. Pearson, J D, Lm, N T, and Regmer, F E (1982) Separation of proteins, in HPLC of Peptldes and Proteins (Heam, M T W, Wehr, C T, and Regmer, F E, eds), Academlc, New York, p 81

    Google Scholar 

  11. Hearn, M T W (1986) Reversed-phase chromatography, in HPLC—Advances and Perspectives, vol 3 (Horvath, C S, ed), Academic, New York, pp 87–91

    Google Scholar 

  12. Burton, W G, Nugent, K D, Slattery, T K, and Summers, B R (1988) Separation of proteins by reversed-phase HPLC I Optlmlsing the column J Chromatogr 443,363–379

    Article  PubMed  CAS  Google Scholar 

  13. Nugent, K D, Burton, W G, Slattery, T K, and Johnson, B F (1988) Separation of proteins by reversed-phase HPLC II Optlmlsing sample pre-treatment and mobile phase condltlons J Chromatogr 443,381–397

    Article  PubMed  CAS  Google Scholar 

  14. GlaJch, J L, Kirkland, J J, and Kohler, J (1987) Effect of column degradation on the reversed-phase HPLC of peptldes and proteins J Chromatogr 384, 81–90

    Article  PubMed  CAS  Google Scholar 

  15. Sagllano, J, Floyd, T R, HartwIck, R A, Dlbussolo, J M, and Miller, N T (1988) Studies on the stablllsatlon of reversed-phases for liquid chromatography J Chromatogr 443,155–172

    Article  Google Scholar 

  16. De Vos, F L, Robertson, D M, and Hearn, MT W (1987) Effect of mass loadablllty, protein concentration and n-alkyl cham length on the reversed-phase HPLC behavtour of bovine serum albumm and bovme follicular fluid mhlbm J Chromatogr 392, 17–32

    Article  PubMed  Google Scholar 

  17. Guo, D, Mant, C T, and Hodges, R S (1987) Effects of ion-pamng reagents on the predlctlon of peptlde retention in reversed phase HPLC J Chromatogr 386,205

    Article  PubMed  CAS  Google Scholar 

  18. Geng, X and Regmer, F E (1984) Retention model for proteins in reversed-phase liquid chromatography J Chromatogr 296, 15–30

    Article  PubMed  CAS  Google Scholar 

  19. Kurntam, M, Johnson, D, and Snyder, L R (1986) Model of protein conformation in the reversed-phase separation of mterleukm-2 mutems J Chromatogr 371,313–333

    Article  Google Scholar 

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Author information

Authors and Affiliations

  1. SmzthKlzne Beecham, The Frythe, Welwyn, UK

    Bill Neville

Authors
  1. Bill Neville
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Editor information

Editors and Affiliations

  1. University of Herfordshire, Hatfield, UK

    Ralph Rapley  & John M. Walker  & 

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© 1998 Humana Press Inc , Totowa, NJ.

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Neville, B. (1998). Reversed-Phase HPLC. In: Rapley, R., Walker, J.M. (eds) Molecular Biomethods Handbook. Springer Protocols Handbooks. Humana Press. https://doi.org/10.1007/978-1-59259-642-3_39

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  • DOI: https://doi.org/10.1007/978-1-59259-642-3_39

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-501-0

  • Online ISBN: 978-1-59259-642-3

  • eBook Packages: Springer Book Archive

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