Skip to main content
SpringerLink
Log in

High-Performance Liquid Chromatography for Hormone Assay

  • Protocol
Hormone Assays in Biological Fluids

Part of the book series: Methods in Molecular Biology ((MIMB,volume 324))

  • 1687 Accesses

Abstract

High-performance liquid chromatography (HPLC) is a refinement of traditional column chromatographic techniques. The speed of analysis and the resolution are increased with new column-packing materials and eluant pumped through the column at high pressure. The potential for achieving measurements of hormones in small volumes of plasma or urine is limited, both in normal and pathological situations. Using HPLC with ultraviolet absorption, the detection limit is only nanogram amounts of hormones per milliliter of blood serum. The applications of the technique to specific hormones from recent and older literature will be used throughout this chapter to illustrate aspects of the technology.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 109.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Schnedl, W. J., Krause, R., Halwachs-Baumann, G, Trinker, M. Lipp, R. W. and Krejs, G. J. (2000) Evaluation of HbA1c determination methods in patients with hemoglobinopathies. Diabetes Care 23, 339–344.

    Article  PubMed  CAS  Google Scholar 

  2. Pappa, A., Seferiades, K., Fotsis, T., et al. (1999) Purification of a candidate gonadotrophin surge attentuating factor from human follicular fluid. Hum. Reprod. 14, 1449–1456.

    Article  PubMed  CAS  Google Scholar 

  3. Slominski, A., Szczesniewski, A., and Wortsman, J. (2000) Liquid chromatography-mass spectrometry detection of corticotrophin-releasing hormone and pro-opiomelanocortin-derived peptides in human skin. J. Clin. Endocrinol. Metab. 85, 3252–3258.

    Google Scholar 

  4. Slominski, A., Heasley, D., Mazurkiewicz, J. E., Ermak, G., Baker, J., and Carlson, J. A. (1999) Expression of proopiomelanocortin (POMC)-derived melanocyte-stimulating hormone (MSH)= and adrenocorticotropic hormone (ACTH) peptides in skin of basal cell carcinoma patients. Hum. Pathol. 30, 208–215.

    Article  PubMed  CAS  Google Scholar 

  5. Gardi, J., Khalil, W. K., Vecsernyes, M. Gaspar, L., Szecsi, M., and Julesz, J. (2000) Molecular forms of alpha-melanocyte-stimulating hormone in phaeochromocytoma tissue. Endocr. Res. 26, 71–79.

    Article  PubMed  CAS  Google Scholar 

  6. O’Connor, J. V. (1999) Chromatography of recombinant proteins. Dev. Biol. Stand. 97, 39–47.

    PubMed  Google Scholar 

  7. Lu, R., Kopeckova, P., and Kopecek, J. (1999) Degradation and aggregation of human calcitonin in vitro. Pharm. Res. 16, 359–367.

    Article  CAS  Google Scholar 

  8. Gonzalo-Lumbreras, R. and Izquierdo-Hornillos, R. (2000) Optimization of the high performance liquid chromatography separation of a complex mixture containing urinary steroids, boldenone and bolasterone: application to urinary samples. J. Chromatogr. B. Biomed. Sci. 26, 47–57.

    Article  Google Scholar 

  9. Linde, S., Welinder, B. S., and Nielsen, J. H. (1993) Analysis of proinsulin and its conversion products by reverse phase high performance liquid chromatography. J. Chromatogr. 614, 185–204.

    Article  PubMed  CAS  Google Scholar 

  10. Ohkubo, T. (1994) High performance liquid chromatographic analysis of peptide hormones in transplanted rat islets. Biomed. Chromatogr. 8, 301–305.

    Article  PubMed  CAS  Google Scholar 

  11. Oliva, A., Farina, J., and Llabres, M. (2000) Development of two high performance liquid chromatographic methods for the analysis and characterisation of insulin= and its degradation products in pharmaceutical preparations. J. Chromatogr..B Biomed. Sci. 749, 25–34.

    Article  CAS  Google Scholar 

  12. Nakamura, K., Nashimoto, M., Hori, Y., and Yamamoto, M. (2000) Serum 25-hydroxyvitamin D concentrations and related dietary factors in peri-and postmenopausal Japanese women. Am. J. Clin. Nutr. 71, 1161–1165.

    PubMed  CAS  Google Scholar 

  13. Kim, J. H. and Moon, S. J. (2000) Time spent outdoors and seasonal variation in serum concentrations of 25-hydroxyvitamin D in Korean women. Int. J. Food Sci. 51, 439–451.

    Article  CAS  Google Scholar 

  14. Bi, M. and Singh, J. (2000) Effect of buffer, pH, buffer concentration and skin with or without enzyme inhibitors on the stability of [Arg(8)]-vasopressin. Int. J. Pharm. 197, 87–93.

    Article  PubMed  CAS  Google Scholar 

  15. Ando, T., Koshika, S., Komura, K., Nakayama, Y., and Hara, S. (1986) Characterisation of packing columns for the liquid chromatographic determination of corticosteroids in human plasma. J. Liquid. Chrom. 9, 2601–2608.

    Article  CAS  Google Scholar 

  16. Fantl, V., Lim, C. K., and Gray, C. H. (1976) in High Pressure Liquid Chromatography in Clinical Chemistry (Dixon, P.F. Gray, C.H. Lim, C.K., and Stoll. M..S. eds.) Academic Press, London.

    Google Scholar 

  17. Musey, P. I., Collins, D. C., and Preedy, J. (1979) Separation of oestrogen conjugates by high performance liquid chromatography. Steroids 31, 583–592.

    Article  Google Scholar 

  18. Adamcztk, M., Gebler, J. C., and Wu, J. (2000) Sequencing of antithyroxine monoclonal antibody fab fragment by ion trap mass spectrometry. Rapid Commun. Mass Spectrom. 14, 999–1007.

    Article  Google Scholar 

  19. O’Hare, M. J., Nice, E. C., Magee-Brown, R., and Bullman, H. (1976) High performance liquid chromatography of steroids secreted by human adrenal and testis cells in monolayer culture J. Chromatogr. 125, 357–367.

    Article  PubMed  Google Scholar 

  20. Schoneshofer, M. and Dulce, H. J. (1979) Comparison of different high performance liquid chromatographic systems for the purification of adrenal and gonadal steroids prior to immunoassay. J. Chromatogr. 164, 17–28.

    Article  PubMed  CAS  Google Scholar 

  21. O’Hare, M. J., Nice, E. C., and Capp, M. (1980) Reversed and normal phase high performance liquid chromatography of 18-hydroxylated steroids and their derivatives. Comparison of selectivity, efficiency and recovery from biological samples. J. Chromatogr. 198, 23–29.

    Article  PubMed  Google Scholar 

  22. O’Hare, M. and Nice, E. C. (1981) Analysis of steroid hormones in adrenal and testicular cells and tissues, in Steroid Analysis by HPLC: Recent Applications (Kautsky, K. P., ed.) Marcel Dekker Inc., NY, pp. 277–322.

    Google Scholar 

  23. Miller, C. and Rivier, J. (1996) Peptide chemistry: development of high performance liquid chromatography and capillary zone electrophoresis. Biopolymers 40, 265–317.

    Article  PubMed  CAS  Google Scholar 

  24. Majid, O., Akhlaghi, F., Lee, T., Holt, D. W., and Trull, A. (2001) Simultaneous determination of plasma prednisolone, prednisone and cortisol levels by high performance liquid chromatography. Ther. Drug Monitor 23, 163–168.

    Article  CAS  Google Scholar 

  25. Hara, S., Fujii, Y., Hirasawa, M., and Miyamoto, S. (1978) Systematic design of binary solvent systems for liquid solid chromatography via retention behaviour of mono-and di-functional steroids on silica gel columns. J. Chromatogr. 149, 680–703.

    Article  Google Scholar 

  26. Capp, M. W. and Simonian, M. H. (1985) Separation of the major adrenal steroids by reverse phase high performance liquid chromatography Anal. Biochem. 147, 374–381.

    Article  PubMed  CAS  Google Scholar 

  27. Wei, J-Q., Wei, J-L., Zhou, X-T., and Cheng, J-P. (1990) Isocratic reversed phase high performance liquid chromatography determination of twelve natural corticosteroids in serum with on-line ultraviolet and fluorescence detection. Biomed. Mass Spectrom. 4, 161–164.

    CAS  Google Scholar 

  28. Burgess, C. (1978) Rapid reverse phase high performance liquid chromatographic analysis of steroid products. J. Chromatogr. 149, 233–240.

    Article  PubMed  CAS  Google Scholar 

  29. Hara, S. and Hayashi, S. (1977) Correlation of retention behaviour of steroid pharmaceuticals in polar and bonded reverse phase liquid column chromatography J. Chromatogr. 142, 689–703.

    Article  PubMed  CAS  Google Scholar 

  30. D’Agostino, G., Castagnetta, L., Mitchell, F., and O’Hare, M. J. (1985) Computer aided mobile phase optimisation and chromatogram simulation in HPLC. A review. J. Chromatogr. 338, 1–23.

    Article  Google Scholar 

  31. Wei, J-Q., Wei, J-L., and Zhou, X-T. (1990) Optimisation of an isocratic reverse phase performance liquid chromatographic system for the separation of fourteen steroids using factorial design and computer simulation. Biomed. Chromatogr. 4, 34–38.

    Article  PubMed  CAS  Google Scholar 

  32. Kreutzmann, D. J. and Silink, M. (1987) Determination of 17-oxosteroid glucuronides and sulphates in urine by liquid chromatography using 2,4-dinitrophenylhydrazine as a prelabelling reagent for spectrophotometric detection. J. Chromatogr. 415, 253–260.

    Article  PubMed  CAS  Google Scholar 

  33. Derks, H. J. G. M. and Drayer, N. M. (1978) The identification and quantification of three new 6α-hydroxylated corticosteroids in human neonatal urine. Steroids 31, 289–305.

    Article  PubMed  CAS  Google Scholar 

  34. Dolan, J. W., Snyder, L. R., Djordjevic, N. M., et al. (1998) Simultaneous variation of temperature and gradient steepness for reversed phase high performance liquid chromatography method development. I Application to 14 different samples using computer simulation. J. Chromatogr. A. 803, 1–31.

    Article  PubMed  CAS  Google Scholar 

  35. Shimada, K., Tanaka, T., and Nambara, T. (1979) Studies on steroids CI Separation of catechol oestrogens by high performance liquid chromatography with electrochemical detection. J. Chromatogr. 178, 350–354.

    Article  PubMed  CAS  Google Scholar 

  36. Shimada, K., Xie, F., and Nambara, T. (1986) Studies on steroids CCXIX. Separation and determination of 4-hydroxyoestriol monoglucuronides and monosulphates in biological fluids by high performance liquid chromatography with electrochemical detection. J. Chromatogr. 378, 17–24.

    Article  PubMed  CAS  Google Scholar 

  37. Tscherne, R. J. and Capitano, G. (1977) High pressure liquid chromatographic separation of pharmaceutical compounds using a mobile phase containing silver nitrate. J. Chromatogr. 136, 337–341.

    Article  PubMed  CAS  Google Scholar 

  38. Andreolini, F., Borra, C., Caccamo, F., Di Corcia, A., and Samperi, R. (1987) Estrogen conjugates in late-pregnancy fluids-extraction and group separation by a graphitised carbon black cartridge and quantification by high performance liquid chromatography. Anal. Chem. 59, 1720–1725.

    Article  PubMed  CAS  Google Scholar 

  39. Shimada, K. and Nonaka, M. (1991) Utility of cyclodextrin in mobile phase for high performance liquid chromatography of C21 steroids. J. Liquid Chromatogr. 14, 2109–2117.

    Article  CAS  Google Scholar 

  40. Agnus, B., Sebille, B., and Gosselet, N. M. (1991) Effect of β-cyclodextrin in the mobile phase on the retention and indirect retention of non-electrolytes in reverse phase liquid chromatography. J. Chromatogr. 552, 583–592.

    Article  CAS  Google Scholar 

  41. Lamparczyx, H., Zarzycki, P. K., and Nowakowska, J. (1994) Application of beta-cyclodextrin for the analysis of estrogenic steroids in human urine by high performance liquid chromatography. Chromatographia 38, 168–172.

    Article  Google Scholar 

  42. Darby, S. M., Miller, M. L., Allen, R. O., and LeBeau, M. (2001) A mass spectrometric method for quantitation of intact insulin in blood samples. J. Anal. Toxic. 25, 8–14.

    CAS  Google Scholar 

  43. Grippa, E., Santini, L., Castellano, G., Gatto, M. T., Leone, M. G., and Saso, L. (2000) Simultaneous determination of hydrocortisone, dexamethasone, indomethacin, phenybutazone and oxyphenbutazone in equine serum by high performance liquid chromatography. J. Chromatogr. B Biomed. Sci. 738, 17–25.

    Article  CAS  Google Scholar 

  44. Naik, G. O., Moe, G. W., and Armstrong, P. W. (2001) Specific and non-specific measurements of angiotensin II cascade members. J. Pharm. Biomed. Anal. 24, 947–955.

    Article  PubMed  CAS  Google Scholar 

  45. Lin, W-J., Her, S-J., Chen, P-F., and Chen, R. R-L. (1998) Determination of medrogestone in plasma by high performance liquid chromatography. J. Chromatogr. B. 714, 263–268.

    Article  CAS  Google Scholar 

  46. Seki, T. and Yamaguchi, Y. (1984) New fluorometric detection method of corticosteroids after high performance liquid chromatography using post-column derivatization bezamidine. J. Chromatogr. 305, 188–193.

    Article  PubMed  CAS  Google Scholar 

  47. Satyaswaroop, P. G., de la Osa, E. L., and Gurpide, E. (1977) High pressure liquid chromatographic separation of C18 and C19 steroids. Steroids 30, 139–145.

    Article  PubMed  CAS  Google Scholar 

  48. Watanabe, K. and Yoshizawa, I. (1985) Clinical analysis of steroids XXXI. Assay of oestradiol-17-sulphate 4 hydroxylase activity by high performance liquid chromatography with electrochemical detection. J. Chromatogr. 337, 114–120.

    Article  PubMed  CAS  Google Scholar 

  49. Fernandez, N., Garcia, J. J., and Diez, M. J. (1993) Rapid high performance liquid chromatographic assay of ethinylestradiol in rabbit plasma. J. Chromatogr. Biomed. Appl. 619, 143–147.

    Article  CAS  Google Scholar 

  50. Sastro Torano, J., Rijn-Baker, P. V., Merkus, P., and Guchelaar, H. J. (2000) Quantitative determination of melatonin in human plasma and cerebrospinal fluid with high performance liquid chromatography and fluorescence detection. Biomed. Chromatogr. 14, 306–310.

    Article  Google Scholar 

  51. Harumi, T. and Matsushima, S. (2000) Separation and assay methods for melatonin= and its precursors. J. Chromatogr. B Biomed. Sci. 747, 95–110.

    Article  CAS  Google Scholar 

  52. Tan, D., Manchester, D. L. C., Reiter, R. J., Qi, W., Hanes, M. A., and Farley, N. J. (1999) High physiological levels of melatonin in bile of mammals. Life Sci. 65, 2523–2529.

    Article  PubMed  CAS  Google Scholar 

  53. Andrisano, V., Bertucci, C., Battaglia, A., and Cavrini, V. (2000) Photostability of drugs: photodegradation of melatonin and its determination in commercial formulations. J. Pharm. Biomed. Anal. 23, 15–23.

    Article  PubMed  CAS  Google Scholar 

  54. Bizanek, R., Manes, J. D., and Fujinari, E. M. (1996) Chemiluminesent nitrogen detection as a new technique for purity assessment of synthetic peptides separated by reversed phase HPLC. Pept. Res. 9, 40–44.

    PubMed  CAS  Google Scholar 

  55. Lam, S., Malikin, G., and Karmen, A. (1988) High performance liquid chromatography of hydroxysteroids detected with post-column immobilised enyme reactors. J. Chromatogr. 441, 81–87.

    Article  PubMed  CAS  Google Scholar 

  56. Hamase, K., Tomita, T., Kiyomizu, A., and Zaitsu, K. (2000) Determination of pineal melatonin by precolumn derivatization reverse phase high performance liquid chromatography and its application to the study of circadian rhythm in rats and mice. Anal. Biochem. 279, 106–110.

    Article  PubMed  CAS  Google Scholar 

  57. Kessler, M. J. (1983) Quantitation of radiolabelled biological molecules by high performance liquid chromatography. J. Chromatogr. 225, 209–217.

    Article  Google Scholar 

  58. Lundmo, P. and Sunde, E. (1984) Rapid analysis of C19 steroid metabolism by high performance liquid chromatography and inline monitoring of radioactivity. J. Chromatogr. 308, 289.

    Article  PubMed  CAS  Google Scholar 

  59. Niiyama, S., Happle, R., and Hofmann, R. (2001) Influence of estrogens on the androgen metabolism in different subunits of human hair follicles. Eur. J. Dermatol. 11, 195–198.

    PubMed  CAS  Google Scholar 

  60. Lonning, P. E., Geisler, J., Johannessen, D. C., et al. (2001) Pharmacokinetics and metabolism of formestane in breast cancer patients. J. Steroid Biochem. Mol. Biol. 77, 39–47.

    Article  PubMed  CAS  Google Scholar 

  61. Asaba, H., Hosoya, K., Takanaga, H., et al. (2000) Blood brain barrier is involved in the efflux transport of a neuroactive steroid, dehydroepiandrosterone sulphate, via organic anion transporting polypeptide 2. J. Neurochem. 75, 1907–1916.

    Article  PubMed  CAS  Google Scholar 

  62. Gentile, D. M., Tomlinson, E. S., Maggsm J. L., Park, K., and Back, D. J. (1996) Dexamethasone metabolism by human liver in vitro. Metabolite identification and inhibition of 6-hydroxylation. J. Pharm. Exper. Ther. 277, 105–112.

    CAS  Google Scholar 

  63. Ohno, M., Yamaguchi, I., Saiki, K., Yamamoto, I., and Azuma, J. (2000) Specific determination of urinary 6-beta-hydroxycortisol and cortisol by liquid chromatography-atmospheric pressure chemical ionisation mass spectrometry. J. Chromatogr. B Biomed. Sci. Appl. 746, 95–101.

    Article  PubMed  CAS  Google Scholar 

  64. Tang, C., Kassahun, K., McIntosh, I. S., Brunner, J., and Rodrigues, A. D. (2000) Simultaneous determination of urinary free cortisol and 6-beta-hydroxycortisol by liquid chromatography atmospheric pressure chemical ionisation tandem mass sepctrometry and its application for estimating hepatic CYP3A induction. J. Chromatogr. B Biomed. Sci. Appl. 742, 303–313.

    Article  PubMed  CAS  Google Scholar 

  65. Mitamura, K., Yatera, M., and Shimada, K. (2000) Studies on neurosteroids XII. Determination of enzymatically formed catechol estrogens and guaiacol estrogens by rat brains using liquid chromatography mass spectrometry mass spectrometry. J. Chromatogr. B Biomed. Sci. Appl. 748, 89–96.

    Article  PubMed  CAS  Google Scholar 

  66. Jia, Q., Hong, M. F., Pan, Z. X., and Orndorff, S. (2001) Quantitation of urine 17-ketosteroid sulfates and glucuronides by high performance liquid chromatography ion trap mass spectrometry. J. Chromatogr. B Biomed. Sci. Appl. 750, 81–91.

    Article  PubMed  CAS  Google Scholar 

  67. Wu, Z., Zhang, C., Yang, C., Zhang, X., and Wu, E. (2000) Simultaneous quantitative determination of norgestrel and progesterone in human serum by high performance liquid chromatography tandem mass spectrometry with atmospheric pressure chemical ionisation. Analyst 125, 2201–2205.

    Article  PubMed  CAS  Google Scholar 

  68. Nassar, A. E., Varshney, N., Getek, T., and Cheng, L. (2001) Quantitative analysis of hydrocortisone in human urine using a high performance liquid chromatographic tandem mass spectrometric atmospheric pressure chemical ionization method. J. Chromatogr. Sci. 39, 59–64.

    PubMed  CAS  Google Scholar 

  69. Wudy, S. A., Hartmann, M., and Svoboda, M. (2000) Determination of 17-hydroxyprogesterone in plasma by stable isotope dilution/benchtop liquid chromatography tandem mass spectrometry. Horm. Res. 53, 68–71.

    Article  PubMed  CAS  Google Scholar 

  70. Desiderio, D. M. (1996) Mass spectrometry, high performance liquid chromatography and brain peptides. Biopolymers 40, 257–264.

    Article  PubMed  CAS  Google Scholar 

  71. van den Burg, E. H., Metz, J. R., Arends, R. J., et al. (2001) Identification of beta-endorphins in the pituitary gland and blood plasma of the common carp (Cyprinus carpio). J. Endocrinol. 169, 271–280.

    Article  Google Scholar 

  72. Qian, M. G. and Lubman, D. M. (1996) Procedures for tandem mass spectrometry on an ion trap storage/reflectron time of flight mass spectrometer. Rapid Commun. Mass Spectrom. 10, 1911–1920.

    Article  PubMed  CAS  Google Scholar 

  73. Naylor, S., Johnson, K. L., Willaimson, B. L., Klarskov, K., and Gleich, G. J. (1999) Structural characterisation of contaminants in commercial preparations of melatonin by on-line HPLC electrospray ionization tandem mass spectrometry. Adv. Exp. Biol. Med. 467, 769–777.

    CAS  Google Scholar 

  74. Jonscher, K., Currie, G., McCormack, A. L., and Yates, J. R. (1993) Matrix assisted laser desorption of peptides and proteins on a quadrupole ion trap mass spectrometer. Rapid Commun. Mass Spectrom. 7, 20–26.

    Article  PubMed  CAS  Google Scholar 

  75. Hensel, R. R., King, R. C., and Owens, K. G. (1997) Electrospray sample preparation for improved quantitation in matrix assisted laser desorption/ioniation mass spectrometry. Rapid Commun. Mass Spectrom. 11, 1785–1793

    Article  PubMed  CAS  Google Scholar 

  76. Fredline, V. F., Kovacs, E. M., Taylor, P. J., and Johnson, A. G. (1999) Measurement of plasma renin activity with use of HPLC electrospray tandem mass spectrometry. Clin. Chem. 45, 659–664.

    PubMed  CAS  Google Scholar 

  77. DeBrandendere, V. I., Hou, P., Stockl, D., Thienpont, L. M., and De Leenheer, A. P. (1998) Isotope dilution liquid chromatography/electrospray ionization tandem mass spectrometry for the determination of thyroxine as a potential reference method. Rapid Commun. Mass Spectrom. 12, 1099–1103.

    Article  Google Scholar 

  78. Fell, A. F., Scott, H. P., Gill, R., and Moffat, A. C. (1983) Novel techniques for peak recognition and deconvolution by computeraided photodiode array detection in high performance liquid chromatography. J. Chromatogr. 282, 123–140.

    Article  CAS  Google Scholar 

  79. Mawer, E. B., Jones, G., Davies, M., et al. (1998) Unique 24-hydroxylated metabolites represent a significant pathway of metabolism of vitamin D2 in humans: 24-hydroxyvitaminDs and 1,24-dihydroyvitamin D2 detectable in human serum. J. Clin. Endocrinol. Metab. 83, 256–266.

    Article  Google Scholar 

  80. Homma, M., Beckerman, K., Hayashi, S., et al. (2000) Liquid chromatographic determination of urinary 6 beta-hydroxycortisol to assess cytochrome P-450 3A activity in HIV positive pregnant women. J. Pharm. Biomed. Anal. 23, 629–635.

    Article  PubMed  CAS  Google Scholar 

  81. Barbaccia, M. L., Lello, S., Sidiropoulo, T., et al. (2000) Plasma 5-alpha-androstane-3-alpha, 17-beta diol, an endogenous steroid that positively modulates GABA (A) receptor function and anxiety: a study in menopausal women. Psychoneuroendocrinology 25, 659–675.

    Article  PubMed  CAS  Google Scholar 

  82. Hampl, R., Hill, M., Sterzi, I., and Star, L. (2000) Immunomodulatory 7-hydroxylated metabolites of dehydroepiandrosterone are present in human semen. J. Steroid Biochem. Mol. Biol. 75, 273–276.

    Article  PubMed  CAS  Google Scholar 

  83. Schonesshofer, M., Fenner, A., and Dulce, H. J. (1981) Assessment of eleven adrenal steroids from a single sample by combination with automated high performance liquid chromatograpy and radioimmunoassay. J. Steroid Biochem. 14, 377–386.

    Article  Google Scholar 

  84. Rasmussen, T. N., Bersani, M., Johnsen, A. H., Kofod, H., and Holst, J. J. (1994) Pigs produce only a single form of CGRP part of which is processed to N-and C-terminal fragments. Peptides 15, 89–94.

    Article  PubMed  CAS  Google Scholar 

  85. Tagliaro, F., Camilot, M., Valentini, R., Mengarda, F., Antoniazzi, F., and Tato, L. (1998) Determination of thyroxine in the hair of neonates by radioimmunoassay with high performance liquid chromatographic confirmation. J. Chromatogr. B Biomed. Anal. 716, 77–82.

    Article  CAS  Google Scholar 

  86. Zhao, X., White, R., Huang, B. S., Van Huysse, J., and Leenen, F. H. (2001) High salt intake and the brain renin-angiotensin system in Dahl salt-sensitive rats. J. Hypertens. 19, 89–98.

    Article  PubMed  CAS  Google Scholar 

  87. Deacon, C. F., Nauck, M. A., Meier, J., Hucking, K., and Holst, J. J. (2000). Degradation of endogenous and exogenous gastric inhibitory polypeptide in healthy and in type 2 diabetic subjects as revealed using a new assay for intact peptide. J. Clin. Endocrinol. Metab. 85, 3575–3581.

    Article  PubMed  CAS  Google Scholar 

  88. Shackleton, C. H. L. and Whitney, J. O. (1980) Use of Sep Pak cartridges for urinary steroid extractions: evaluation of the method for use prior to gas chromatographic analysis. Clin. Chim. Acta 107, 231–243.

    Article  PubMed  CAS  Google Scholar 

  89. Morris, D. J. and Tsai, R. (1981) Chromatographic separation of aldosterone and its metabolites. Adv. Chromatogr. 19, 261–285.

    PubMed  CAS  Google Scholar 

  90. Schoneshofer, M., Kager, A., and Weber, B. (1983) New “on-line” sample pre-treatment procedure for routine liquid chromatographic assay of low concentration compounds in body fluids, illustrated by triamcinolone assay. Clin. Chem. 29, 1367–1371

    PubMed  CAS  Google Scholar 

  91. Turnell, D. C. and Cooper, J. D. H. (1987) Automated sequential process for preparing samples for analysis by high performance liquid chromatography. J. Chromatogr. 395, 613–621.

    Article  PubMed  CAS  Google Scholar 

  92. Turnell, D. C., Cooper, J. D. H., Green, B., Hughes, G., and Wright, D. J. (1988) Totally automated liquid chromatographic assay for cortisol and associated glucocorticoids in serum with ASTED® sample preparation. Clin. Chem. 34, 1816–1820.

    PubMed  CAS  Google Scholar 

  93. Andersson, S. H. G., and Sjovall, J. (1985) Analysis of profiles of unconjugated steroids in testicular tissue by gas chromatography mass spectrometry. J. Steroid Biochem. 23, 469–475.

    Article  PubMed  CAS  Google Scholar 

  94. Creaser, C. S., Feely, S.J., Houghton, E., and Seymour, M. (1998) Immunoaffinity chromatography combined on-line with highperformance liquid chromatography-mass spectrometry for the determination of corticosteroids. J. Chromatogr. A. 794, 37–43.

    Article  PubMed  CAS  Google Scholar 

  95. Novotny, M., Karlsson, K-E., and Konishi, M. (1984) New biochemical separations using pre-columnn derivatisation and microcolumm liquid chromatography. J. Chromatogr. 305, 159–167.

    Article  Google Scholar 

  96. Kawasaki, T., Maeda, M., and Tsuli, A. (1982) Determination of 17-oxosteroid glucuronides and sulphates in urine and serum by fluoresence high performance liquid chromatography using dansyl hydrazine as a pre-labelling reagent. J. Chromatogr, 233, 61–68.

    Article  PubMed  CAS  Google Scholar 

  97. Iohan, F., Vincze, I., Moonder, C., and Cohen, S. (1991) High performance liquid chromatographic determination of cortolic and cortolonic acids as pyrenyl ester derivatives. J. Chromatogr. 564, 27–41.

    Article  PubMed  CAS  Google Scholar 

  98. Kayama, M. and Taniguchi, H. (1993) Determination of estrogens in plasma by high performance liquid chromatography after precolumn derivatization with 2-(4-carboxyphenyl)-5.6 dimethylbezimidazole. J. Chromatogr. 616, 317–322.

    Article  Google Scholar 

  99. Katayama, M., Masuda, Y., and Taniguchi, H. (1993) Determination of corticosteroids in plasma by high performance liquid chromatography after pre-column derivatization with 2-(4-carboxyphenyl)-5.6 dimethylbezimidazole. J. Chromatogr, 612, 33–39.

    Article  PubMed  CAS  Google Scholar 

  100. Fredline, V. F., Kovacs, E. M., Taylor, P. J., and Johnson, A. G. (1999) Measurement of plasma rennin activity with use of HPLC electrospray tandem mass spectrometry. Clin. Chem. 45, 659–664.

    PubMed  CAS  Google Scholar 

  101. Burch, J. B., Reif, J. S., Noonan, C. W., and Yost, M. G. (2000) Melatonin levels in workers exposed to 60-HZ magnetic fields: work in substations and with 3-phase conductors. J. Occup. Environ. Med. 42, 136–142.

    Article  PubMed  CAS  Google Scholar 

  102. Sanz-Nebot, V., Toro, I., Castillo, A., and Barbosa, J. (2001) Investigation of synthetic peptide hormones by liquid chromatography couples to pneumatically assisted electrospray ionization mass spectrometry: analysis of a synthesis of crude peptide triptorelin. Rapid Commun. Mass Spectrom. 15, 1031–1039.

    Article  PubMed  CAS  Google Scholar 

  103. Cirimele, V., Kintz, P., Dumestre, V., Goulle, J. P., and Ludes, B. (2000) Identification of ten corticosteroids in human hair by liquid chromatrography ionspray mass spectrometry. Forensic Sci. Int. 107, 381–388.

    Article  PubMed  CAS  Google Scholar 

  104. Fitzpatrick, J. L., Ripp, S. I., Smith, N. B., Pierce, W. M., and Prough, R. A. (2001) Metabolism of DHEA by cytochromes P450 in rat and human liver microsomal fractions. Arch. Biochem. Biophys. 389, 278–287.

    Article  PubMed  CAS  Google Scholar 

  105. Garcia, S. I., Porto, P. I., Martinez, V. N., et al. (2000) Expression of TRH and TRH-like peptides in a human glioblastoma-astrocytoma cell line (U-373-MG). J. Endocrinol. 166, 697–703.

    Article  PubMed  CAS  Google Scholar 

  106. Ghilchik, M.W. Tobaruela, M. del Rio-Garcia, J., and Smyth, D.G. (2000) Characterization of neural TRH-like peptides in mammary gland and mammary tumors and milk. Biochim. Biophys. Acta 1475, 55–60.

    PubMed  CAS  Google Scholar 

  107. Krishnan, K. A., Proudman, G. A., and Bahr, J. M. (1994) Purification and partial characterization of isoforms of luteinizing hormone from the chicken pituitary gland. Comp. Biochem. Physiol. Biochem. Mol. Biol. 108, 253–264.

    Article  PubMed  CAS  Google Scholar 

  108. Tanabe, S., Shimohigashi, Y., Nakayama, Y., et al. (1999) In vivo and in vitro evidence of blood brain barrier transport of a novel cationic arginine-vasopressin fragment 4–9 analog. J. Pharmacol. Exp. Ther. 290, 561–568.

    PubMed  CAS  Google Scholar 

  109. Amoresano, A., Siciliano, R., Orru, S., et al. (1996) Structural characterisation of human recombinant glycohormones follitropin, lutropin and chorionic gonadotropin expressed in Chinese hamster ovary cells. Eur. J. Biochem. 242, 608–618.

    Article  PubMed  CAS  Google Scholar 

  110. Secchi, C., Berrini, A., Gaggioli, D., and Borromeo, V. (2001) Amino acid modifications in canine, equine and porcine pituitary growth hormones identified by peptide-mass mapping. J. Chromatogr. B Biomed. Sci. Appl. 757, 237–245.

    Article  PubMed  CAS  Google Scholar 

  111. Janaky, T., Szabo, P., Kele, Z., et al. (1998) Identification of oxytocin and Vasopressin from neurohypophyseal cell culture. Rapid Commun. Mass Spectrom. 12, 1765–1768.

    Article  PubMed  CAS  Google Scholar 

  112. Liu, C. L. and Bowers, L. D. (1996) Immunoaffinity trapping of urinary chorionic gonadotrophin and its high performance liquid chromatography mass spectrometric confirmation. J. Chromatogr. B Biomed. Sci. Appl. 687, 213–220.

    Article  CAS  Google Scholar 

  113. Liu, C. L. and Bowers, L. D. (1997) Mass spectromic characterization of nicked fragments of the beta-subunit of human chorionic gonadotropin. Clin. Chem. 43, 1172–1181.

    PubMed  CAS  Google Scholar 

  114. Black, R. S. and Bowers, L. D. (2000) Mass spectromic characterization on the beta-subunit of human chorionic gonadotropin. Methods Mol. Biol. 146, 337–354.

    PubMed  CAS  Google Scholar 

  115. de Kock, S. S., Rodgers, J. P., and Swanepoel, B. C. (2001) Growth hormone abuse in the horse: preliminary assessment of a mass spectrometric procedure for IGF-1 identification and quantitation. Rapid Commun. Mass Spectrom. 15, 1191–1197.

    Article  PubMed  Google Scholar 

  116. Rafferty, B., Corran, P., and Bristow, A. (2001) Multicenter collaborative study to calibrate salmon calcitonin by bioassay and high performance liquid chromatography: establishment of the third international standard. Bone 29, 84–89.

    Article  PubMed  CAS  Google Scholar 

  117. Iitake, M., Kawasaki S., Sakurai, S., et al. (1998) Serum substances that interfere with thyroid hormone assays in patients with chronic renal failure. Clin. Endocrinol. (Oxf) 48, 739–746.

    Article  Google Scholar 

  118. Fredline, V. F., Taylor, P. J., Dodds, H. M., and Johnson, A. G. (1997) A reference method for the analysis of aldosterone in blood by high performance liquid chromatography atmospheric pressure chemical ionization tandem mass spectrometry. Anal. Biochem. 252, 308–313.

    Article  PubMed  CAS  Google Scholar 

  119. Kippen, A. D., Cerini, F., Vadas, L., Stocklin, R., Vu, L., Offord, R. E., and Rose, K. (1997) Development of an isotope dilution assay for precise determination of insulin, C-peptide and proinsulin levels in non-diabetic and type II diabetic individuals with comparison to immunoassay. J. Biol. Chem. 272, 12,513–12,522.

    Article  PubMed  CAS  Google Scholar 

  120. Kobayashi, N., Kanai, M., Seta, K., and Nakamura, K. (1995) Quantitative analysis of synthetic human calcitonin by liquid chromatography mass spectrometry. J. Chromatogr. B Biomed. Sci. Appl. 672, 17–23.

    Article  CAS  Google Scholar 

  121. Peran, S., Garriga, M. G., Morreale de Esscobar, G., Asuncion, M., and Peran, M. (1997) Increase in thyrotropin levels in hypothyroid patients during treatment due to a defect in the commercial preparation. J. Clin. Endocrinol. Metab. 82, 3192–3195.

    Article  PubMed  CAS  Google Scholar 

  122. Thienpont, L. M., Fierens, C., De Leenheer, A. P., and Przywara, L. (1999) Isotope dilution gas chromatrography mass spectrometry and liquid chromatography electrospray ionization tandem mass spectrometry for the determination of triiodo-L-thyronine in serum. Rapid Commun. Mass Spectrom. 13, 1924–1931.

    Article  PubMed  CAS  Google Scholar 

  123. Oliva, A., Farina, J., and Llabres, M. (2000) Development of two high performance liquid chromatographic methods for the analysis and characterisation of insulin and its degradation products in pharmaceutical preparations. J. Chromatogr. B Biomed. Sci. Appl. 749, 25–34.

    Article  PubMed  CAS  Google Scholar 

  124. Witowska, E., Orlowska, A., Sagan, B., Smoluch, M., and Izdebski, J. (2001) Tryptic hydrolysis of hGH-RH(1-29)-NH2 analogues containing Lys or Orn in positions 12 and 21. J. Pept. Sci. 7, 166–172.

    Article  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Clinical Biochemistry, University College London Hospitals, London, UK

    John W. Honour PhD, FRCPath

Authors
  1. John W. Honour PhD, FRCPath
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. St. Thomas’ Hospital, Guy’s and St. Thomas’ NHS Foundation Trust, London, UK

    Michael J. Wheeler

  2. University of Aberdeen, Aberdeen, UK

    J. S. Morley Hutchinson

Rights and permissions

Reprints and permissions

Copyright information

© 2006 Humana Press Inc.

About this protocol

Cite this protocol

Honour, J.W. (2006). High-Performance Liquid Chromatography for Hormone Assay. In: Wheeler, M.J., Hutchinson, J.S.M. (eds) Hormone Assays in Biological Fluids. Methods in Molecular Biology, vol 324. Humana Press. https://doi.org/10.1385/1-59259-986-9:25

Download citation

  • DOI: https://doi.org/10.1385/1-59259-986-9:25

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-005-2

  • Online ISBN: 978-1-59259-986-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation