Skip to main content
SpringerLink
Log in

RP-HPLC determination of phytoplankton pigments—comparison of calibration results for two columns

  • Research Article
  • Published:
Marine Biology Aims and scope Submit manuscript

Abstract

In the experiment reported the results obtained with two different columns for the RP-HPLC analysis of phytoplankton pigment components were compared. LiChroCART Hypersil ODS and LiChroCART LiChrospher 100 RP18e columns were used exchangeably. Two groups of high-purity pigment standards of reference phytoplankton strains (23 chlorophylls and carotenoids in the first case and 25 in the second case) were subjected to chromatographic analysis. All calibration parameters (retention times, calibration curves, spectrum shapes, detection thresholds) for qualitative and quantitative identification of pigments were determined. The results indicate that both columns used in the analysis ensure sufficient resolution and can be used interchangeably for the analysis of pigments present in naturally existing phytoplankton, even when present in only trace amounts.

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

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  • Andersen RA, Bidigare RR, Keller MD, Latasa M (1996) A comparison of HPLC pigment signatures and electron microscopic observations for oligotrophic waters of the North Atlantic and Pacific Oceans. Deep-Sea Res II 43:517–537

    Google Scholar 

  • Barlow RG, Mantoura RFC, Gough MA, Fileman TW (1993) Pigment signatures of the phytoplankton composition in the north-eastern Atlantic during the 1990 spring bloom. Deep-Sea Res II 40:459–477

    Google Scholar 

  • Barlow RG, Aiken J, Holligan PM, Cummings DG, Maritorena S, Hooker S (2002) Phytoplankton pigment and absorption characteristics along meridional transects in the Atlantic Ocean. Deep-Sea Res I 49:637–660

    Google Scholar 

  • Bidigare RR, Kennicutt II MC, Brooks JM (1985) Rapid determination of chlorophylls and their degradation products by high-performance liquid chromatography. Limnol Oceanogr 30:432–435

    Google Scholar 

  • Bjørnland T (1997) UV-vis spectroscopy of carotenoids. In: Jeffrey SW, Mantoura RFC, Wright SW (eds) Phytoplankton pigments in oceanography: guidelines to modern methods. UNESCO, Paris, pp 578–594

  • Bjørnland T, Haxo FT, Liaaen-Jensen S (2003) Carotenoids of the Florida red tide dinoflagellate Karenia brevis. Biochem Syst Ecol 31:1147–1162

    Google Scholar 

  • Brotas V, Plante-Cuny MR (2003) The use of HPLC pigment analysis to study microphytobenthos communities. Acta Oecol 24:109–115

    Google Scholar 

  • Claustre H, Hooker SB, Heukelem L, Berthon JF, Barlow R, Ras J, Sessions H, Targa C, Thomas CS, Linde D, Marty JC (2004) An intercomparison of HPLC phytoplankton pigments methods using in situ samples: application to remote sensing and database activities. Mar Chem 85:41–61

    Google Scholar 

  • Gieskes WWC, Kraay GW (1983) Dominance of Cryptophyceae during the phytoplankton spring bloom in the central North Sea detected by HPLC analysis of pigments. Mar Biol 75:179–185

    Google Scholar 

  • Gieskes WWC, Kraay GW (1989) Estimating the carbon-specific growth rate of the major algal species groups in eastern Indonesian waters by 14C labeling of taxon-specific carotenoids. Deep-Sea Res 36:1127–1139

    Google Scholar 

  • Gieskes WWC, Kraay GW, Nontji A, Setiapermana D, Sutomo (1988) Monsoonal alternation of a mixed and a layered structure in the phytoplankton of the euphotic zone of the Banda Sea (Indonesia): a mathematical analysis of algal pigment fingerprints. Neth J Sea Res 22:123–137

    Google Scholar 

  • Goericke R, Repeta DJ (1992) The pigments of Prochlorococcus marinus: presence of divinyl chlorophyll a and b in marine prokaryotes. Limnol Oceanogr 37:425–433

    Google Scholar 

  • Heukelem L, Thomas CS (2001) Computer-assisted high-performance liquid chromatography method development with applications to the isolation and analysis of phytoplankton pigments. J Chromatogr A 910:31–49

    Google Scholar 

  • Heukelem L, Lewitus AJ, Kana TM (1992) High-performance liquid chromatography of phytoplankton pigments using a polymeric reverse-phase C18 column. J Phycol 28:867–872

    Google Scholar 

  • Heukelem L, Lewitus AJ, Kana TM, Craft NE (1994) Improved separations of phytoplankton pigments using temperature-controlled high performance liquid chromatography. Mar Ecol Prog Ser 114:303–313

    Google Scholar 

  • Jeffrey SW (1997a) Chlorophyll and carotenoid excitation coefficients. In: Jeffrey SW, Mantoura RFC, Wright SW (eds) Phytoplankton pigments in oceanography: guidelines to modern methods. UNESCO, Paris, pp 595–596

  • Jeffrey SW (1997b) Preparation of chlorophyll standards. In: Jeffrey SW, Mantoura RFC, Wright SW (eds) Phytoplankton pigments in oceanography: guidelines to modern methods. UNESCO, Paris, pp 207–238

  • Jeffrey SW, Humphrey GF (1975) New spectrophotometric equations for determining chlorophylls a, b, c1 and c2 in higher plants, algae, and natural phytoplankton. Biochem Physiol Pflanz (BPP) 167:191–194

    Google Scholar 

  • Jeffrey SW, Vesk M (1997) Introduction to marine phytoplankton and their pigment signatures. In: Jeffrey SW, Mantoura RFC, Wright SW (eds) Phytoplankton pigments in oceanography: guidelines to modern methods. UNESCO, Paris, pp 37–84

  • Kraay GW, Zapata M, Veldhuis MJW (1992) Separation of chlorophylls c 1 , c 2 and c 3 of marine phytoplankton by reversed-phase C18 high-performance liquid chromatography. J Phycol 28:708–712

    Google Scholar 

  • Latasa M, Bidigare RR, Ondrusek ME, Kennicutt II MC (1996) HPLC analysis of algal pigments: a comparison exercise among laboratories and recommendations for improved analytical performance. Mar Chem 51:315–324

    Google Scholar 

  • MacIntyre HL, Kana TM, Geider RJ (2000) The effect of water motion on the short-term rates of photosynthesis by marine phytoplankton. Trends Plant Sci 5:12–17

    CAS  PubMed  Google Scholar 

  • Mackey DJ, Blanchot J, Higgins HW, Neveux J (2002) Phytoplankton abundances and community structure in the equatorial Pacific. Deep-Sea Res II 49:2561–2582

    Google Scholar 

  • Mantoura RFC, Llewellyn CA (1983) The rapid determination of algal chlorophyll and carotenoid pigments and their breakdown products in natural waters by reverse-phase high-performance liquid chromatography. Anal Chim Acta 151:297–314

    Article  CAS  Google Scholar 

  • Mantoura RFC, Repeta DJ (1997) Calibration methods for HPLC. In: Jeffrey SW, Mantoura RFC, Wright SW (eds) Phytoplankton pigments in oceanography: guidelines to modern methods. UNESCO, Paris, pp 407–428

  • Marty JC, Chiavlerini J, Pizay MD, Avril B (2002) Seasonal and interannual dynamics of nutrients and phytoplankton pigments in the western Mediterranean Sea at the DYFAMED time-series station (1991–1999). Deep-Sea Res II 49:1965–1985

    Google Scholar 

  • Obayashi Y, Tanoue E (2002) Growth and mortality rates of phytoplankton in the northwestern North Pacific estimated by the dilution method and HPLC pigment analysis. J Exp Mar Biol Ecol 280:33–52

    Google Scholar 

  • Qian Y, Jochens AE, Kennicutt II MC, Biggs DC (2003) Spatial and temporal variability of phytoplankton biomass and community structure over the continental margin of the northeast Gulf of Mexico based on pigment analysis. Contin Shelf Res 23:1–17

    Google Scholar 

  • Repeta DJ, Bjørland T (1997) Preparation of carotenoid standards. In: Jeffrey SW, Mantoura RFC, Wright SW (eds) Phytoplankton pigments in oceanography: guidelines to modern methods. UNESCO, Paris, pp 239–260

  • Rodriguez F, Varela M, Zapata M (2002) Phytoplankton assemblages in the Gerlache and Bransfield Straits (Antarctic Peninsula) determined by light microscopy and CHEMTAX analysis of HPLC pigment data. Deep-Sea Res II 49:723–747

    Google Scholar 

  • Roy S, Chanut J, Gosselin M, Sime-Ngando T (1996) Characterization of phytoplankton communities in the lower St. Lawrence Estuary using HPLC—detected pigments and cell microscopy. Mar Ecol Prog Ser 142:55–73

    Google Scholar 

  • Schmid H, Stich HB (1995) HPLC-analysis of algal pigments: comparison of columns, column properties and eluents. J Appl Phycol 7:487–494

    Google Scholar 

  • Stoń J, Kosakowska A (2002) Phytoplankton pigment designation—an application of RP-HPLC in qualitative and quantitative analysis. J Appl Phycol 14:205–210

    Google Scholar 

  • Stoń J, Kosakowska A, Łotocka M, Łysiak-Pastuszak E (2002) Pigment composition in relation to phytoplankton community structure and nutrient content in the Baltic Sea. Oceanologia 44:419–437

    Google Scholar 

  • Vidussi F, Claustre H, Bustillos-Guzman J, Cailliau C, Marty J (1996) Determination of chlorophylls and carotenoids of marine phytoplankton: separation of chlorophyll a from divinyl-chlorophyll a and zeaxanthin from lutein. J Plankton Res 18:2377–2382

    Google Scholar 

  • Woźniak B, Dera J, Majchrowski R, Ficek D, Koblentz-Mishke OI, Darecki M (1997) PAS initial model of marine primary production for remote sensing application. Oceanologia 39:377–395

    Google Scholar 

  • Woźniak B, Dera J, Ficek D, Majchrowski R, Kaczmarek S, Ostrowska M, Koblentz-Mishke OI (2000) Model of the in vivo spectral absorption of algal pigments, part 1. Mathematical apparatus. Oceanologia 42:177–190

    Google Scholar 

  • Wright SW, Enden RL (2000) Phytoplankton community structure and stocks in the eastern Antarctic marginal ice zone (BROKE survey, January–March 1996) determined by CHEMTAX analysis of HPLC pigment signatures. Deep-Sea Res II 47:2363–2400

    Google Scholar 

  • Wright SW, Jeffrey SW (1997) High-resolution HPLC system for chlorophylls and carotenoids of marine phytoplankton. In: Jeffrey SW, Mantoura RFC, Wright SW (eds) Phytoplankton pigments in oceanography: guidelines to modern methods. UNESCO, Paris, pp 327–342

  • Wright SW, Jeffrey SW, Mantoura RFC, Llewellyn CA, Bjornland T, Repeta D, Welschmeyer N (1991) Improved HPLC method for the analysis of chlorophylls and carotenoids from marine phytoplankton. Mar Ecol Prog Ser 77:183–196

    Google Scholar 

  • Zapata M, Rodriguez F, Garrido JL (2000) Separation of chlorophylls and carotenoids from marine phytoplankton: a new HPLC method using reversed phase C8 column and pyridine-containing mobile phases. Mar Ecol Prog Ser 195:29–45

    CAS  Google Scholar 

  • Zapata M, Edvardsen B, Rodriguez F, Maestro MA, Garrido JL (2001) Chlorophyll c2 monogalactosyldiacylglyceride ester (chl c2-MGDG). A novel marker pigment for Chysochromulina species (Haptophyta). Mar Ecol Prog Ser 219:85–98

    Google Scholar 

Download references

Acknowledgements

This work was carried out within the framework of the PAS Institute of Oceanology statutory activities, grant no. I.2.2, and was financed in part from the National Project funded by the Polish State Committee for Scientific Research no. PBZ-KBN 056/P04/2001 and grant no. 2 PO4 052 26/2004–2006.

Author information

Authors and Affiliations

  1. Institute of Oceanology, Polish Academy of Sciences, Powstańców Warszawy 55, 81-712, Sopot, Poland

    J. Stoń-Egiert & A. Kosakowska

Authors
  1. J. Stoń-Egiert
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. A. Kosakowska
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to J. Stoń-Egiert.

Additional information

Communicated by O. Kinne, Oldendorf/Luhe

Rights and permissions

Reprints and permissions

About this article

Cite this article

Stoń-Egiert, J., Kosakowska, A. RP-HPLC determination of phytoplankton pigments—comparison of calibration results for two columns. Marine Biology 147, 251–260 (2005). https://doi.org/10.1007/s00227-004-1551-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00227-004-1551-z

Keywords

Search

Navigation