Analytical and Bioanalytical Chemistry

, Volume 411, Issue 11, pp 2317–2326 | Cite as

Determination of glyphosate and aminomethylphosphonic acid by sequential-injection reversed-phase chromatography: method improvements and application in adsorption studies

  • Erico A. Oliveira Pereira
  • Vander Freitas Melo
  • Gilberto Abate
  • Jorge C. MasiniEmail author
Research Paper


This paper describes a low-cost reversed-phase sequential injection chromatography method for the determination of glyphosate and aminomethylphosphonic acid in environmental samples. The method is based on the pre-column conversion of glyphosate to glycine by hypochlorite, followed by reaction with o-phthaldialdehyde in presence of 2-mercaptoethanol in borate buffer (pH 9.5) to produce the fluorescent 1-(2′-hydroxyethylthio)-2-N-alkylisoindole. In addition to producing detectable fluorescent indoles, the pre-column derivatization also decreases the polarity of the analytes, favoring their retention on a C18 monolithic column. The isocratic reversed-phase chromatography enabled the separation of both glyphosate and aminomethylphosphonic acid derivatives from polar compounds such as organic acids, humic substances, and carbohydrates which are commonly found in waters and soil extracts. This separation minimizes the laborious sample preparation procedures prior to the analysis. The linear response was observed for concentrations between 0.10 and 12.8 μM. The limits of detection and quantification were 0.03 and 0.10 μM (glyphosate), and 0.015 and 0.050 μM (aminomethylphosphonic acid). At the 0.10 μM concentration level, the relative standard deviations were 21 and 25% for aminomethylphosphonic acid and glyphosate, respectively (n = 5). Recoveries between 80 and 120% were found in the determination of glyphosate and aminomethylphosphonic acid in spiked lake waters (0.80 to 6.4 μM). The method was applied in the determination of kinetic and thermodynamic parameters related to the adsorption of glyphosate on two horizons of an Alfisol from the Paraná State in South Brazil.


Monolithic column Liquid chromatography Herbicides Soil Waters Fluorescence 


Funding information

This work was funded by grants 2013/18507-4 from the São Paulo Research Foundation (FAPESP) and 303940/2017-4 from the National Council for Scientific and Technological Development (CNPq). EAOP received MSc fellowship from CNPq (Grant 134790/2016-2).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This article does not contain any studies made with humans or animals.

Supplementary material

216_2019_1672_MOESM1_ESM.pdf (1.9 mb)
ESM 1 (PDF 1.89 mb)


  1. 1.
    Huhn C. More and enhanced glyphosate analysis is needed. Anal Bioanal Chem. 2018;410:3041–5.CrossRefPubMedGoogle Scholar
  2. 2.
    Saunders L, Pezeshki R. Glyphosate in runoff waters and in the root-zone: a review. Toxics. 2015;3:462–80.CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Koskinen WC, Marek LJ, Hall KE. Analysis of glyphosate and aminomethylphosphonic acid in water, plant materials and soil. Pest Manag Sci. 2016;72:423–32.CrossRefPubMedGoogle Scholar
  4. 4.
    Arkan T, Molnár-Perl I. The role of derivatization techniques in the analysis of glyphosate and aminomethyl-phosphonic acid by chromatography. Microchem J. 2015;121:99–106.CrossRefGoogle Scholar
  5. 5.
    Chen M, Cao Z, Jiang Y, Zhu Z. Direct determination of glyphosate and its major metabolite, aminomethylphosphonic acid , in fruits and vegetables by mixed-mode hydrophilic interaction / weak anion-exchange liquid chromatography coupled with electrospray tandem mass spectrometry. J Chromatogr A. 2013;1272:90–9.CrossRefPubMedGoogle Scholar
  6. 6.
    Hao C, Morse D, Morra F, Zhao X, Yang P, Nunn B. Direct aqueous determination of glyphosate and related compounds by liquid chromatography/tandem mass spectrometry using reversed-phase and weak anion-exchange mixed-mode column. J Chromatogr A. 2011;1218:5638–43.CrossRefPubMedGoogle Scholar
  7. 7.
    Gauglitz G, Wimmer B, Melzer T, Huhn C. Glyphosate analysis using sensors and electromigration separation techniques as alternatives to gas or liquid chromatography. Anal Bioanal Chem. 2018;410:725–46.CrossRefPubMedGoogle Scholar
  8. 8.
    Ruzicka J, Marshall GD. Sequential injection: a new concept for chemical sensors, process analysis and laboratory assays. Anal Chim Acta. 1990;237:329–43.CrossRefGoogle Scholar
  9. 9.
    Idris AM. The second five years of sequential injection chromatography: significant developments in the technology and methodologies. Crit Rev Anal Chem. 2014;44:220–32.CrossRefPubMedGoogle Scholar
  10. 10.
    Hartwell SK, Kehling A, Lapanantnoppakhun S, Grudpan K. Flow injection/sequential injection chromatography: a review of recent developments in low pressure with high performance chemical separation. Anal Lett. 2013;46:1640–71.CrossRefGoogle Scholar
  11. 11.
    Chocholouš P, Solich P, Šatínský D. An overview of sequential injection chromatography. Anal Chim Acta. 2007;600:129–35.CrossRefPubMedGoogle Scholar
  12. 12.
    Šatínský D, Solich P, Chocholouš P, Karlíček R. Monolithic columns - a new concept of separation in the sequential injection technique. Anal Chim Acta. 2003;499:205–14.CrossRefGoogle Scholar
  13. 13.
    dos Santos LBO, Infante CMC, Masini JC. Development of a sequential injection chromatography (SIC) method for determination of simazine, atrazine, and propazine. J Sep Sci. 2009;32:494–500.CrossRefPubMedGoogle Scholar
  14. 14.
    De Prá Urio R, Infante CMC, Masini JC. Online sequential-injection chromatography with stepwise gradient elution: a tool for studying the simultaneous adsorption of herbicides on soil and soil components. J Agric Food Chem. 2013;61:7909–15.CrossRefGoogle Scholar
  15. 15.
    De Miranda Colombo S, Masini JC. A sequential-injection reversed-phase chromatography method for fluorimetric determination of glyphosate and aminomethylphosphonic acid. Anal Methods. 2014;6:490–6.CrossRefGoogle Scholar
  16. 16.
    Rigobello-Masini M, Masini JC. Improvements in the separation capabilities of sequential injection chromatography: determination of intracellular dissolved free amino acid profiles in three taxonomic groups of microalgae. Phytochem Anal. 2013;24:224–9.CrossRefPubMedGoogle Scholar
  17. 17.
    Colombo S de M, Masini JC. Developing a fluorimetric sequential injection methodology to study adsorption/desorption of glyphosate on soil and sediment samples. Microchem J. 2011;98:260–6 d.CrossRefGoogle Scholar
  18. 18.
    Hanke D, Melo V de F, Dieckow J, Dick DP, Bognola IA. Influência da matéria orgânica no diâmetro médio de minerais da fração argila de solos desenvolvidos de basalto no sul do Brasil. Rev Bras Cienc do Solo. 2015;39:1611–22.CrossRefGoogle Scholar
  19. 19.
    Liebman M. Ecological Management of Agricultural Weeds. Cambridge: Cambridge University Press; 2001.CrossRefGoogle Scholar
  20. 20.
    Abdullah MP, Daud J, Hong KS, Yew CH. Improved method for the determination of glyphosate in water. J Chromatogr A. 1995;697:363–9.CrossRefGoogle Scholar
  21. 21.
    Mallat E, Barceló D. Analysis and degradation study of glyphosate and of aminomethylphosphonic acid in natural waters by means of polymeric and ion-exchange solid-phase extraction columns followed by ion chromatography-post-column derivatization with fluorescence detection. J Chromatogr A. 1998;823:129–36.CrossRefPubMedGoogle Scholar
  22. 22.
    Reis BF, Giné MF, Zagatto EAG, Lima JLFC, Lapa RA. Multicommutation in flow analysis. Part 1. Binary sampling: concepts, instrumentation and spectrophotometric determination of iron in plant digests. Anal Chim Acta. 1994;293:129–38.CrossRefGoogle Scholar
  23. 23.
    Vieira JA, Raimundo IM, Reis BF, Zagatto EAG, Lima JLFC. Sampling strategies in sequential injection analysis: exploiting the monosegmented-flow approach. Anal Chim Acta. 1998;366:257–62.CrossRefGoogle Scholar
  24. 24.
    Sierra MMD, Giovanela M, Parlanti E, Soriano-Sierra EJ. Fluorescence fingerprint of fulvic and humic acids from varied origins as viewed by single-scan and excitation/emission matrix techniques. Chemosphere. 2005;58:715–33.CrossRefPubMedGoogle Scholar
  25. 25.
    Waiman C, Avena MJ, Garrido M, Fernández Band B, Zanini GP. A simple and rapid spectrophotometric method to quantify the herbicide glyphosate in aqueous media. Application to adsorption isotherms on soils and goethite. Geoderma. 2012;170:154–8.CrossRefGoogle Scholar
  26. 26.
    Kaczyński P, Łozowicka B. Liquid chromatographic determination of glyphosate and aminomethylphosphonic acid residues in rapeseed with MS/MS detection or derivatization/fluorescence detection. Open Chem. 2015;13:1011–9.CrossRefGoogle Scholar
  27. 27.
    Gübeli T, Christian GD, Ruzicka J. Fundamentals of sinusoidal flow sequential injection spectrophotometry. Anal Chem. 1991;63:2407–13.CrossRefPubMedGoogle Scholar
  28. 28.
    Ho YS, McKay G. The sorption of lead(II) ions on peat. Water Res. 1999;33:578–84.CrossRefGoogle Scholar
  29. 29.
    Fierro V, Torné-Fernández V, Montané D, Celzard A. Adsorption of phenol onto activated carbons having different textural and surface properties. Microporous Mesoporous Mater. 2008;111:276–84.CrossRefGoogle Scholar
  30. 30.
    Vereecken H. Mobility and leaching of glyphosate: a review. Pest Manag Sci. 2005;61:1139–51.CrossRefPubMedGoogle Scholar
  31. 31.
    Pignatello JJ, Xing B. Mechanisms of slow sorption of organic chemicals to natural particles. Environ Sci Technol. 1996;30:1–11.CrossRefGoogle Scholar
  32. 32.
    Kosmulski M, Maczka E, Jartych E, Rosenholm JB. Synthesis and characterization of goethite and goethite-hematite composite: experimental study and literature survey. Adv Colloid Interf Sci. 2003;103:57–76.CrossRefGoogle Scholar
  33. 33.
    Gómez Ortiz AM, Okada E, Bedmar F, Costa JL. Sorption and desorption of glyphosate in mollisols and ultisols soils of Argentina. Environ Toxicol Chem. 2017;36:2587–92.CrossRefPubMedGoogle Scholar
  34. 34.
    Ghafoor A, Jarvis NJ, Stenström J. Modelling pesticide sorption in the surface and subsurface soils of an agricultural catchment. Pest Manag Sci. 2013;69:919–29.CrossRefPubMedGoogle Scholar
  35. 35.
    Gros P, Ahmed A, Kühn O, Leinweber P. Glyphosate binding in soil as revealed by sorption experiments and quantum-chemical modeling. Sci Total Environ. 2017;586:527–35.CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  • Erico A. Oliveira Pereira
    • 1
  • Vander Freitas Melo
    • 2
  • Gilberto Abate
    • 3
  • Jorge C. Masini
    • 1
    Email author
  1. 1.Departamento de Química Fundamental, Instituto de QuímicaUniversidade de São PauloSão PauloBrasil
  2. 2.Departamento de Solos e Engenharia Agrícola, Setor de Ciências AgráriasUniversidade Federal do ParanáCuritibaBrazil
  3. 3.Departamento de QuímicaUniversidade Federal do Paraná, Centro PolitécnicoCuritibaBrazil

Personalised recommendations