Amino Acids

, Volume 51, Issue 10–12, pp 1623–1631 | Cite as

Evaluation of polyamines as marker of melanoma cell proliferation and differentiation by an improved high-performance liquid chromatographic method

  • Bruno Provenzano
  • Alessandro Lentini
  • Roberta Tatti
  • Angelo De Martino
  • Ilaria Borromeo
  • Carlo Mischiati
  • Giordana Feriotto
  • Cinzia Forni
  • Claudio TabolacciEmail author
  • Simone Beninati
Original Article


The differentiation therapy is focused on the identification of new agents able to impair the proliferative and metastatic potential of cancer cells through the induction of differentiation. Although several markers of cell differentiation on tumor cells have been identified, their causal relationship with neoplastic competence has not been characterized in sufficient detail to propose their use as new pharmacological targets useful for the design of new differentiation agents. Polyamine level in cancer cells and in body fluids was proposed as potential marker of cell proliferation and differentiation. The main advantage of this marker is the possibility to evaluate the antineoplastic activity of new drugs able to induce cell differentiation and consequently to inhibit tumor growth and metastasis. The presented report shows a simply and highly reproducible reverse-phase high-performance liquid chromatographic (HPLC) method for the determination of ortho-phthalaldehyde (OPA) derivatives of polyamines: putrescine (PUT), cadaverine (CAD), spermidine (SPD) and spermine (SPM). The novelty of this method is the fluorescence response for OPA-derivate of SPM, generally low in other procedures, that has been significantly improved by the use of a fully endcapped packing material with minimal silanol interactions. The limits of detection for PUT, CAD, SPD and SPM were 0.6, 0.7, 0.8, and 0.4 pmol/mL, respectively. The analysis time was ≤ 20 min, and the relative recovery rate was of about 97%. To verify the usefulness of this method, it has been validated in a murine melanoma cell line (B16-F10) treated with two theophylline derivatives (namely 8-chlorotheophylline and 8-bromotheophylline). These two compounds increased the activity of tissue transglutaminase (TG2) and the synthesis of melanin, two recognized markers of melanoma cell differentiation, and significantly reduced the levels of intracellular polyamines.


Polyamines OPA derivatization Melanoma cells Differentiation therapy 



Tabolacci C. was supported by Fondazione Umberto Veronesi that is gratefully acknowledged. This work was supported by grants from MIUR (Project PRIN no. 2004067819) and from COST Action 922.

Compliance with ethical standards

Conflict of interest

Authors declare that no conflicts of interest exist.

Research involving human and animal participants

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. This work did not involve experiments with human participants.


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Copyright information

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

Authors and Affiliations

  • Bruno Provenzano
    • 1
  • Alessandro Lentini
    • 1
  • Roberta Tatti
    • 2
  • Angelo De Martino
    • 1
  • Ilaria Borromeo
    • 1
  • Carlo Mischiati
    • 3
  • Giordana Feriotto
    • 4
  • Cinzia Forni
    • 1
  • Claudio Tabolacci
    • 5
    Email author
  • Simone Beninati
    • 1
  1. 1.Department of BiologyUniversity of Rome “Tor Vergata”RomeItaly
  2. 2.Lab. Biomarker Studies and Structure Analysis for Health, Center for Materials and MicrosystemsFondazione Bruno KesslerTrentoItaly
  3. 3.Department of Biomedical Sciences and Surgical SpecialtiesUniversity of FerraraFerraraItaly
  4. 4.Department of Chemical and Pharmaceutical SciencesUniversity of FerraraFerraraItaly
  5. 5.Department of Oncology and Molecular MedicineIstituto Superiore di SanitàRomeItaly

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