Indirect determination of pepsin activity in human seric protein samples by polyphenol oxidase electrodes

Erica S. Forzani, María E. Bernardi, Ada M. Sisti, Jorge A. Zarzur, Velia M. Solis

Research output: Contribution to journalArticlepeer-review

4 Scopus citations

Abstract

We present a new chronoamperometric methodology for the indirect determination of pepsin activity in proteic solutions. The high-sensitive polyphenol oxidase (PPO) carbon paste bioelectrodes, prepared with 150-200U of PPO adsorbed on the carbon paste surface and trapped behind a dialysis membrane were able to detect free L-tyrosine (L-Tyr) as well as L-Tyr-containing peptides, the product of pepsin hydrolysis on haemoglobin samples. The electroactive enzymatic products were determined by their reduction current at -0.050V, a working potential low enough for preventing electrochemical interferences. The external diffusional barrier posed by the dialysis membrane and the high PPO-superficial concentrations allowed us to prevent electrode surface fouling and to reach stationary diffusional currents after 3min. The electrochemical L-Tyr concentrations measured with proper internal standards, were correlated to those determined by the spectrophotometric reference method (Folin-Ciocalteu), reaching a linear relationship with a slope of 1.05±0.04 and a linear regression coefficient of 0.9951. These results as well as a recovery assay of 98% and the detection limit lower than that of the spectrophotometric method indicate that the proposed methodology is very satisfactory and potentially useful for indirect determination of pepsin in biological samples. The technique was also applied for the determination of pepsin activity in hydrolysed immuno globulin solutions of different composition.

Original languageEnglish (US)
Pages (from-to)163-175
Number of pages13
JournalAnalytica Chimica Acta
Volume460
Issue number2
DOIs
StatePublished - Jun 5 2002
Externally publishedYes

Keywords

  • Enzymatic electrodes
  • L-Tyr-containing peptides
  • L-Tyrosine
  • Pepsin activity
  • Polyphenol oxidase

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Environmental Chemistry
  • Spectroscopy

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