What are the proteolytic enzymes of honey and what they do tell us? A fingerprint analysis by 2-D zymography of unifloral honeys

PLoS One. 2012;7(11):e49164. doi: 10.1371/journal.pone.0049164. Epub 2012 Nov 7.


Honey is a sweet and healthy food produced by honeybees (Apis mellifera L.) from flower nectars. Using bidimensional zymography, we have detected the, until now unrevealed, proteolytic activities present in row honey samples. The resulting zymograms were specific for each type of the four unifloral honey under study, and enzymes were identified as serine proteases by the use of specific inhibitors. Further, using bidimensional electrophoresis, we have shown that honey proteases are able to degrade the major Royal Jelly proteins and in particular MRPJ-1, the protein that promotes queen differentiation in honeybees. Our findings open new perspectives for the better understanding of honeybee development, social behaviour and role in honey production. The now discovered honey proteases may influence honey properties and quality, and bidimensional zymograms might be useful to distinguish between different honey types, establish their age and floral origin, and allow honey certification.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bees / enzymology*
  • Chromatography, High Pressure Liquid
  • Honey*
  • Humans
  • Peptide Hydrolases / isolation & purification*
  • Peptide Hydrolases / metabolism
  • Serine Proteases* / classification
  • Serine Proteases* / isolation & purification
  • Serine Proteinase Inhibitors / pharmacology


  • Serine Proteinase Inhibitors
  • Peptide Hydrolases
  • Serine Proteases

Grants and funding

This study was supported by grants from University of Basilicata (Fondo Ateneo, 2008) for the research in food and in part by the project 2010/R/35 of PR on “The molecular basis of nutritional intervention in Multiple Sclerosis”, funded by the Italian Foundation for Multiple Sclerosis (FISM)] for the development of bidimensional zymography (2-DZ). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.