Chlorhexidine preserves the hybrid layer in vitro after 10-years aging

Lorenzo Breschi, Tatjana Maravic, Allegra Comba, Sandra Ribeiro Cunha, Alessandro D. Loguercio, Alessandra Reis, Viviane Hass, Milena Cadenaro, Edoardo Mancuso, Eric Mayer-Santos, Lina Niu, David H. Pashley, Franklin R. Tay, Annalisa Mazzoni

Research output: Contribution to journalArticlepeer-review

36 Scopus citations


Objective: The present study investigated the ability of a chlorhexidine (CHX)-containing primer (0.2% aqueous solution) to inhibit dentinal enzymes, preserve the hybrid layer (HL) and remain within the HL, after 10 years of aging in artificial saliva at 37 °C. Methods: Non-carious extracted molars were assigned to two groups, cut into slabs exposing middle/deep dentin, etched and bonded with Adper Scotchbond 1XT (SB1XT) with or without 0.2% CHX aqueous solution pretreatment. Composite build-ups were made, and the specimens were cut in 1-mm thick bonded sticks. In situ zymography was performed on freshly prepared specimens (T0) and specimens aged for 10 years (T10-yr) at 37 °C in artificial saliva, to investigate endogenous gelatinolytic activity within the HL. At T10-yr, specimens were also decalcified and embedded in epoxy resin for TEM analysis. Micro-Raman spectroscopy was performed at T0 and T10-yr to evaluate the chemical profiles in intertubular dentin and the HL. Results: In situ zymography showed less pronounced enzymatic activity in the CHX-pretreated group (p < 0.05) regardless of aging, maintaining a similar level of fluorescence at T0 and T10-yr (p > 0.05). TEM results showed that 98% of the HL had been degraded in the control group, while 95% of the HL was intact in the experimental group. Moreover, all the Raman spectra peaks assigned to CHX could be identified only in the CHX-pretreated group (T0 and T10-yr). Significance: In vitro, CHX remains in the HL after 10 years with its inhibitory effect preserved. This may be the underlying factor for HL preservation after this long aging period.

Original languageEnglish (US)
Pages (from-to)672-680
Number of pages9
JournalDental Materials
Issue number5
StatePublished - May 2020


  • Dentin bonding agents
  • Hybrid layer
  • In situ zymography
  • In vitro aging
  • MMPs inhibitors
  • Raman spectroscopy
  • TEM

ASJC Scopus subject areas

  • General Materials Science
  • General Dentistry
  • Mechanics of Materials


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