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(1) Background: For any antibacterial oral formulation to be successful, it must present effects in the presence of biofilms. Therefore, our aim is to analyze the drug release and the antibiofilm effects of a semi-solid formulation containing chlorhexidine (CHX) in the presence of pathogenic biofilms. (2) Methods: The biofilms of Streptococcus mutans ( n = 6) or Porphyromonas gingivalis ( n = 3) were formed for 6 and 4 days, respectively, being exposed to: 1) a CHX system or 2) vehicle control without CHX. A group without treatment was included as negative control. The acidogenicity, CHX quantification and bacterial viability were determined. A dissolution assay in a buffer and culture medium in the absence of bacteria was also performed. (3) Results: Although the CHX quantification in the culture medium of both biofilms was lower compared to the buffer ( p &lt; 0.05) and the culture medium in the absence of bacteria, the CHX system was able to display antibiofilm effects until 96 h for the S. mutans biofilms ( p &lt; 0.05) and 72 h for the P. gingivalis biofilms ( p &lt; 0.05). (4) Conclusions: The experimental formulation is able to extend chlorhexidine effects, even in challenging conditions such as in the presence of bacteria, allowing the in vitro control of cariogenic biofilms for 4 days and periodontopathogenic biofilms for 3 days.

Effect of an Experimental Formulation Containing Chlorhexidine on Pathogenic Biofilms and Drug Release Behavior in the Presence or Absence of Bacteria