The study on the effect of amino acid porphyrin conjugate-mediated antimicrobial photodynamic therapy on Streptococcus mutans biofilm

Abstract

Antimicrobial Photodynamic Therapy (aPDT) based on the action of visible light and photosensitizers has emerged as a promising microbial reduction and alternative to antibiotics resistance to cariogenic pathogens. The present research aims to evaluate the antimicrobial effect of aPDT mediated by a new photosensitizer (amino acid porphyrin conjugate 4i) on Streptococcus mutans (S. mutans) biofilm. Qualitative morphologic characteristics of S. mutans biofilms are shown by scanning electron microscopy (SEM). The colony plate counting method is used to measure the dark toxicity and the phototoxicity of different concentrations of 4i-aPDT to S. mutans biofilms. MTT assay is conducted to investigate the effect of 4i mediated aPDT on the metabolic activity of S. mutans biofilm. Changes in structure morphology, bacterial density and extracellular matrix of S. mutans biofilm are observed by SEM. The distribution of living and dead bacteria in biofilm is detected using Confocal laser microscopy (CLSM). The results indicate that single laser irradiation has no antibacterial effect on S. mutans biofilms. With the increase of 4i concentration or the prolongation of laser irradiation time, the antibacterial effect of 4i-mediated aPDT on S. mutans biofilm is more statistically significant compared to the control. When the concentration of 62.5 µmol/L 4i is continuously illuminated for 10 min, the logarithm of the colonies in the biofilm shows a reduction of 3.4 log10. MTT assay detected absorbance values of biofilm by 4i-mediated aPDT are the lowest, indicating a significant decrease in biofilm metabolic activity. SEM analysis shows that 4i mediated aPDT reduced the quantity and density of S. mutans. A dense red fluorescence image of the 4i-aPDT treated biofilm is observed under CLSM, indicating that the dead bacteria are widely distributed.