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Molecules 2019, 24, 2692 13 of 17 30%. Therefore, the reduction in EPS via aPDT could reduce/destroy the protection of all three species of the bacteria. Figure 6. Polysaccharide production by 4-day biofilms of (A) P. gingivalis, (B) P. intermedia and (C) F. nucleatum on dentin squares (mean ± sd; n = 6). Dissimilar letters indicated significant differences between each group (p < 0.05). Clinically, the subgingival biofilm is an aggregation of multispecies bacteria. Multispecies biofilms are more challenging to eradicate than single species biofilms and planktonic bacteria [46]. The early attached dominant species of bacteria are streptococci and members of the yellow and purple complexes, such as Actinomycess pp. which soon develop a polymicrobial community [47]. However, most studies also have been done in planktonic or single-species biofilms without regard for the complex microbial and biochemical changes occurring simultaneously [24]. Besides the standardized simple model for culturing in vitro, intact single-species biofilms were easy to realize in a normal laboratory. Within the limitations of this in vitro study, the biofilm model gave a simple means of determining the antimicrobial efficacy of novel nanocomposite with Mn doping upon NIR irradiation against three key periodontal pathogens. This methodology may be more clinically representative than the methods, which do not consider the microorganism in biofilms [48]. However, it still does not reproduce what happens clinically in the periodontal pocket. In such an environment, several mechanisms allow the growth and selection of several microorganisms, even after the treatment. Therefore, further studies should focus on the susceptibility of novel nanocomposite based aPDT against the multispecies biofilms and confirm the effects in animal studies. 4. Conclusions NaYF4@Ce6@silane NPs with upconversion triggered aPDT function were designed in the treatment of periodontal disease. This upconversion fluorescent nanomaterial can overcome the limited tissue penetration depth of visible light in conventional PDT due to the infrared light excitation. The efficient photosensitizer Ce6 molecules were combined with NaYF4:Yb,Er NPs by amphiphilic silane method, through a hydrophobic-hydrophobic interaction between the hydrophobic side chain of the silane and the hydrophobic groups of the UCNPs, avoiding the leakage of the Ce6 molecules. With 980 nm NIR light excitation, the upconversion red emission can efficiently trigger the aPDT effect, because the primary excitation band of Ce6 molecules is totally overlapped with the upconversion red emission band of its UCNPs carrier, and, also, the Ce6 molecule is located in the hydrophobic thin layer of the NPs. Mn ions were doped into the UCNPs for the enhanced red emission and obtained improved energy transfer to Ce6 molecules for stimulating the aPDT function. The novel UCNPs had excellent biocompatibility low cytotoxicity with above 90% cell viability at the concentration of 200 μg/mL. By investigating the aPDT function on three bacterial of P. gingivalis, P. intermedia and F. nucleatum, an enhanced bacteriological therapeutic effect on bacterial and the corresponding biofilms was obtained with the increased Mn doping amount from 0 to 30%. Especially for Mn30% doped UCNPs, the three bacterial CFU counts of 4-day biofilms all showed a logarithmic reduction of more than 2 log. Moreover, Mn30% doped UCNPs had the lowest polysaccharide production among allPDF Image | Anti-Biofilm Property of Bioactive Upconversion Nanocomposites
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