Department of Restorative Dentistry, Ribeirão Preto School of Dentistry, University of São Paulo (USP), Avenida do Café, s/n°, Monte Alegre, Ribeirão Preto, São Paulo, 14040-904, Brazil.
Department of Chemistry, Center of Nanotechnology and Tissue Engineers, Photobiology and Photomedicine Research Group, FFCLRP-University of São Paulo (USP), Avenida do Café, s/n°, Monte Alegre, Ribeirão Preto, São Paulo, 14040-904, Brazil.
Lasers Med Sci. 2022 Apr;37(3):2033-2043. doi: 10.1007/s10103-021-03466-z. Epub 2021 Nov 23.
This study aimed to characterize the aluminum phthalocyanine chloride (AlClPc) encapsulated in chitosan nanoparticles (CN) and apply it in antimicrobial photodynamic therapy (aPDT) on multispecies biofilm composed of Streptococcus mutans, Lactobacillus casei, and Candida albicans to analyze the antimicrobial activity and lactate production after treatment. Biofilms were formed in 24-well polystyrene plates at 37 °C for 48 h under microaerophilia. The following groups were evaluated (n = 9): as a positive control, 0.12% chlorhexidine gluconate (CHX); phosphate-buffered saline (PBS) as a negative control; 2.5% CN as release vehicle control; the dark toxicity control of the formulations used (AlClPc and AlClPc + CN) was verified in the absence of light; for aPDT, after 30 min incubation time, the photosensitizers at a final concentration of 5.8 × 10 mg/mL were photoirradiated for 1 min by visible light using a LED device (AlClPc + L and AlClPc + CN + L) with 660 nm at the energy density of 100 J/cm. An in vitro kit was used to measure lactate. The biofilm composition and morphology were observed by scanning electron microscopy (SEM). The antimicrobial activity was analyzed by quantifying colony forming units per mL (CFU/mL) of each microorganism. Bacterial load between groups was analyzed by ANOVA and Tukey HSD tests (α = 0.05). A lower lactate dosage was observed in the aPDT AlClPc + CN + L and CHX groups compared to the CN and AlClPc groups. The aPDT mediated by the nanoconjugate AlClPc + CN + L showed a significant reduction in the viability of S. mutans (3.18 log10 CFU/mL), L. casei (4.91 log10 CFU/mL), and C. albicans (2.09 log10 CFU/mL) compared to the negative control PBS (p < 0.05). aPDT using isolated AlClPc was similar to PBS to the three microorganisms (p > 0.05). The aPDT mediated by the nanoconjugate AlClPc + CN + L was efficient against the biofilm of S. mutans, L. casei, and C. albicans.
本研究旨在表征包裹在壳聚糖纳米粒子（CN）中的铝酞菁氯（AlClPc），并将其应用于由变形链球菌、干酪乳杆菌和白色念珠菌组成的多物种生物膜的抗菌光动力疗法（aPDT）中，以分析治疗后的抗菌活性和乳酸产量。生物膜在 37°C 下于微需氧条件下在 24 孔聚苯乙烯板中形成 48 小时。评估了以下组（n=9）：作为阳性对照，0.12%葡萄糖酸氯己定（CHX）；磷酸盐缓冲盐水（PBS）作为阴性对照；2.5%CN 作为释放载体对照；在没有光的情况下验证了制剂的暗毒性对照（AlClPc 和 AlClPc+CN）；对于 aPDT，在 30 分钟孵育时间后，将终浓度为 5.8×10mg/mL 的光敏剂用可见光照射 1 分钟，使用 LED 装置（AlClPc+L 和 AlClPc+CN+L），波长为 660nm，能量密度为 100J/cm。使用体外试剂盒测量乳酸。通过扫描电子显微镜（SEM）观察生物膜的组成和形态。通过定量每毫升（mL）每个微生物的菌落形成单位（CFU/mL）来分析抗菌活性。通过方差分析和 Tukey HSD 检验（α=0.05）分析组间细菌负荷。与 CN 和 AlClPc 组相比，aPDT AlClPc+CN+L 组和 CHX 组的乳酸剂量较低。与阴性对照 PBS 相比，纳米复合物 AlClPc+CN+L 介导的 aPDT 显著降低了变形链球菌（3.18 log10 CFU/mL）、干酪乳杆菌（4.91 log10 CFU/mL）和白色念珠菌（2.09 log10 CFU/mL）的活力（p<0.05）。单独使用 AlClPc 的 aPDT 对三种微生物与 PBS 相似（p>0.05）。纳米复合物 AlClPc+CN+L 介导的 aPDT 对变形链球菌、干酪乳杆菌和白色念珠菌的生物膜有效。
