In vitro bactericidal effects of 625, 525, and 425 nm wavelength (red, green, and blue) light-emitting diode irradiation

Abstract

Objective: The purpose of this study was to evaluate the relationship of 625, 525, and 425 nm wavelengths, providing average power output and effects on three common pathogenic bacteria.

Background data: Ultraviolet (UV) light kills bacteria, but the bactericidal effects of UV may not be unique, as 425 nm produces a similar effect. The bactericidal effects of light-emitting diode (LED) wavelengths such as 625 and 525 nm have not been described. Before conducting clinical trials, the appropriate wavelength with reasonable dose and exposure time should be established.

Materials and methods: The bactericidal effects of 625, 525, and 425 nm wavelength LED irradiation were investigated in vitro for the anaerobic bacterium Porphyromonas gingivalis and two aerobes (Staphylococcus aureus and Escherichia coli DH5α). Average power output was 6 mW/cm(2) for 1 h. The bacteria were exposed to LED irradiation for 1, 2, 4, and 8 h (21.6, 43.2, 86.4, and 172.8 J/cm(2), respectively). LED irradiation was performed during growth on agar and in broth. Control bacteria were incubated without LED irradiation. Bacterial growth was expressed in colony-forming units (CFU) and at an optical density at 600 nm in agar and broth.

Results: The bactericidal effect of LED phototherapy depended upon wavelength, power density, bacterial viable number, and bacteria species. The bactericidal effect of 425 and 525 nm irradiation varied depending upon the bacterial inoculation, compared with unirradiated samples and samples irradiated with red light. Especially, P. gingivalis and E. coli DH5α were killed by 425 nm, and S. aureus growth was inhibited by 525 nm. However, the wavelength of 625 nm was not bactericidal for P. gingivalis, E. coli DH5α, or S. aureus.

Conclusions: Irradiation at 625 nm light was not bactericidal to S. aureus, E. coli, and P. gingivalis, whereas wavelengths of 425 and 525 nm had bactericidal effects. S. aureus was also killed at 525 nm.

Figures

FIG. 1.

Schematic diagram of setup for the three-in-one approach mounting the RGB light-emitting diode (LED) package (a) agar plate and (b) broth setup. The LED was positioned 50 mm directly over the Petri dish and Erlenmeyer flask. The three-in-one approach mounting the RGB LED package for light irradiation and the power supply system was set in an incubator for the experiment (see Fig. 2). The electrical circuit system for experiment controlled time, color of RGB, current, and voltage.

FIG. 2.

(a) Power supply system and (b) three-in-one approach mounting the RGB light-emitting diode (LED) package for light irradiation. Red LED peak at 625 nm presented spectral width 620∼630 nm. Green LED peak at 525 nm presented spectral width 520∼530 nm. Blue LED peak at 425 nm presented spectral width 420∼430 nm.

FIG. 3.

Bacterial survival by LED irradiation on Porphyromonas gingivalis. (A) Bacterial survival is shown in optical density (OD600) of 0.1 mL of 1.6×109 colony-forming units (CFU)/mL. The X-axis represents irradiated time, and OD600 is shown on the Y-axis. (B) Bacterial survival is shown in OD600 of 0.1 mL of 2×108 CFU/mL. The X-axis represents irradiated time, and OD600 is shown on the Y-axis. The data are expressed as the mean±SEM and analyzed by using the two way analysis of variance (ANOVA). *p<0.05, **p<0.01, ***p<0.001.

FIG. 4.

Bacterial survival of light-emitting diode (LED) irradiation by 6 mW/cm2; for 1 h irradiation in Porphyromonas gingivalis. (A) Bacterial survival is shown in colony forming units (CFU). The X-axis represents irradiated time, and CFU are shown on the Y-axis. (B) Representative photographs showing colonies of P. gingivalis irradiated at 48 mW/cm2 for 8 h in agar plates. (a) Control, (b) 625 nm red, (c) 525 nm green, and (d) 425 nm blue. (C) Bacterial survival is shown in optical density (OD600). The X-axis represents irradiated time, and OD600 is shown on the Y-axis. (D) After P. gingivalis was irradiated at 48 mW/cm2 for 8 h in broth, representative photographs showing colonies were shown in CFU. (a) Control, (b) 625 nm red, (c) 525 nm green, and (d) 425 nm blue. The data are expressed as the mean±SEM and analyzed by using the two way analysis of variance (ANOVA). *p<0.05, **p<0.01, ***p<0.001.

FIG. 5.

Bacterial survival of light-emitting diode (LED) irradiation at 6 mW/cm2 for 1 h irradiation in DH5α. (A) Bacterial survival is shown in colony forming units (CFU). The X-axis represents irradiated time, and CFU are shown on the Y-axis. (B) Representative photographs showing colonies of DH5α irradiated at48 mW/cm2 for 8 h in agar. (a) Control, (b) 625 nm red, (c) 525 nm green, (d) 425 nm blue. (C) Bacterial survival is shown in optical density (OD600). The X-axis represents irradiated time, and OD600 is shown on the Y-axis. (D) After DH5α irradiated at 48 mW/cm2 for 8 h in broth, representative photographs showing colonies were shown in CFU. (a) Control, (b) 625 nm red, (c) 525 nm green, and (d) 425 nm blue. The data are expressed as the mean±SEM and analyzed by using the two way analysis of variance (ANOVA). *p<0.05, **p<0.01, ***p<0.001.

FIG. 6.

Bacterial survival of LED irradiation at 6 mW/cm2 for 1 h in Staphylococcus aureus. (A) Bacterial survival is shown in colony forming units (CFU). The X-axis represents irradiated time and CFU are shown on the Y-axis. (B) Representative photographs showing colonies of S. aureus irradiated at 48 mW/cm2 for 8 h in agar. (a) Control, (b) 625 nm red, (c) 525 nm green, and (d) 425 nm blue. (C) Bacterial survival is shown in optical density (OD600). The X-axis represents irradiated time and OD600 is shown on the Y-axis. (D) After S. aureus was irradiated at 48 mW/cm2 for 8 h in broth, representative photographs showing colonies were shown in CFU. (a) Control, (b) 625 nm red, (c) 525 nm green, and (d) 425 nm blue. The data are expressed as the mean±SEM and analyzed by using the two way analysis of variance (ANOVA). *p<0.05, **p<0.01, ***p<0.001.