Development of a new water sterilization device with a 365 nm UV-LED

3.1

The relation between the UV irradiation time and the inactivation rate

We estimated the ability of UVA-LED light to inactivate E. coli DH5α (Fig. 2). At 54 J/cm2 of UVA irradiation, the inactivation efficiency reached a maximum log10 reduction of 3.9. These data indicate that UVA-LED can inactivate bacteria in water.

UVA-LED irradiation inactivates E. coli DH5α in a UVA dose-dependent manner. The initial number of cells was 104 CFU ml−1. The log survival ratio is described in the Materials and methods. (filled circle); Consecutive irradiation of E. coli DH5α by UVA-LED. The current was set at 500 mA as described in the text. (open circle); intermittent irradiation of E. coli DH5α by UVA-LED. The bacteria were irradiated with 10 ms 1A pulses with 100 ms between each pulse (duty ratio 1/10). (filled square); Non-irradiated control samples (in the dark at 25°C). The data represent means ± SD (n = 5)

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3.2

Pulse irradiation and continuous irradiation

The sterilization rate using pulse irradiation and continuous irradiation was compared used by E. coli DH5α (Fig. 2). We irradiated using a 1 A current pulse: 10 ms ON and 100 ms OFF (duty ratio 1/10). Under continuous irradiation, the current was set to 500 mA as described in the experiment above. Applying such high currents to UV-LEDs can generate high amounts of heat, and can cause problems with the experimental device; to prevent this, pulse irradiation must be performed, particularly when using 1A currents .

The inactivation rate was 100% for an irradiation dose of 27 J/cm2 at both 500 mA continuous irradiation and 1 A pulse irradiation. At low irradiation doses, a continuous irradiation of 500 mA was sufficient to inactivate the bacteria, whereas pulse irradiation was not. However, the advantages of using pulse irradiation include the prevention of heat generation and the capability to use higher outputs of irradiation, which penetrate deeper into the sample as compared to lower outputs. Therefore, it is beneficial to use pulse irradiation for sterilization. For the remaining experiments, we performed sterilization using pulse irradiation.

3.3

Comparison of the sterilization ability by wavelength

In order to evaluate the sterilization ability of 365 nm UVA-LED, we conducted an experiment using an LED that emits light at 405 nm and a low-pressure UV lamp that emits light at 254 nm used by E. coli DH5α (Fig. 3). No sterilization effect was observed by irradiating with light at 405 nm. This suggests that the sterilization effect of the UVA-LED is due to light emitted at a wavelength of 365 nm. Indeed, similar sterilization abilities were observed with the 364 nm wavelength UVA-LED and the 254 nm wavelength UV lamp which sterilized at a significantly low energy (J/cm2). Although a UVA-LED can sterilize at an irradiation dose of 27 J/cm2, it can not sterilize at irradiation doses as small as those used by low-pressure UV lamps.

The log survival ratio depends on different wavelengths of irradiation. The initial number of E. coli DH5α was 104 CFU ml−1. Log Survival Ratio is described in the Materials and methods. A total of 405 nm wavelength of light emitted by an LED (prototype; Nichia Corporation, Japan,) and 254 nm wavelength of light emitted by a low-pressure UV lamp (3UV Multi-Wavelength Lamp, 3UV-38; UVP, Inc. CA, USA) were used to compare the sterilization ability of different wavelengths of light. White colum; 15 min exposured of each of light. Black column; 30 min exposure of each of light. The data represent means ± SD (n = 5)

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3.4

Effects of temperature and pH of the water on inactivation percentage

Next, we estimated the effect on the environmental condition, such as temperature (Fig. 4a) and pH (Fig. 4b) of the bacterial suspension, because there were reported that temperature and pH effect on sterilization [3, 4, 9]. Sterilization abilities were indicated higher at 20°C and the pH 8. Moreover, different bacteria, such as E. coli DH5α, Enteropathogenic E. coli, V. parahaemolyticus, S. aureus, and S. enterica serovar Enteritidis had each of different sensitivity for temperature and pH (data not shown), it will be necessary to consider sensitivity for each of bacteria.

Effects of the environmental condition on sterilization ability. The initial number of E. coli DH5α was 104 CFU ml−1. Log survival ratio is described in the Materials and methods. Consecutive irradiation of E. coli DH5α by UVA-LED for 15 min. The current was set at 500 mA. a Temperture dependency of sterilization ability. The equipment and the bacterial solution had been kept for 1 h prior to the experiment. The data represent means ± SD (n = 5). b pH dependency sterilization ability. The pH of PBS for making the bacterial suspension was adjusted to each of indicated pH by the NaCl or HCl. The data represent means ± SD (n = 5)

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The purpose of this research was to investigate the possibility of sterilizing by using a UVA-LED and to determine the possibility of applying UVA-LED to a sterilization device. From our results, we show that irradiating with a UVA-LED for about 30 minutes can almost completely sterilize nonpathogenic and pathogenic bacteria. It is difficult to instantly sterilize as with UV lamps, however, the bacteria were fully sterilized by a UVA-LED sterilizer after a certain time. In a water drinking guideline by World Health Organization, E. coli or thermotolerant coliform bacteria must not be detectable in any 100 ml sample in the bacteriological quality of drinking water [13]. Thus, it will be possible to use this system for drinking water sterility. It is hard to estimate “suitability” for animal or human consumption or surgery, because there are a lot of factors included for estimating the suitability, such as taste, a smell, an appearance, not only bacterial existence. Thus, we need future experiments about estimating suitability of this irradiated water for animal or human consumption or surgery.

Compared to the low-pressure UV lamp, a UVA-LED is considerably smaller and operates at a higher intensity, which would make it more useful over a broader range of applications. In addition, 365 nm UV can penetrate further than 254 nm UV. Therefore, it is conceivable that 365 nm UV is more effective than 254 nm UV for sterilizing cloudy or colored water.

Because the UVA-LED disinfection system studied here was a small, ELISA plate system, challenges will be encountered when trying to apply this system to a large volume of flowing water. Because much of the LED light leaked out of the ELISA device used this study, most of the light from the LED was not used for inactivating the bacteria. Clearly, the geometry of the light source will need to be adapted to develop an effective device for a larger-scale UVA-LED water disinfection system.

In the future, we will conduct experiments using larger volumes of water to develop UVA-LED into practical use in a circulating water system to take advantage of the safety and compact size of UVA-LED sterilization devices.