22P004-O
Sterilization Treatment by Using Atmospheric Pressure Plasma Jet
大気圧プラズマジェットによる殺菌・滅菌処理
Takeshi Homma, Katsuko Koike1, Masakazu Furuta1, and Yuichiro Takemura
本間 壮, 小池佳都子1, 古田雅一1, 武村祐一朗
Graduated School of Sciences and Engineering, Kinki University
3-4-1 Kowakae, Higashiosaka City, Osaka 577-8502, Japan
近畿大学大学院エレクトロニクス系工学専攻 〒577-8502 大阪府東大阪市小若江3-4-1
1Radiation Research Center, Osaka Prefecture University
1-1 Gakuen-cho, Nakaku, Sakai, Osaka 599-8531, Japan
大阪府立大学放射線研究センター 〒599-8531 大阪府堺市中区学園町1-2
Germicidal treatments of Escherichia coli (E. coli) have been performed using an atmospheric pressure Ar
plasma jet sources. While the bactericidal effect confirmed under plasma treatment, the surface temperatureof Luria-Bertani (LB) agar was not increased and ultraviolet (UV) is low emission. Therefore, it isconsidered that ions and radical species generated by atmospheric pressure plasma jet have bactericidaleffects.

1. Introduction
Sterilization treatment is of importance toprevent the infection disease and food poisoning.
Today, it is necessary to select the sterilization
method according to microbial species or thecharacter of sterilizing objects. Therefore,
research of the new sterilization method is
attractive. The plasma sterilization also start togarner attention as a new sterilization method.1,2)
In addition, atmospheric pressure plasma isstudied for various uses these days.3,4)
We have evaluated the bactericidal effect of theplasma using E. coli as a biological indicator theindex in UV, radical species and heat by use of
atmospheric pressure plasma jet.
2. Experimental Methods

Fig.1. Schematic of the germicidal treatment by plasma jet,spectroscopic measurement, and thermometry.

A schematic of our experimental set-up is shownin Fig.1. The plasma device is an atmosphericpressure plasma jet source (Plasma-Pre-treatment
System; Plasmatreat), which consists of a conicalinner electrode, and a grounded outer electrode witha nozzle of 5 mm diameter. The inner electrode iscoupled to stepped high frequency pulse current
power supply, about 280 V, 8 A, and 16-20 kHz,through a high voltage transformer (HTR1001;Plasmatreat). The working gas is Ar, and flows
spirally at 20-50 l/min through two electrodes.
The spectroscopic diagnostic system usedconsists of a space-resolving slit of 1 mm width, acollecting lens, and spectrograph (Maya2000PRO;oceanoptics).
The
spectrographallowsmeasurement wavelength between 200 and 1000
nm. The schematic layout of the spectroscopic
diagnostic system is also shown in Fig.1.
In this research, E. coli was employed as theindicator bacterium in order to evaluate thebactericidal effect of the plasma. Indicator samples
were prepared by the following procedures. In thefirst place, 10ml LB agar were poured into glass
laboratory dishes (inside diameter: 40 mm, height:
20 mm). Then, 100 µl suspensions of overnightgrown E. coli culture (108 colony forming unit(CFU)/ml) were evenly spread over LB agar in thedishes. The dishes were exposed to the plasma withincreasing time. Temperature of the surface of theagar was monitored during the exposure using theinfrared
thermometer(THERMO-HUNTER;OPTEX). After the exposure, the exposed portionof the LB agar was excavated from the dish andcells were recovered and diluted serially withsterilized liquid. One hundred microliter of eachdilute was spread in LB agar plates and incubatedfor 24 hours at 37 °C.5)

Fig. 2 UV/visible emission spectra of Ar plasma jet
sources in the wavelength range from 250 to 1000 nm,
where distance from nozzle

Bacterial count (CFU/ml)

3. Results and Discussion.
UV and visible emission spectra are shown in
Fig.2 in the wavelength range from 200 to 1000 nm,
where the observation positions are 5, 10 and 15mm from nozzle, and the working gas was Ar. The
strong emission line of the Ar I was observed in thewavelength range from 650 to 900 nm. Moreover,
the OH A-X transition lines were found around 310
nm, therefore radical species were generated fromnot only Ar gas but atmosphere. In other words,
reactive oxygen species (•OH, •O2 and etc) may begenerated from H2O contained in the surrounding
atmosphere by plasma exposure. When theobservation position was near from nozzle, theintensity of the emission spectra was strong.
Fig.3 shows that the total counts of the surviving
bacteria on the LB agar after plasma treatment (theworking gas was Ar gas, the gas flow rate was 20l/min, the plasma exposure time was 60 seconds,
the distance from nozzle was 5 mm, 10 mm, and 15mm). As a result, significant bactericidal effect wasseen on E. coli by the plasma treatment.
Furthermore, when the distance between the nozzle
and LB agar was small, the bactericidal effect washigher. However, as shown in Fig.2. emission
intensity of UV (200-300 nm) was low. Moreover,measured temperature on the surface of the LB agar
under each plasma treatment was less than 40 °C.

910810Untreated71061051041031051015Distance from nozzle (mm)Fig.3. Total bacterial count of Escherichia coli after
plasma treatment.

For this reason, it is considered that this bactericidal
effect was not high within the range of UV emission
and heat in this expriment.6,7) As we discussed in
the results, the bactericidal effects on E. coli would
be caused by ions and radical species produced in
the plasma.
4. Conclusion
In this study, we have investigated thebactericidal effects on E. coli using atmosphericpressure plasma generated from Ar gas. The
bactericidal effects caused by the plasma would beother than UV or heat from the plasma. When thedistance from nozzle was small, ions or radicalspecies increase and the bactericidal effect washigher. It is considered that bactericidal effects inions and radical species generated by atmosphericpressure plasma jet might be more significant.
References[1] Lassen, K. S., Nordby, B., and Grun, R: J. Biomed
Mater Res Part B, Appl Biomater, 65B (2003) 239.
[2] S. Takashima, and M. Kamitani: Japanese Society ofMedical Instrumentation, 78 (2008) 49.
[3] Y. Takemura, N. Yamaguchi, and T. Hara: J. Appl.
Phys. 47 (2008) 5644.
[4] Y. Takemura, Y. Kubota, N. Yamaguchi, and T. Hara:
Ieee Transactions on Plasma Science, 37 (2009)1604.
[5] R. F. Harris, and L. E. Sommers: American Society
for Microbiology, 16 (1968) 330.
[6] S. Kinoshita, and K. Yoshino: Society for
Antibacterial and Antifungal Agents, Japan, 38(2010) 521.
[7] T. Tatsuta, and T. Ogura: Biophysical Society ofJapan, 39 (1999) 295