Study of Microbial Sterilization by Atmospheric Remote Microplasma
S. Tatematsu,Tatematsu, N. masamura,masamura, M. Blajan and K. Shimizu
Organization for Innovation and Social Collaboration, Shizuoka University

P-75
Jouoku,, Hamamatsu, 432Jouoku
432-8561, Japan
E-mailmail：：shimizu@cjr.shizuoka.ac.jp

Fig. 3 shows experimental setup for sterilization of E.
coli. E. coli was diluted in the liquid medium and theINTRODUCTION
An effective and economical sterilization method isrequired in the recent years. Therefore plasma
sterilization received a lot of attention andintensively studied as a replacement of traditional
sterilization
methods.
Inaddition,“Plasmamedicine“ is got a lot of attention as new application
field of plasma technology.
Microplasma is studied in our group is atmospheric
non thermal plasma which can be generated relatively
low discharge voltage. It is a kind of dielectricbarrier discharge and discharge gap is in the orderof micro meters. Remote sterilization of surfaceassociated bacteria by atmospheric microplasma wasinvestigated.

culture medium was applied on the agar medium. Thenthe agar medium was inserted in the microplasma
reactor and exposed to remote microplasma. Thedistance between agar medium and electrodes could be
adjusted by Z-axis stage.

1 µm(a) Before treatment(b) Ar microplasma 650 VFig. 8 Image of E. coli before and after Ar microplasma
treatment(x9,000,, treatment time 60 s).
treatment(x9,000RESULTS(1)Electric(1) Electric Characteristics
Discharge voltage

Discharge voltage
GNDGND

1 kV/div

1 kV/div

10 µs/div

10 µs/div
Discharge current

Discharge current
GNDGNDMETHODS

1 µm

100 mA/divmA/div

1 µm

10 µs/div

1 µm

100 mAmA/div/div
10 µs/div(1)Sterilization(1)Sterilization process and electrodes(a) Before treatment
Ar or air

Ar

O2

H2O

N2

OH*

O3O(1D)

OH*

O3

OH*

OH*

Fig. 9 Image of Alicyclobacillus before and after Ar
microplasma treatment(x30,000, treatment time 60 s).

Spike-like currents occurred at the steepest slopesof the discharge voltage. This is a typical waveform of
dielectric barrier discharge shown in Fig. 4.

Fig. 8 shows the image of E. coli which was thenursing cell state and Fig. 9 shows the image ofgram-positive bacteria A. acidoterrestris was spore
state taken by SEM. E. coli and A. acidoterrestris
were physically damaged by applying microplasma as
shown in Figs. 8 and 9 (b). This could be consideredthat active species derived from oxygen e. g. OH
radicals exert sterilization effect.

Spacer
100 µmArI(b) Ar microplasma 650 V(a) Ar microplasma(b) Air microplasma
Fig. 4 currentcurrent-voltage characteristic

N2(A)OH*(2) Sterilization result
Agar(a) Sterilization process(b) Image of microplasmaFig. 1 Sterilization process for E. coli and image ofmicroplasma during discharge.
discharge.
Fig. 1 shows the electrodes and sterilization process.
Electrodes are perforated metal plates covered with adielectric layer and faced together. Microplasma wasgenerated between the electrodes by the applied
voltage thus active species and radicals were produced
according to gas composition. Table 1 describes thedetail of electrodes.

(3) Electron spin resonance method(a) No treatment(b)Ar microplasma 850 V
Fig
Fig.. 5 The experiment result of sterilization by using
Ar microplasma. Treatment time was 60 s, distance
between electrodes and E. coli was 2 mm.
mm.

Table 1 The detail of electrodes.
Diameter
Hole diameter
Aperture ratio
Discharge gap
Discharge area
Gas flow rate

45 mm
1.6 mm30%
100 µm
100 mm2
10 L/minFig. 10 OH signal detected by the ESR measurement(treatment time 3 min.).

(2) Image of streamer of discharge

100 µmHigh voltage electrode (negative)(a) Electrode surface (b) Streamer from discharge
Fig. 2 Image of a streamer from microplasma discharge.
Microplasma is typically dielectric barrier discharge.
The image of emitting of streamer is shown in fig. 2.
Streamer diameter was thinner than previously
reported.

(3) Experimental Setup(a) No treatment(b)Air microplasma 1.5 kV
Fig
Fig.. 6 The experiment result of sterilization by using
air microplasma. Treatment time was 60 s, distance
between electrodes and E. coli was 2 mm.
Figs. 5 and 6 show the sterilization result of E. coli.
Ar microplasma treatment could sterilize near thecenter of Petri dish. However, air microplasma
treatment could sterilize more widely area than Arone. This could be due to the long lifetime activespecies derived from O2.

100
Sterilization rate [%]Grounded electrode

O·+H2Oe·-+H2O·OH+·OH→→→·OH+·OH·OH+·H+ e·H2O2

(1)
(2)
(3)

Among this active species and radicals, OH and O3
could play an important role in the sterilization
process of bacteria.

80

CONCLUSIONS
60
40

Microplasma generated at atmospheric pressure andlow discharge voltage was used for sterilization andmeasured surface potential. The following conclusions
were obtained by the series of experiment.
experiment.

Air microplasma 1.4 kVAir microplasma 1.5 kV
Air microplasma 1.6 kV

20002040
Exposure time [s]60Fig
Fig.. 7 The experiment result of sterilization rate byusing air microplasma.
microplasma. Treatment time was 0 - 60 s,distance between electrodes and E. coli was 2 mm.
mm.

Fig. 3 Experimental setup

Spin trap agent 5,5-dimethyl-1-pyrroline N-oxide
was used to perform ESR analysis. Generated OH
radical by microplasma were observed at 3445, 3460,3430, 3475 gauss.
In water these active were generated during
discharge via reactions (1), (2), and (3)[1].

Fig
Fig.. 7 shows the sterilization rate by air microplasma.
microplasma.
The culture solution of E. coli was diluted to 104 andapplied on the agar medium due to make the colonies.
colonies.
The colonies was counted after microplasma exposureand sterilization rate was calculated.
calculated.

(1)Air microplasma sterilized a wider area than Ar
microplasma
treatment.
Almost100%ofsterilization rate was completed by 60 s of air
microplasma treatment.
(2)The photos taken by SEM indicate that microplasma sterilization acts like an etching process.
(3)OH radical was one of the causes of indigo
carmine decomposition. It was dissolved in thewater as confirmed by using ESR.
[1].K Takahashi, Y Sasaki, S Mukaigawa and K Takaki
IEEE Trans. on Plasma Sci. 38 2694-2700 (2010).