cold atmospheric plasma for surface disinfection

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Cold Atmospheric Plasma for SurfaceDisinfection

Yang-Fang Li,* Tetsuji Shimizu, Julia L. Zimmermann, Gregor E. Morfill

A new design evolved from the surface microdischarge electrode system has been developedfor the application of surface disinfection. Having the electrode encapsulated in a dielectricsurface the plasma is produced directly on the surface of the electrode which thereforebecomes disinfected. Experiments with bacteria werecarried out and results show that the surface can bedisinfected within 30 s with typical operating conditions.Even sterilization appears possible in a reasonable time.Possible applications of this electrode design are dis-cussed for regular disinfection of work surfaces, labora-tory benches, etc.

1. Introduction

Recent progress in the development of cold atmospheric

plasmas (CAPs) has accelerated the focus on plasma

applications in health care.[1–10] CAPs are designed to

operate at normal atmospheric temperature and pressure

in air. As shown in various studies,[11–18] CAPs have

bactericidal (including antibiotic resistant bacteria, for

example, Methicillin-resistant Staphylococcus aureus

(MRSA)) and fungicidal properties. These plasma sources

have many advantages compared with conventional

disinfection methods, e.g., low cost, simple design, and

easy handling. Moreover, recent experiments with fibrous

material[19] show that the plasma disinfection and

sterilization works at the atomic/molecular level. Therefore

the reactive plasma species can reach all surfaces and can

penetrate into small openings, including the interior of

hollow syringe which is not accessible for fluid disin-

fectants.[20] In addition, plasma can be used as a prospective

Y.-F. Li, T. Shimizu, J. L. Zimmermann, G. E. MorfillMax-Planck-Institute for Extraterrestrial Physics, 85748 Garching,GermanyFax: þ49 89 30000 3950; E-mail: [email protected]

Plasma Process. Polym. 2011, 8, 000–000

� 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim wileyonlin

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tool for medical applications instead of or together with

current standard treatments with fluids and ointments.

In health care, CAPs can be used in two fields: hygiene and

medicine application. In the field of hygiene, disinfection

and sterilization are the main issues, i.e., to kill bacteria,

fungi, viruses, and spores. Treatment with plasma allows

efficient disinfection in a very short period of time

(seconds). Therefore, the plasma technique is very promis-

ing for the containment of infectious diseases in both public

and personal areas. In the field of medicine application,

there are several potential applications. Plasma produces

biologically reactive species (of limited life time) – charged

particles, chemically reactive species and UV light – which

affect tissue. By ‘‘designing’’ these components, CAPs can in

principle be used as a therapeutic tool, e.g., for treatment of

chronic wounds.[13,21]

Our group has developed and tested a large area plasma

dispenser based on surface microdischarge (SMD) electrode

technology.[20] This technology provides a robust and

scalable design. In this paper, we describe a modified

SMD electrode which is completely encapsulated in a thin

insulating layer. Plasma is then produced on the surface of

this insulator. Such a plasma device is then water-tight,

elibrary.com DOI: 10.1002/ppap.201100090 1

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Y.-F. Li, T. Shimizu, J. L. Zimmermann, G. E. Morfill

easy to clean, and free of certain chemical contaminations

and corrosion. Using this design, objects which are not in

contact with the insulating surface as well as the insulating

surface itself can be disinfected. Such electrodes (provided it

is sufficiently large) can be used as a work surface in

laboratories, the food industry, private homes, etc.

Figure 1. (a) The sketch shows the electrode arrangement of theSSS (HV: high voltage signal). The size of the large area electrodeis 9 cm in diameter. (b) The photography shows the device withplasma ignited on the Epoxy surface. The size of the discharge isapproximately 9 cm in diameter and the Epoxy board has adimension of 12 cm� 12 cm. (c) The optical emission spectrummeasured (HAMAMATSU, TM-UV/VIS: C10082CA, MiniSpectrom-eter, 200–800 nm) for the discharge. It is from the secondpositive system of Nitrogen molecules. The main peaks arelabeled with their wavelengths.

2. The Self-Sterilizing Surface

The electrode arrangement of the self-sterilizing surface

(SSS) is based on the principle of the SMD electrode as

reported by Morfill et al.[20] The SMD electrode has a

sandwich structure which consists typically of a large-area

powered electrode, a dielectric layer, and a grounded mesh

electrode. The plasma is ignited on the side of the mesh

electrode. A printed circuit board (PCB) with laminated

copper foil on both sides is utilized in our study.

The SSS device is fabricated from a commercialized PCB.

The dielectric layer of the PCB is made of Epoxy with

dimensions of 12 cm� 12 cm and has a thickness of 1.5 mm.

The copper foils which are laminated on both sides of the

dielectric layer are 0.1 mm thick and with size of 9 cm in

diameter. The sketch in Figure 1 shows the electrode

arrangement of the SSS used in this study. Below the Epoxy

board the copper foil with a large area functions as the

powered electrode. Above the Epoxy board the copper foil is

fabricated into two electrically isolated wire electrodes, i.e.,

‘‘wire electrode 1’’ and ‘‘wire electrode 2’’ in the sketch. The

distance between two wire electrodes is 2 mm. The wire

electrodes are covered by a thin layer of glass Epoxy

(https://www.epotek.com/SSCDocs/datasheets/920-

FL.PDF) with a thickness of approximately 0.2 mm. The large

area electrode and one of the two wire electrodes are

powered. The other wire electrode is grounded. The plasma

is ignited on the surface of the glass Epoxy and the height of

the plasma glow above the Epoxy glass is <1 mm. The

device is presented as the lower image in Figure 1 when the

plasma was ignited on the Epoxy surface.

The voltage signal is provided by a Voltcraft 7202 Sweep/

Function Generator and amplified by a TREK PM04015 High

Voltage AC/DC Generator. A square waveform was used

for igniting the plasma. Compared with sinusoidal and

triangular waveforms the voltage for igniting plasma was

lower when a square waveform was used.

3. Results and Discussion

In our study, Escherichia coli (E. coli, DSM1116) were used.

Number of bacteria on agar plate, which is about 90 mm in

diameter, was estimated by counting the colony forming

units (CFUs) after 16-h incubation at a temperature of 35 8C.

A voltage signal of 2 kHz with peak-to-peak amplitude of


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18 kV was applied to the electrode. Figure 1c shows the

optical emission spectrum of the discharge measured by a

spectrometer (HAMAMATSU, TM-UV/VIS: C10082CA) with

integration time of 0.1 s. The emission is from the second

positive system of Nitrogen molecules. No detectable

signals from NO were found. The possible reason can be

that the discharge produced large amount of O3 and the

lifetime of NO was too short to be detected because of

oxidation by O3. No emission signals from the first positive

system of the Nitrogen molecules were detected. The B3Pg

state of N2 may have interacted with other species instead

of decaying to the meta-stable state. The UV emission

measured by a UV power meter (HAMAMATSU, C8026) at a

DOI: 10.1002/ppap.201100090

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Figure 3. Surviving number of CFUs on the agar samples byapplying plasma treatment with different plasma-on times (firstnumber in the horizontal coordinate) and afterglow time (secondnumber in the horizontal coordinate). The control shows thenumber of CFUs on the agar plates without applying any plasmatreatment. The numbers inserted above each column (withplasma treatment) refer to the log-reductions.

Figure 4. Survival ratio of bacteria after plasma treatment by theSSS with respect to different treatment times and for differentbacteria dilutions.

Figure 2. The voltage and the current waveforms for the dischargeare shown with light gray line and the black line, respectively, andthe power density absorbed by the discharge is about 1 W � cm�2.

The Self-Sterilizing Surface

distance of 1 cm from the discharge was approximately

0.5 mW � cm�2. The current as well as the voltage waveforms

were measured and presented in Figure 2. To get the current

waveform, a pure resistor of 3.3 V was connected to the

discharge circuit in serial and the voltage signal on this

resistor was detected with a LeCroy active voltage probe

(Model: HFP2500, Bandwidth: 2.5 GHz). Both the voltage

and the current waveforms were recorded by a LeCroy

WavePro 725Zi oscilloscope (Bandwidth: 2.5 GHz) with a

sampling rate of 20 G � s�1. The power density absorbed by

the discharge is about 1 W � cm�2. Bacterial sample was

placed upside down on the SSS. The distance between the

surface where the plasma was ignited and the bacterial

surface was 7 mm. We first did a series of experiments to

check the effect of afterglow and the result is presented in

Figure 3. Every treatment was repeated three times and also

three plates were used for the control. The error bars shown

in the plot were calculated as the standard deviations. The

ambient temperature and relative humidity for the after-

glow experiment are 24 8C and 60%, respectively and the

cell density on the agar plates before the plasma treatment

was about 3.3� 105 cm�2. The first four treatments have the

same total treatment time, including the plasma-on time

and the afterglow time. It shows that, by including 28 s of

afterglow effect, even 2 s plasma production can achieve

approximately 5-log reduction. Given the same total

treatment time, increasing the plasma-on time until 10 s

does not significantly increase the log-reduction of the

bacterial killing. In addition, when the plasma-on time for

the treatments is the same (15 s, the last two treatments in

Figure 2), including 15 s afterglow time can obviously

increase the bacterial killing rate.

A recent report showed that the bactericidal efficiency by

CAPs also depends on initial bacteria concentration of

cultured samples.[22] Therefore we performed the bacterial




experiments with different initial concentrations of E. coli

on the agar plates. In this experiment, the afterglow was not

used (The bacterial samples were immediately removed

from the SSS electrode after the plasma treatment). The

ambient temperature and relative humidity for this

experiment were 24 8C and 55%, respectively. To improve

the statistics, every treatment was repeated three times

and standard deviation of the CFUs was calculated. The

obtained results are presented in Figure 4. Before plasma

treatment, approximately 107 E. coli bacteria were cultured

on the agar surface. The survival curves for different

dilutions of this basic culture show a bi-phasic structure

similar to that found in ref. [23], although the survival ratios

of bacteria are different for a given treatment time. The first

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Figure 5. Direct bacteria inactivation experiment on the SSS usingmembrane filters. Image (a) and (b), respectively, show the CFUson the membrane filter without and with plasma treatment. Thearea covered by the CFUs (normalized by the filter area) is plottedin (c).

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Y.-F. Li, T. Shimizu, J. L. Zimmermann, G. E. Morfill

phase takes place in first 2 s where a very high rate of

bacteria inactivation is obtained. For longer treatment

times, the inactivation rate increases comparatively slowly.

For the master suspension a 4-log reduction of the bacteria

number is achieved within 2 s. This result indicates that the

decimal time (D-value, time to reduce the number of

survivors by one log) of the plasma is much <1 s. Although

not presented here, it was found all bacteria were killed

after 30 s plasma exposure. Figure 4 shows higher

inactivation rates for higher initial bacteria concentrations.

The explanation can be straightforward. Even for the

highest initial concentration, the agar surface was not

covered totally by E. coli. Given that bacteria were

homogeneously distributed on the agar surface, the

shielding and stacking effects of bacteria played no

role. The size of the SMD plasma was larger than the size

of the agar plate. The plasma treatment was applied to

the whole agar plate and each bacterium had the same

probability to receive the same dosage of plasma treatment

from the beginning of the plasma treatment. Therefore the

plasma treatment had high probability to kill more bacteria

when the initial concentration was higher. The experiment

presented in ref. [22] has a major difference from ours. In

ref. [22], a plasma jet was used and size of plasma (cross

section of the plasma plume) was much smaller than the

surface area of the bacterial samples. The plasma inactiva-

tion started from the center of the bacterial samples and

then dispersed to larger sample area when longer plasma

treatment was applied. Most of the bacteria were shielded

from the plasma treatment by the bacteria closer to the

sample center. The shielding effect played a major role and

their results showed higher bacterial inactivation rate for

lower initial bacteria concentration.

In order to test for bacteria inactivation directly on the

SSS, bacterial experiment was conducted by using mem-

brane filters: [24] In this case, the master bacteria suspension

was applied directly to the SSS. After a drying time of

10 min, the plasma was switched on for a chosen treatment

time and subsequently the bacteria on the SSS were

collected using a membrane filter (analogous to the

technique used in ref. [13]). The result is shown in

Figure 5. Without plasma treatment, more than 75% of

the filter area is covered by overlapping CFUs. After a

plasma treatment time of 30 s only isolated CFUs were

found on the filter and the CFU covered area was <10% of

the filter area. We are therefore able to show that the

bacteria concentration is significantly decreased after 30 s

of plasma exposure. This experiment was also repeated

three times. The results led to a good reproducibility and the

standard deviation is presented in Figure 5c. Although the

electrode surface was kept drying for 10 min, the humidity

in the ambient air (ambient conditions for the experiment:

relative humidity of 60% and temperature of 25 8C) was

sufficient to keep the bacteria alive at least for short period.



rly View Publication; these are NOT the fina

This was confirmed by the non-exposed sample presented

in Figure 5a.

4. Conclusion

It could be shown that a fully encapsulated SMD electrode –

the SSS – has high bactericidal effect directly on the electrode

surface. The design of the SSS can be easily improved by

optimizing the distance between the electrodes, the thick-

ness of the surface and the material for the dielectric layer.

Thus the SSS has a potentially large range of application

areas, such as in food hygiene, biological laboratories,

domestic kitchens etc. – i.e., in any surface which is

intensively used and therefore needs a regular disinfection.

Acknowledgements: This work was carried out in the frame of theplasma health care project, a collaboration initiated by the MaxPlanck Institute for Extraterrestrial Physics. We would like tothank Hans-Ulrich Schmidt for providing us the stock cultures ofE. coli and Georg Isbary for the membrane filters.

Received: May 24, 2011; Revised: October 5, 2011; Accepted:October 20, 2011; DOI: 10.1002/ppap.201100090

DOI: 10.1002/ppap.201100090

l page numbers, use DOI for citation !!

The Self-Sterilizing Surface

Keywords: atmospheric pressure glow discharge; self-steriliza-tion; surface disinfection; surface micro-discharge

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cold atmospheric plasma for surface disinfection

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