International Journal of

Molecular Sciences

Review
Disinfection and Sterilization Using Plasma
Technology: Fundamentals and Future Perspectives
for Biological Applications
Akikazu Sakudo 1, *, Yoshihito Yagyu 2 and Takashi Onodera 3
1 Faculty of Veterinary Medicine, Okayama University of Science, Imabari, Ehime 794-8555, Japan
2 Department of Electrical and Electric Engineering, National Institute of Technology Sasebo College,
Nagasaki 857-1193, Japan; yyagyu@sasebo.ac.jp
3 Research Center for Food Safety, Graduate School of Agricultural and Life Sciences, the University of Tokyo,
Bunkyo-ku, Tokyo 113-8657, Japan; takashi.onodera@riken.jp
* Correspondence: akikazusakudo@gmail.com




Received: 8 October 2019; Accepted: 21 October 2019; Published: 21 October 2019


Abstract: Recent studies have shown that plasma can efficiently inactivate microbial pathogens
such as bacteria, fungi, and viruses in addition to degrading toxins. Moreover, this technology is
effective at inactivating pathogens on the surface of medical and dental devices, as well as agricultural
products. The current practical applications of plasma technology range from sterilizing therapeutic
medical devices to improving crop yields, as well as the area of food preservation. This review
introduces recent advances and future perspectives in plasma technology, especially in applications
related to disinfection and sterilization. We also introduce the latest studies, mainly focusing on
the potential applications of plasma technology for the inactivation of microorganisms and the
degradation of toxins.

Keywords: discharge; disinfection; inactivation; sterilization; toxins

1. Introduction
Irving Langmuir first coined the term ‘plasma’ in 1927 to describe an ionized gas [1].
Early applications of plasma technology mainly focused in the field of engineering, such as nuclear
fusion and plasma etching [2–4]. However, over the past 20 years, there has been a plethora of patents
and scientific papers describing the microbicidal properties of plasma [5]. Recently accumulated
knowledge has led to improvements in the efficiency of the disinfection and sterilization using plasma
technology and a growing awareness of its potential utility [5–7].
In this review, we summarize the fundamentals of the methods for plasma generation and
their application, as well as the efficacy of these disinfection/sterilization methods against various
microorganisms. Furthermore, we also discuss the possible future application of this technology in the
area of medicine and dentistry as well as agriculture.

2. Fundamentals of Plasma and Methods for Its Generation

There are three commonly encountered states of matter: solid, liquid, and gas. When a solid is
heated, it transforms into a liquid and then from a liquid into a gas. If enough energy is applied to
gas, it becomes an ionized gas known as plasma, which represents the fourth fundamental state of
matter [8]. The plasma contains reactive chemical species such as electrons, ions, neutral molecules,
and atoms, as well as charged species [9]. In addition, the emission of radiation occurs in the ultraviolet
(UV) as well as visible and near-infrared regions during plasma generation.

Int. J. Mol. Sci. 2019, 20, 5216; doi:10.3390/ijms20205216 www.mdpi.com/journal/ijms

Int. J. Mol. Sci. 2019, 20, 5216 2 of 17

A state of plasma could be typically classified according to temperature [10–17] (Table 1). In a
high-temperature plasma, which is a strong or fully ionized plasma, the temperature of the electrons Te
and ions Tion are the same, so they are in thermal equilibrium with each other by collision due to thermal
motion. The gas temperature Tgas of high-temperature plasma and thermal plasma is too extreme for
treating living organisms. Alternatively, in non-thermal plasma, comprising partially ionized plasmas,
the temperature of the electrons Te is much higher than that of the ions Tion and neutrons Tn . The energy
transfer of the kinematics of a collision between electrons (light particles) and ions or neutrals (heavy
particles) tends to be very slow by elastic collision, but electron-electron collisions readily achieve
thermodynamic equilibrium. Therefore, the ionized gas temperature keeps the normally ambient
temperature in non-thermal plasma. As a result, the gas temperature of non-thermal plasma remains
low, making it suitable for biological applications.

Table 1. Typical classification of plasma [10,17].

Electron Density
Classification Temperature [K] Discharge Type Examples
[m−3 ]
High-temperature
Te ≈ Tion ≈ Tgas =
plasma ne ≥ 1020 Laser fusion Tokamak Fusion plasma for energy
106 –108
(Equilibrium plasma)
Arc plasma, Plasma torch,
Thermal plasma Radiation, welding and
Te ≈ Tion ≈ Tn ≈ Radio-frequency (RF)
(Quasi-equilibrium ne ≥ 1020 cutting, Waste treatment,
Tgas ≤ 2 × 104 Plasma, Microwave
plasma) Material processing, etc.
plasma etc.
Ozonizer, Plasma
medicine, Volatile
Glow discharge, Corona
organic compound
discharge, atmospheric
(VOC) treatment, Plasma
pressure plasma jet (APPJ),
Non-thermal plasma agriculture, Surface
Te ≥ Tion ≥ Tn ≈ dielectric barrier discharge
(Non-equilibrium ne ≈ 1010 modifications (coating,
Tgas = 300–1000 (DBD), micro-hollow
plasma) etching, activation,
cathode discharge
cleaning, nitration, etc.),
(MHCD), Plasma needle,
Illumination (plasma
Low-pressure plasma etc.
screen, fluorescent
lamps, etc.)
Te = electron temperature, Tion = ion temperature, Tgas = gas temperature, ne = electron density.

Electrical discharge methods commonly utilized for non-thermal plasma generation in biological
applications are generally categorized into one of the following: glow discharge, corona discharge,
atmospheric pressure plasma jet (APPJ), dielectric barrier discharge (DBD), micro-hollow cathode
discharge (MHCD), DC discharge, pulse discharge, or high/low-frequency discharge (Table 2). The type
of discharge depends on the frequency of the power source, such as direct current (DC) and alternating
current (AC) discharge, as well as ambient gas pressure, such as low-pressure and atmospheric pressure
plasma, and the precise shape and configuration of the electrodes [9]. In addition, the waveform may
also affect the type of discharge. Different types of plasma can be used in various biological fields,
including disinfection/sterilization.
Int. J. Mol. Sci. 2019, 20, 5216 3 of 17

Table 2. Various types of electrical discharge methods for non-thermal plasma generation.

Representative
Discharge Type * Conditions (V, A, Freq, Pressure Gas Temperature Application References
Gas)
5–30 kV direct current
(DC) (positive and
Direct current (DC) Biomedical
negative); 10–250 µA; 1 atm Room temperature [18]
corona discharge applications
dry or wet; O2 , N2 , Ar,
He at 10 L/min
Max. 50.8 ◦ C on a
Atmospheric P = 2.5 W; 2.45 GHz;
dentin surface; Biomedical
pressure plasma jet He/O2 /N2 at 2.0/1.2/1.5 1 atm [19]
20 ◦ C on an agar applications
(APPJ) microwave L/min
surface
Dielectric barrier
±2.5 kV; 5 kHz; air, Approximately Biomedical
discharge (DBD) 1 atm [20]
humidity 64.4% 50 ◦ C applications
(Flexible sheet-type)
Room temperature
Micro-hollow (220 mL/min);
1.5–2.5 kV DC; 20 mA; Medical
cathode discharge >1 atm >55 ◦ C (5 mm from [21]
air (0.1–8 L/min) applications
(MHCD) jet nozzle,
220 mL/min)
Pin-to-hole spark 9030 ± 320 K (by Medical
4 kV DC; average ~1.8
discharge (PHD) 1 atm Boltzmann applications [22]
J/pulse
plasma calculation) (wound healing)
* These types of electrical discharge are for atmospheric pressure plasmas.

Non-thermal plasma is easy to obtain under low-pressure conditions because the collisions between
electrons, ions, and neutral molecules occur infrequently. Low-pressure plasma can be generated by a
low breakdown voltage in a vacuum chamber evacuated with a vacuum pump. Low-pressure plasma
systems are important for the manufacture of semiconductor components. Furthermore, research into
low-pressure plasma systems has also focused on the decontamination and sterilization of medical
devices [23,24]. Although low pressure plasma can generate high concentrations of active species
with a uniform glow plasma, it involves high maintenance costs because of the requirement for a
vacuum system. Atmospheric pressure plasma requires a high voltage and relatively high temperature
due to frequent collisions between electrons and ions accompanying the high particle density [9].
However, it is possible to generate plasma under non-thermal conditions by using a pulse discharge
and APPJ [25], DBD [26], and floating electrode barrier discharge (FE-DBD) [27,28], or MHCD [21,29].
These non-thermal conditions allow applications involving exposure of the plasma with tissues such
as skin [30].
Similarly, non-thermal plasma can be used to disinfect agricultural products and medical devices
with relatively little impact on their structural integrity [31,32]. Alternatively, the plasma could
be transferred to a target site where the object for treatment is located using a plasma afterglow
(Figure 1). Indirect treatment using solutions treated with plasma, known as “plasma-activated
water (PAW)” [33], “plasma-activated medium (PAM)” [34], “plasma-stimulated medium (PSM)” [35],
“plasma-treated water (PTW)” [36,37], “plasma-treated phosphate-buffered saline (pPBS)” [38], or
“non-thermal plasma-conditioned media (NTP media)” [39], is also possible. The constituents in these
solutions react with samples and act as disinfectants [40] or anti-cancer agents [41]. In cases where
samples are in contact with plasma bulk in the discharging area, plasma components such as UV
radiation and reactive chemical species directly interact with the samples. Thus, short-life reactive
chemical species, such as reactive oxygen species (ROS) and reactive nitrogen species (RNS), efficiently
interact with the sample components [42]. By contrast, in cases where the sample is in contact with
plasma bulk away from the discharging area, the contribution of UV radiation is significantly lower.
In addition, there is a greatly reduced concentration of reactive chemical species in the post discharging
 Int. J. Mol. Sci. 2019, 20, x FOR PEER REVIEW 4 of 17

UV radiation and reactive chemical species directly interact with the samples. Thus, short-life reactive
chemical species, such as reactive oxygen species (ROS) and reactive nitrogen species (RNS),
efficiently interact with the sample components [42]. By contrast, in cases where the sample is in
Int. J. Mol. Sci. 2019,
contact with20,plasma
5216 bulk away from the discharging area, the contribution of UV radiation is4 of 17
significantly lower. In addition, there is a greatly reduced concentration of reactive chemical species
in the post discharging area due to their short half-life at ambient temperature. In the case of plasma-
area due to their
treated short half-life
solutions, freezing at
canambient temperature.
extend the Inand
storage time the case of plasma-treated
minimize solutions,
the loss of reactive freezing
chemical
can extend the
species [37].storage time and minimize the loss of reactive chemical species [37].

Figure
Figure 1. Types
1. Types of plasma
of plasma treatment
treatment can canbe
beclassified
classified as
as follows.
follows.(a)(a)Discharging
Discharging areaarea
is iniscontact
in contact
withsample,
with the the sample, or (b)
or (b) thethe sample
sample is isinincontact
contact with
with plasma
plasma transferred
transferred to to
thethe
target site site
target fromfrom
the the
discharging area. Alternatively, (c) solutions previously subjected to plasma treatment could be used
discharging area. Alternatively, (c) solutions previously subjected to plasma treatment could be used as
as reagents to apply to samples. Photographic images of the different modes of treatment are shown
reagents to apply to samples. Photographic images of the different modes of treatment are shown in
in the upper section with corresponding illustrations in the lower section.
the upper section with corresponding illustrations in the lower section.
3. Inactivation of Microorganisms by Plasma
3. Inactivation of Microorganisms by Plasma
Some microorganisms such as bacteria, viruses, and fungi act as pathogens and cause diseases.
Some microorganisms such as bacteria, viruses, and fungi act as pathogens and cause diseases.
There is a resistance hierarchy of microorganisms against disinfection/sterilization that can be
Theredivided
is a resistance
into the hierarchy of microorganisms
following five categories: most against
resistant,disinfection/sterilization that can
highly resistant, intermediately be divided
resistant,
into the
lessfollowing five very
resistant, and categories: most
susceptible resistant,
[43]. The mosthighly resistant,
resistant intermediately
infectious resistant,
agents are prions less resistant,
(proteinaceous
and very susceptible
infectious [43].
particles), Theare
which most
the resistant infectious
causative agents agents
of prion are prions
diseases, such as(proteinaceous infectious
the Creutzfeldt–Jakob
particles), which are the causative agents of prion diseases, such as the Creutzfeldt–Jakob disease (CJD)
disease (CJD) in humans, bovine spongiform encephalopathy (BSE) in cattle, and chronic wasting
diseasebovine
in humans, (CWD)spongiform
in cervids (deer family). Bacterial
encephalopathy spores,
(BSE) protozoan
in cattle, oocysts,wasting
and chronic and helminth
diseaseeggs are in
(CWD)
categorized
cervids as highly
(deer family). resistant
Bacterial microorganisms.
spores, Intermediately
protozoan oocysts, resistant
and helminth microorganisms
eggs are categorized include
as highly
mycobacteria,
resistant protozoan
microorganisms. cysts, small non-enveloped
Intermediately viruses, and fungal
resistant microorganisms spores.
include Vegetative bacteria,
mycobacteria, protozoan
protozoa, helminths, fungi, and algae, as well as large non-enveloped viruses are less resistant.
cysts, small non-enveloped viruses, and fungal spores. Vegetative bacteria, protozoa, helminths, fungi,
Enveloped viruses such as human immunodeficiency virus (HIV) are generally highly susceptible to
and algae, as well as large non-enveloped viruses are less resistant. Enveloped viruses such as human
various disinfectants.
immunodeficiency
However, in virus (HIV)
reality, theare generally
situation highly
is more susceptible
complex. to various
For example, disinfectants.
although the enveloped virus
However, in reality, the situation is more complex. For example, although
HIV is very sensitive to disinfection and is rapidly inactivated even at room temperature the enveloped
without anyvirus
HIV is very sensitive
treatment, to disinfection
the influenza and isenveloped
virus (another rapidly inactivated even atinfectious
virus) can remain room temperature
for up to 48without
h.
any treatment, the influenza
Orthopoxviruses virus vaccinia)
(e.g., smallpox, (anotherand enveloped virus)
filoviruses (e.g., can remain
Ebola virus, infectious for up
Marburg virus) are to
all 48 h.
enveloped viruses
Orthopoxviruses (e.g.,that remain infectious
smallpox, vaccinia)for up filoviruses
and to several weeks.
(e.g.,Furthermore,
Ebola virus,susceptibility
Marburg virus)againstare all
enveloped viruses that remain infectious for up to several weeks. Furthermore, susceptibility against
biocides depends on the environment where the microorganisms are present (e.g., blood, serum, spinal
fluid, saliva). Certain soils can prevent drying and stabilize the viral structure, extending the survival
time of viruses. For example, viruses normally interact with external materialss including proteins,
lipids, salts, and cell debris. In other cases, viruses cause the aggregation of host cells [44] or make
aggregates themselves [45], which could reduce the virucidal effect of disinfectants. Therefore, the
susceptibility of microorganisms to disinfection/sterilization should be examined under a range of
different conditions.
 Int. J. Mol. Sci. 2019, 20, x FOR PEER REVIEW 5 of 17

biocides depends on the environment where the microorganisms are present (e.g., blood, serum,
spinal fluid, saliva). Certain soils can prevent drying and stabilize the viral structure, extending the
Int. J. Mol. Sci. 2019, 20, 5216 5 of 17
survival time of viruses. For example, viruses normally interact with external materialss including
proteins, lipids, salts, and cell debris. In other cases, viruses cause the aggregation of host cells [44]
or make aggregates themselves [45], which could reduce the virucidal effect of disinfectants.
The most impressive results in terms of microbicidal activity have come from studies using bacteria,
Therefore, the susceptibility of microorganisms to disinfection/sterilization should be examined
including bacterial spores [4–49]. Although some bacteria can form biofilms in certain environments,
under a range of different conditions.
this doesThe not most
prevent their successful inactivation by plasma treatment [50,51]. Furthermore, the
impressive results in terms of microbicidal activity have come from studies using
recent emergence of drug-resistant
bacteria, including bacterial spores pathogens is now asome
[4–49]. Although serious public
bacteria canhealth concern in
form biofilms that has been
certain
acknowledged by both the World Health Organization (WHO) [52] and the
environments, this does not prevent their successful inactivation by plasma treatment [50,51]. US Centers for Disease
Control Furthermore,
(CDC) [53]. Theemergence
the recent extensiveofand indiscriminate
drug-resistant pathogensuseis of
now antibiotics mayhealth
a serious public haveconcern
altered the
that has been
environmental acknowledged
microbiome, by both thetoWorld
contributing Health Organization
the emergence (WHO)bacteria
of drug-resistant [52] and [54].
the US Centers
Consequently,
therefor Disease
is an Control
urgent (CDC) [53].
requirement The extensive
to devise and indiscriminate
novel methods to eliminateuse these
of antibiotics may have altered
multidrug-resistant bacteria
from the food production process. Plasma treatment is an especially promising bacteria
the environmental microbiome, contributing to the emergence of drug-resistant method[54]. because
Consequently, there is an urgent requirement to devise novel methods to eliminate these multidrug-
the mechanism of bactericidal action is unlikely to differ between multidrug-resistant and normal
resistant bacteria from the food production process. Plasma treatment is an especially promising
bacteria. The main mechanisms of bactericidal action in plasma are thought to involve exposure
method because the mechanism of bactericidal action is unlikely to differ between multidrug-
to reactive
resistantchemical
and normal species for which
bacteria. The main multidrug-resistant bacteria
mechanisms of bactericidal are unlikely
action in plasmato arebethought
resistantto [55].
Furthermore, plasma pre-treatment enhances the sensitivity of methicillin-resistant
involve exposure to reactive chemical species for which multidrug-resistant bacteria are unlikely to Staphylococcus
aureus beto antibiotics
resistant [55]. [56].
Furthermore, plasma pre-treatment enhances the sensitivity of methicillin-resistant
Several reports
Staphylococcus suggest
aureus that plasma
to antibiotics [56]. can be effective in inactivating fungi [57–59]. However,
our own investigations have shown thatcan
Several reports suggest that plasma thebeviable
effective
cellinnumber
inactivating fungi [57–59].
of Aspergillus However, our
brasiliensis was not
own investigations have shown that the viable cell number of
significantly impacted after 5 min plasma treatment using a nitrogen gas plasma device, Aspergillus brasiliensis was BLP-TES
not
significantly impacted after 5 min plasma treatment using a nitrogen gas plasma device,
(bi-polar and low-pressure plasma-triple effects sterilization) [57], whereas Salmonella enterica serovar BLP-TES (bi-
polar and low-pressure plasma-triple effects sterilization) [57], whereas Salmonella enterica serovar
Abony was completely inactivated after employing the same treatment regime [55] (Figure 2). Indeed,
Abony was completely inactivated after employing the same treatment regime [55] (Figure 2). Indeed,
a 15 min treatment with nitrogen gas plasma was required to reduce the viability of A. brasiliensis.
a 15 min treatment with nitrogen gas plasma was required to reduce the viability of A. brasiliensis.
Thus,Thus,by comparison
by comparison to bacteria, extended
to bacteria, extendedtreatment timewith
treatment time withnitrogen
nitrogen gasgas plasma
plasma must must be used
be used to to
inactivate fungi.
inactivate fungi.

Figure
Figure 2. Direct
2. Direct plasma plasma treatment
treatment inactivates
inactivates bacteria
bacteria andand fungi.
fungi. (a) (a) A sample
A sample of Salmonella
of Salmonella was was
treated
treated with plasma using the BLP-TES (bi-polar and low-pressure plasma-triple effects
with plasma using the BLP-TES (bi-polar and low-pressure plasma-triple effects sterilization) device, sterilization)
device, which generates nitrogen gas plasma using a fast high-voltage pulse by a static induction (SI)
which generates nitrogen gas plasma using a fast high-voltage pulse by a static induction (SI) thyristor
thyristor power supply, for the indicated time. Colony-forming units (CFU) per ml of culture reduced
power supply, for the indicated time. Colony-forming units (CFU) per ml of culture reduced with the
with the plasma treatment in a time-dependent manner. (b) Viable cell numbers of Aspergillus reduced
plasma treatment in a time-dependent manner. (b) Viable cell numbers of Aspergillus reduced after
after plasma treatment using the BLP-TES device. Differences where * p < 0.05 and ** p < 0.01 versus
plasma treatment
control (0 min)using
were the BLP-TES
considered device. Differences
significant. where
Modified from * p <et0.05
Maeda and[55]
al. 2015 < 0.01
** pand versus
Sakudo control
et al.
(0 min) were considered significant. Modified
2017 [57] with permission from Elsevier. from Maeda et al. 2015 [55] and Sakudo et al. 2017 [57]
with permission from Elsevier.
The resistance of fungi to plasma treatment has also been studied. Soušková et al. reported the
The resistance
susceptibility of offungi
fungiis to plasmabetween
different treatment has also
species, been studied.
including AspergillusSoušková et al. reported
oryzae, Cladosporium
the susceptibility of fungi is different between species, including Aspergillus oryzae, Cladosporium
sphaerospermum, and Penicillium crustosum despite no significant differences in susceptibility against
plasma generated by corona discharge among bacteria, including Escherichia coli and Staphylococcus
epidermidis [58–60]. Among these fungi, Aspergillus displayed the greatest resistance to plasma
inactivation, possibly due to the presence of spores. Therefore, the resistance of fungi to plasma
treatment appears to be related to spore generation.
Several studies have investigated the effect of plasma on the inactivation of both enveloped and
non-enveloped viruses. Representative studies showed that nitrogen plasma generated by BLP-TES
 sphaerospermum, and Penicillium crustosum despite no significant differences in susceptibility against
plasma generated by corona discharge among bacteria, including Escherichia coli and Staphylococcus
epidermidis [58–60]. Among these fungi, Aspergillus displayed the greatest resistance to plasma
inactivation, possibly due to the presence of spores. Therefore, the resistance of fungi to plasma
Int. J. Mol. Sci. 2019,
treatment 20, 5216to be related to spore generation.
appears 6 of 17
Several studies have investigated the effect of plasma on the inactivation of both enveloped and
non-enveloped viruses. Representative studies showed that nitrogen plasma generated by BLP-TES
inactivated enveloped viruses, such as the influenza virus [61] and respiratory syncytial virus (RSV) [42],
inactivated enveloped viruses, such as the influenza virus [61] and respiratory syncytial virus (RSV)
as well as as
[42], non-enveloped viruses, viruses,
well as non-enveloped such assuch
the adenovirus [62] (Figure
as the adenovirus 3). In3).addition,
[62] (Figure there
In addition, areare
there several
studies usingstudies
several bacteriophages as model as
using bacteriophages objects
modelofobjects
viral of
inactivation by plasma
viral inactivation [63–65].
by plasma [63–65].

Figure
Figure 3. Direct
3. Direct plasmaplasma treatment
treatment inactivates
inactivates viruses.
viruses. (a) Viral
(a) Viral titers titers of median
of median tissue tissue
cultureculture
infectious
infectious dose (TCID ) per ml were calculated in duplicate (No 1 and No 2) for
dose (TCID50 ) per ml were calculated in duplicate (No 1 and No 2) for the influenza virus. The
50 the influenza virus.
TCID50
The
values TCID50 values
reduced reduced after
after treatment treatment
with with low-pressure
low-pressure nitrogennitrogen
gas plasmagas plasma
usingusing the BLP-TES
the BLP-TES device.
device. (b) Nitrogen gas plasma treatment with the BLP-TES device resulted in a decrease in viral titer
(b) Nitrogen gas plasma treatment with the BLP-TES device resulted in a decrease in viral titer [plaque
[plaque forming units (PFU) per ml] of adenovirus. Differences where * p < 0.05 versus control (0 min)
forming units (PFU) per ml] of adenovirus. Differences where * p < 0.05 versus control (0 min) were
were considered significant. Cited from Sakudo et al. 2013 [61] and Sakudo, Toyokawa, and Imanishi
considered significant. Cited from Sakudo et al. 2013 [61] and Sakudo, Toyokawa, and Imanishi
2016 [62] under the terms of the Creative Commons Attribution license.
2016 [62] under the terms of the Creative Commons Attribution license.
Furthermore, a DBD plasma torch inactivated the non-enveloped virus, feline calicivirus [66].
Furthermore, a DBD was
Inactivation of viruses plasma torchbyinactivated
achieved the non-enveloped
a relatively short exposure to plasma.virus, feline calicivirus
According to the U.S. [66].
Inactivation of viruses was achieved by a relatively short exposure
Environmental Protection Agency (USEPA) “Guide Standard and Protocol for Testingto plasma. According to the U.S.
Environmental Protection Agency (USEPA) “Guide Standard and Protocol for Testing
Microbiological Water Purifiers,” the minimum performance standards of the inactivation efficiency Microbiological
Waterare a six-log reduction/inactivation
Purifiers,” the minimum performance of bacteria, or a four-log
standards reduction/inactivation
of the inactivation efficiencyof viruses
are a[67].
six-log
Treatment using nitrogen
reduction/inactivation gas or
of bacteria, plasma generated
a four-log by BLP-TES showed
reduction/inactivation an approximate
of viruses two-logusing
[67]. Treatment
reduction
nitrogen in influenza
gas plasma virus titer
generated by after 1 min showed
BLP-TES and four-log reduction of virus
an approximate titer ofreduction
two-log adenovirusinwithin
influenza
4 min [61,62]. A 1-min treatment with the DBD plasma torch resulted in a greater than two-log
virus titer after 1 min and four-log reduction of virus titer of adenovirus within 4 min [61,62]. A 1-min
reduction of virus titer for feline calicivirus [66]. Lengthening the treatment to 2 min reduced the viral
treatment with the DBD plasma torch resulted in a greater than two-log reduction of virus titer for feline
titer to an undetectable level (3.81 × 104 ± 1.58 × 103 median tissue culture infectious dose (TCID50) /mL
calicivirus [66]. Lengthening the treatment to 2 min reduced the viral titer to an undetectable level
at 0 min; below the detection limit at 2 min). These results suggest that the 2 min treatment meets the
(3.81 × 10 4 ± 1.58 × 103 median tissue culture infectious dose (TCID )/
performance standards set by USEPA as outlined earlier. However,50asmL far at
as0the
min; beloware
authors the detection
aware,
limit at
there are a limited number of studies on the inactivation of plant viruses using plasma [68].set by
2 min). These results suggest that the 2 min treatment meets the performance standards
USEPA as outlinedthere
Furthermore, earlier. However,
are no published as far as the
studies on authors are aware,
the plasma thereofare
inactivation a limited
viroid’s, number
which are of
infectious
studies on the RNAs that cause
inactivation plant diseases.
of plant Indeed,
viruses using to develop
plasma [68].aFurthermore,
plasma disinfection
theresystem
are nofor the
published
agricultural
studies sector, it
on the plasma would be necessary
inactivation to determine
of viroid’s, which arethe effectiveness of thisthat
infectious RNAs technology on a range
cause plant diseases.
of plant pathogens.
Indeed, to develop a plasma disinfection system for the agricultural sector, it would be necessary to
determinePlasma is also effective against other microorganisms besides the ones mentioned above. For
the effectiveness of this technology on a range of plant pathogens.
example, plasma inactivation of the yeast-like algae Prototheca zopfii [69] and water-borne helminth
Plasma is also effective against other microorganisms besides the ones mentioned above.
Schistosoma japonicum [70], Acanthamoeba species (spp.), and other ocular pathogens, as well as water-
For example, plasma inactivation of the yeast-like algae Prototheca zopfii [69] and water-borne helminth
borne protozoan enteroparasite Cryptosporidium parvum when combined with pulsed UV [71] was
Schistosoma japonicum [70], Acanthamoeba species (spp.), and other ocular pathogens, as well as
water-borne protozoan enteroparasite Cryptosporidium parvum when combined with pulsed UV [71]
was confirmed. These results suggest that plasma has the potential to inactivate cysts and protozoal
oocysts as well as trophozoites of protozoon parasites. Although prions are known to be the most
resistant pathogens, they are nonetheless inactivated by radio-frequency (RF) plasma treatment using
an Ar/O2 gas mixture [72] and by plasma from a microwave discharge [73]. In addition, plasma
treatment can efficiently degrade toxins produced by both bacteria and fungi (Figure 4).
 Int. J. Mol. Sci. 2019, 20, x FOR PEER REVIEW 7 of 17

confirmed. These results suggest that plasma has the potential to inactivate cysts and protozoal
oocysts as well as trophozoites of protozoon parasites. Although prions are known to be the most
resistant pathogens, they are nonetheless inactivated by radio-frequency (RF) plasma treatment using
an Ar/O
Int. J. Mol. 2 gas
Sci. 2019, 20,mixture
5216 [72] and by plasma from a microwave discharge [73]. In addition, plasma7 of 17
treatment can efficiently degrade toxins produced by both bacteria and fungi (Figure 4).

Figure 4. Direct plasma treatment inactivates toxins. Quantitative measurement of aflatoxin B1 (AFB1)
Figure 4. Direct plasma treatment inactivates toxins. Quantitative measurement of aflatoxin B1 (AFB1)
(a,b), Shiga toxin 1 (Stx1) (c), Shiga toxin 2 (Stx2) (D) after low-pressure nitrogen gas plasma treatment
(a,b), Shiga toxin 1 (Stx1) (c), Shiga toxin 2 (Stx2) (D) after low-pressure nitrogen gas plasma treatment
with BLP-TES
with BLP-TES at 1.5 at
kpps1.5 for
kppsthefor
indicated times times
the indicated (a,c,d)(a,c,d)
and atand 0–1.5
at kpps
0–1.5 for 15 for
kpps min15(b)min
was(b)performed
was
by anperformed
enzyme-linked by an immunosorbent assay (ELISA)assay
enzyme-linked immunosorbent using an MytoJudge
(ELISA) Total Aflatoxin
using an MytoJudge Totalkit (NH Foods
Aflatoxin
Ltd.) kit
and(NHa RIDASCREEN ® ** p < 0.01
Foods Ltd.) and aVerotoxin
RIDASCREEN kit (R-Biopharm
® Verotoxin kitAG, Darmstadt).
(R-Biopharm AG,Differences
Darmstadt). where
Differences
versus control
where ** p (0 min)
< 0.01 werecontrol
versus considered
(0 min)significant.
were considered(a) and (b) are(a)
significant. cited
andfrom Sakudo
(b) are et al.
cited from [57] with
Sakudo
et al. [57]
permission withElsevier,
from permission from(c)
while Elsevier,
and (d) while
are (c) andfrom
cited (d) are cited from
Sakudo Sakudo
et al. et al. [74]
[74] under theunder
termsthe of the
terms of the Creative Commons Attribution
Creative Commons Attribution 4.0 International license. 4.0 International license.

Thus,Thus,
plasmaplasma
hashas applications
applications notonly
not onlyfor
fordisinfection/sterilization
disinfection/sterilization but butalso for for
also the the
degradation
degradation
of toxins [75]. This includes not only exotoxins but also endotoxins, lipopolysaccharides
of toxins [75]. This includes not only exotoxins but also endotoxins, lipopolysaccharides (LPS) by
inactivating their lipid A [76]. Consequently, plasma technology is a novel advanced
(LPS) by inactivating their lipid A [76]. Consequently, plasma technology is a novel
disinfection/sterilization system that can simultaneously inactivate pathogens and their associated
advanced disinfection/sterilization system that can simultaneously inactivate pathogens and their
toxins.
associatedThetoxins.
inactivation mechanisms of action of plasma remain to be determined [5,6,31,77]. The
The
mechanisms dependmechanisms
inactivation on the types ofofgases
action
usedof plasma the
to generate remain
plasma. toInbe
thedetermined
case of nitrogen[5,6,31,77].
gas
The mechanisms depend
plasma, at least threeon the types
major of gases(reactive
mechanisms used tochemical
generatespecies,
the plasma. In the case
UV radiation of nitrogen
exposure, and gas
plasma, at least
electric three
fields) major
are mechanisms
thought (reactive
to be involved [31].chemical species,
In addition, UV radiation
etching effects may exposure, and electric
also contribute,
especially in the case of oxygen plasmas [78]. Specifically, shrinking of bacterial spores
fields) are thought to be involved [31]. In addition, etching effects may also contribute, especially in were observed
in oxygen
the case gas plasma-treated
of oxygen plasmas [78].spores but not shrinking
Specifically, in nitrogen ofgasbacterial
plasma-treated
sporesspores
were [79–81].
observed in oxygen
Overall, reactive chemical species, UV radiation, and
gas plasma-treated spores but not in nitrogen gas plasma-treated spores [79–81]. electric fields contribute to the
antimicrobial effects of plasma, depending on the type of gases as well as the methods employed to
Overall, reactive chemical species, UV radiation, and electric fields contribute to the antimicrobial
generate the plasma. Reactive chemical species seem to be the principal inactivation factor in most
effects of plasma, depending on the type of gases as well as the methods employed to generate the
plasma. Reactive chemical species seem to be the principal inactivation factor in most cases, although
this may vary depending on the method of plasma generation and whether the sample is exposed to
direct or indirect plasma treatment.
In addition, inactivation mechanisms may vary depending on the target microorganism. However,
it should be noted that most studies on the inactivation mechanisms of plasma have been conducted
using bacterial spores. Therefore, further studies are required on the inactivation of various
microorganisms using plasma to understand the underlying mechanisms involved fully.
Int. J. Mol. Sci. 2019, 20, 5216 8 of 17

4. Future Perspectives in Agriculture and the Food Industry

There are extensive applications of plasma technology in the field of agriculture and in the food
sector. For example, plasma technology could be applied to the disinfection of foods, packaging
materials and equipment as well as agricultural sources such as seeds, fertilizer, water, and soil.
Agricultural products, such as fruits and vegetables, are prone to contamination from agricultural
sources, including seeds, fertilizers, water, and soil. Moreover, the agricultural products come into
contact with dust, insects, animal feces, field workers, and equipment during pre-harvest/harvest,
transport, packaging and food processing stages of the supply chain. These individual risk factors
may contribute to a microbial hazard. The application of innovative disinfection techniques, including
plasma technology, may help reduce the potential risk from these factors. A recent review has shown
that food products subjected to plasma disinfection are becoming widespread, which include fresh
fruits, vegetables, dry fruits, nuts, seeds, spices, eggshells, as well as protein products such as meat
and cold cuts [82]. However, further advances in plasma disinfection technology are required before
this method can be applied to the food industry (e.g., food processing and distribution system as well
as agricultural products).
We anticipate the wide-ranging application of plasma technology in the field of agriculture once
the equipment has been fully developed. The current priority is the development of an efficient
open-air system suitable for disinfecting both large objects and high numbers of samples. Moreover,
such a device will be readily scalable. Based on this background, we recently developed a novel roller
conveyer plasma device (Figure 5).
Int. J. Mol. Sci. 2019, 20, x FOR PEER REVIEW 9 of 17

Figure 5. (a) Schematic representation of a roller conveyer plasma device for the continuous
Figure 5. (a) Schematic representation of a roller conveyer plasma device for the continuous disinfection
disinfection of fruits and vegetables using atmospheric pressure plasma. As an example, oranges are
of fruits and vegetables using atmospheric pressure plasma. As an example, oranges are shown.
shown. (b) Oranges on rollers during operation of the device. (c) Enlarged image of (b) showing the
(b) Oranges on rollers during operation of the device. (c) Enlarged image of (b) showing the plasma
plasma (Arrow) generated between the orange and roller during operation of the device. The image
(Arrow) generated
is modified frombetween
Toyokawa theetorange
al. 2017and
[83]roller during operation
with permission of the device. The image is modified
from Elsevier.
from Toyokawa et al. 2017 [83] with permission from Elsevier.
In addition, a critical factor to consider is the safe application of this novel technology. The
This plasma
European device isstated
Commission well suited to the
that there are disinfection
no restrictionsofinfruits and vegetables
the regulations during
regarding sorting
the use of on
rollers [83]. as
plasma Theanapparatus is an original
electronic preservative designfor
practice that generates
organic foodsatmospheric
[84]. However, plasma
plasmaby the mechanism
treatment of
aqueous
of DBD. Thissolutions
unique canplasmapotentially generate
apparatus hydrogen peroxide,
is composed of rollingnitrates, and nitrites
electrodes [85,86].
comprising These rod
a plastic
(diameter = 30 mm) covered with a thin aluminum and silicon sheet positioned at an interval of 50
compounds might react to form other toxic compounds such as peroxynitrous acid. Therefore, a mm
comprehensive evaluation of the effect of plasma on foods and human health is
between the high -voltage electrode and earth electrode. The high-voltage electrode is then connectednecessary before this
to annew disinfection technology can be fully utilized. Indeed, a range of applications of plasma
alternating power supply. Plasma is generated in the silicon sheet when electrically conductive
technology in the field of agriculture is currently being assessed.
samples, such as fruits and vegetables as well as metals, make contact with both the high-voltage
Recently, the use of plasma technology has been reported to enhance seed germination and the
growth of plants [87]. In addition, the removal of volatile organic compounds, such as ethylene gas,
by plasma treatment may be useful during the transportation of agricultural products in containers
[88]. Therefore, the application of plasma technology could also contribute to higher crop yields as
well as the preservation of foods.

5. Future Perspectives in Medicine and Dentistry

Int. J. Mol. Sci. 2019, 20, 5216 9 of 17

electrode and earth electrode. Our findings suggest the device could have practical applications for the
disinfection of agricultural products during the sorting process on rollers. Disinfection of Xanthomonas
campestris p.v. campestris-contaminated cabbage leaves using the roller conveyer plasma device has been
achieved. In addition, our preliminary study has shown that the surface of Penicillium-contaminated
oranges could be disinfected using the device (Sakudo and Yagyu, unpublished results). Nonetheless,
to achieve broad applicability with a variety of agricultural food products, the device needs to be
scaled-up and its performance fully evaluated.
In addition, a critical factor to consider is the safe application of this novel technology. The European
Commission stated that there are no restrictions in the regulations regarding the use of plasma as
an electronic preservative practice for organic foods [84]. However, plasma treatment of aqueous
solutions can potentially generate hydrogen peroxide, nitrates, and nitrites [85,86]. These compounds
might react to form other toxic compounds such as peroxynitrous acid. Therefore, a comprehensive
evaluation of the effect of plasma on foods and human health is necessary before this new disinfection
technology can be fully utilized. Indeed, a range of applications of plasma technology in the field of
agriculture is currently being assessed.
Recently, the use of plasma technology has been reported to enhance seed germination and the
growth of plants [87]. In addition, the removal of volatile organic compounds, such as ethylene gas, by
plasma treatment may be useful during the transportation of agricultural products in containers [88].
Therefore, the application of plasma technology could also contribute to higher crop yields as well as
the preservation of foods.

5. Future Perspectives in Medicine and Dentistry

Sterilization of medical instruments contaminated with pathogens is crucial in preventing
secondary infections. Currently, medical instruments are sterilized by autoclaving, gamma-ray
treatment, UV exposure, and the use of gases such as ethylene oxide, hydrogen peroxide, formaldehyde,
peracetic acid [43]. Each sterilization method has both advantages and disadvantages. Autoclaving
is relatively quick, highly penetrative, and generates no toxic residues, but temperatures of 121 ◦ C
could damage the material being sterilized. Treatment with gamma-rays is highly penetrative, and
involves low temperatures with no associated residues, but it could induce changes in the properties
of the materials and is a relatively slow process. UV treatment is fast, low cost with no toxic residues,
and involves low temperatures, but the effectiveness of the sterilization is poor and may result in
damage to the material. Chemical treatments are low temperature and effective, but these procedures
generally involve the use of toxic gases that may be carcinogenic and flammable. Furthermore, these
gases sometimes induce unwanted biochemical changes. Although novel techniques have been
developed, such as chemical treatment with supercritical carbon dioxide as well as freeze-drying and
other methods, these procedures are often ineffective and may damage the material being sterilized.
Thus, it is necessary to evaluate each sterilization method for a particular purpose carefully. Plasma
technology is a promising new method that enables rapid processing at low temperatures without any
associated chemical residues [6]. Plasma is effective against a broad spectrum of pathogens, including
bacterial spores and prions, both of which display a high level of resistance to chemical and physical
treatments [47,89].
The potential of plasma technology in medical and dental applications is extremely broad. As well
as disinfection/sterilization of medical and dental devices, the technology could be used to treat beds,
desks, and floors [90]. Plasma technology may also have therapeutic potential [91–93]. Therapeutic uses
include the treatment of skin diseases [94], blood coagulation [95] as well as dental treatment [96] and
applications in dermatology such as chronic wound healing [97]. Recently, the potential application of
plasma technology as a novel anticancer therapy has been assessed [98]. Induction of cancer cell death
by both direct and indirect exposure of plasma has been reported [99,100]. However, as actual therapy,
plasma is often difficult to apply to cancer cells. As an alternative approach, a plasma-irradiated
medium (PAW, PAM, PSM, PTW, pPBS, NTP media) has been used for the treatment of cancer cells
Int. J. Mol. Sci. 2019, 20, 5216 10 of 17

and tumor tissues that resulted in cell killing and tumor-shrinking effects [41,101–103]. In addition,
PAM is reported to inhibit the MAP (Mitogen-activated protein) kinase (MAPK) pathway, which is an
important signaling pathway for cell proliferation [104]. Cell death is induced by the suppression of
these signaling cascades [105]. Moreover, ROS and RNS in plasma are key factors for the induction of
cancer cell death, although the mechanisms of action have not been fully elucidated [106,107].

6. Conclusions
In conclusion, plasma disinfection covers almost all of the resistance hierarchy of microorganisms.
The susceptibility to the plasma of microorganisms categorized as being most resistant, highly resistant,
intermediate resistant, less resistant, and very susceptible, have already been studied. Therefore, the
applicability of plasma technology in disinfection/sterilization is potentially wide-ranging (Figure 6).
However, to date, no studies investigating plasma treatment of viroids have been reported. Investigation
of the effect of plasma on various microorganisms would potentially contribute to further expanding
the applicability of this technology. In addition to the field of agriculture and medicine, plasma
technology has also been utilized in a range of environmental applications, including water purification
and remediation, as well as the treatment of exhaust gases [108,109]. We anticipate that the utilization
of
Int.this technology
J. Mol. Sci. 2019, 20,will continue
x FOR to expand.
PEER REVIEW 11 of 17

Figure6.6.Recent
Figure Recent andand potential
potential applications
applications of plasma
of plasma disinfection
disinfection technologytechnology
in the fieldinofthe field of
agriculture,
agriculture,
medicine, medicine,
dentistry, anddentistry, and environment.
environment. In thefield,
In the agricultural agricultural field, plasma
plasma technology technology
could be appliedcould
to
be applied
the to the
disinfection of disinfection of foods and
foods and packaging packaging
materials as wellmaterials as wellsources
as agricultural as agricultural sources
such as seeds, such as
fertilizers,
waters and soils. In
seeds, fertilizers, the medical
waters field,
and soils. Inplasma is useful
the medical forplasma
field, disinfection/sterilization of medical devices,
is useful for disinfection/sterilization
as
of well as the
medical degradation
devices, of the
as well as toxins and otherofpathological
degradation contaminants.
toxins and other pathological Potential applications
contaminants. of
Potential
this technology
applications of also
this includes skinalso
technology antisepsis
includesas skin
well antisepsis
as the treatment
as well of as
pathogen-based
the treatment skin diseases.
of pathogen-
In dentistry,
based plasma treatment
skin diseases. hasplasma
In dentistry, been used for microbicidal
treatment decontamination,
has been used for microbicidalincluding root canal
decontamination,
disinfection
including rootandcanal
toothdisinfection
disinfection.and
Plasma technology
tooth disinfection.mayPlasma
also betechnology
utilized in the
may environmental
also be utilizedfield,
in
including cleaning of
the environmental wastewater
field, includingascleaning
well as the treatment ofasexhaust
of wastewater well as gases.
the treatment of exhaust gases.

Finally, it should be mentioned that an increase in the disinfection efficiency and improved cost
performance is required before the true potential of plasma technology can be fully realized. This
may be achieved by optimization of the plasma generating conditions, including the use of different
gas mixtures and careful control of the relative humidity as well as plasma generation methods.
Author Contributions: For A.S., Y.Y., and T.O. contributed to the conceptualization and wrote the manuscript.
Int. J. Mol. Sci. 2019, 20, 5216 11 of 17

Finally, it should be mentioned that an increase in the disinfection efficiency and improved cost
performance is required before the true potential of plasma technology can be fully realized. This may
be achieved by optimization of the plasma generating conditions, including the use of different gas
mixtures and careful control of the relative humidity as well as plasma generation methods.

Author Contributions: For A.S., Y.Y., and T.O. contributed to the conceptualization and wrote the manuscript.
Funding: This work was supported by JSPS (Japan Society for the Promotion of Science) KAKENHI Grant number
JP16K04997 as well as Grant-in-aid from Takahashi Industrial and Economic Research Foundation.
Conflicts of Interest: The authors declare no conflict of interest.

Abbreviations
AC Alternating current
APPJ Atmospheric plasma jet
BLP-TES Bi-polar and low- pressure plasma-triple effects sterilization
BSE Bovine spongiform encephalopathy
CDC Centers for Disease Control
CJD Creutzfeldt-Jakob disease
CWD Chronic wasting disease
DBD Dielectric barrier discharge
DC Direct current
ELISA Enzyme-linked immunosorbent assay
FE-DBD Floating electrode barrier discharge
HIV Human immunodeficiency virus
LPS Lipopolysaccharides
MAP Mitogen-activated protein
MAPK MAP kinase
MHCD Micro hollow cathode discharge
ne Electron density
NTP Non-thermal plasma
PAM Plasma-activated medium
PAW Plasma-activated water
PHD Pin-to-hole spark discharge
pPBS Plasma-treated phosphate-buffered saline
PSM Plasma-stimulated medium
PTW Plasma-treated water
RF Radio-frequency
RNS Reactive nitrogen species
ROS Reactive oxygen species
RSV Respiratory syncytial virus
Te Electron temperature
Tgas Gas temperature
Tion Ion temperature
Tn Neutron temperature
USEPA U.S. Environmental Protection Agency
UV Ultraviolet
VOC Volatile organic compound
WHO World Health Organization

References
1. Langmuir, I. Oscillations in ionized gases. Proc. Natl. Acad. Sci. USA 1928, 14, 627–637. [CrossRef] [PubMed]
2. Ichimaru, S. Nuclear fusion in dense plasmas. Rev. Mod. Phys. 1993, 65, 255–259. [CrossRef]
3. Team, J. Fusion energy production from a deuterium-tritium plasma in the JET tokamak. Nuc. Fus. 1992,
32, 187–203. [CrossRef]
Int. J. Mol. Sci. 2019, 20, 5216 12 of 17

4. Coburn, J.W.; Winters, H.F. Plasma etching-A discussion of mechanisms. J. Vac. Sci. Technol. 1979, 16, 391–403.
[CrossRef]
5. Sakudo, A.; Shintani, H. Sterilization and Disinfection by Plasma: Sterilization Mechanisms, Biological and Medical
Applications (Medical Devices and Equipment); Nova Science Publishers: New York, NY, USA, 2010.
6. Shintani, H.; Sakudo, A. Gas Plasma Sterilization in Microbiology: Theory, Applications, Pitfalls and New
Perspectives; Caister Academic Press: London, UK, 2016.
7. Sakudo, A.; Shintani, H.; Yagyu, Y. Plasma sterilization. In Block’s Disinfection, Sterilization, and Preservation;
McDonnell, G.E., Ed.; Lippincott Williams & Wilkins: Philadelphia, PA, USA, 2019; in press.
8. Nandkumar, N. Plasma-The fourth state of matter. Int. J. Sci. Technol. Res. 2014, 3, 49–52.
9. Fridman, A. Plasma Chemistry; Cambridge University Press: Cambridge, UK, 2012.
10. Nehra, V.; Kumar, A.; Dwivedi, H. Atmospheric non-thermal plasma sources. Int J. Eng. 2008, 2, 53–68.
11. Boulos, M.I.; Fauchais, P.; Pfender, E. Thermal Plasma: Fundamental and Applications, Vol. 1; Plenum Press:
New York, NY, USA, 1994.
12. Boulos, M.I. New frontiers in thermal plasma processing. Pure Appl. Chem. 1996, 68, 1007–1010. [CrossRef]
13. Heberlein, J. New approaches in thermal plasma technology. Pure Appl. Chem. 2002, 74, 327–335. [CrossRef]
14. Bonnizzoni, G.; Vassallo, E. Plasma physics and technology; industrial applications. Vacuum 2002, 64, 327–336.
[CrossRef]
15. Hippler, R.; Kersten, H.; Schmidt, M.; Schoenbach, K.H. Low Temperature Plasmas. Fundamentals, Technologies,
and Techniques, 2nd ed.; Wiley-VCH: Weinheim, Germany, 2008.
16. Meichsner, J.; Schmidt, M.; Schneider, R.; Wagner, H.E. Nonthermal Plasma Chemistry and Physics; CRC Press,
Taylor & Francis Group: London, UK, 2013.
17. Taylor, P.R.; Pirzada, S.A. Thermal plasma processing of materials: A review. Adv. Perform. Mater. 1994,
1, 35–50. [CrossRef]
18. Dobrynin, D.; Friedman, G.; Fridman, A.; Starikovskiy, A. Inactivation of bacteria using dc corona discharge:
Role of ions and humidity. New J. Phys. 2011, 13, 103033. [CrossRef] [PubMed]
19. Rupf, S.; Lehmann, A.; Hannig, M.; Schafer, B.; Schubert, A.; Feldmann, U.; Schindler, A. Killing of adherent
oral microbes by a non-thermal atmospheric plasma jet. J. Med. Microbiol. 2010, 59, 206–212. [CrossRef]
[PubMed]
20. Eto, H.; Ono, Y.; Ogino, A.; Nagatsua, M. Low-temperature sterilization of wrapped materials using flexible
sheet-type dielectric barrier discharge. Appl. Phys. Lett. 2008, 93, 221502. [CrossRef]
21. Kolb, J.F.; Mohamed, A.A.H.; Price, R.O.; Swanson, R.J.; Bowman, A.; Chiavarini, R.L.; Stacey, M.;
Schoenbach, K.H. Cold atmospheric pressure air plasma jet for medical applications. Appl. Phys. Lett. 2008,
92, 241501. [CrossRef]
22. Dobrynin, D.; Arjunan, K.; Fridman, A.; Friedman, G.; Morss Clyne, A. Direct and controllable nitric oxide
delivery into biological media and living cells by a pin-to hole spark discharge (PHD) plasma. J. Phys. D
Appl. Phys. 2011, 44, 075201. [CrossRef]
23. Rossi, F.; Kylian, O.; Rauscher, H.; Gilliland, D.; Sirghi, L. Use of a low-pressure plasma discharge for the
decontamination and sterilization of medical devices. Pure Appl. Chem. 2008, 80, 1939–1951. [CrossRef]
24. Stapelmann, K.; Fiebrandt, M.; Raguse, M.; Awakowicz, P.; Reitz, G.; Moeller, R. Utilization of low-pressure
plasma to inactivate bacterial spores on stainless steel screws. Astrobiology 2013, 13, 597–606. [CrossRef]
25. Teschke, M.; Kedzierski, J.; Finantu-Dinu, E.G.; Korzec, D.; Engemann, J. High-speed photographs of a
dielectric barrier atmospheric pressure plasma jet. IEEE Trans. Plasma Sci. 2005, 33, 310–311. [CrossRef]
26. Kalghatgi, S.U.; Fridman, G.; Cooper, M.; Nagaraj, G.; Peddinghaus, M.; Balasubramanian, M.; Vasilets, V.N.;
Gutsol, A.F.; Fridman, A.; Friedman, G. Mechanism of blood coagulation by nonthermal atmospheric
pressure dielectric barrier discharge plasma. IEEE Trans. Plasma Sci. 2007, 35, 1559. [CrossRef]
27. Fridman, G.; Peddinghaus, M.; Balasubramanian, M.; Ayan, H.; Fridman, A.; Gutsol, A.; Brooks, A. Blood
coagulation and living tissue sterilization by floating-electrode dielectric barrier discharge in air. Plasma Chem.
Plasma Process. 2006, 26, 425. [CrossRef]
28. Cooper, M.; Fridman, G.; Fridman, A.; Joshi, S. Biological responses of Bacillus stratosphericus to floating
electrode-dielectric barrier discharge plasma treatment. J. Appl. Microbiol. 2010, 109, 2039. [CrossRef]
[PubMed]
Int. J. Mol. Sci. 2019, 20, 5216 13 of 17

29. Schoenbach, K.; Verhappen, R.; Tessnow, T.; Peterkin, F.; Byszewski, W. Microhollow cathode discharges.
Appl. Phys. Lett. 1996, 68, 13. [CrossRef]
30. Fridman, G.; Fridman, G.; Gutsol, A.; Shekhter, A.B.; Vasilets, V.N.; Fridman, A. Applied plasma medicine.
Plasma Process. Polym. 2008, 5, 503–533. [CrossRef]
31. Shintani, H.; Sakudo, A.; Burke, P.; McDonnell, G. Gas plasma sterilization of microorganisms and mechanisms
of action. Exp. Ther. Med. 2010, 1, 731–738. [CrossRef] [PubMed]
32. Pankaj, S.K.; Wan, Z.; Keener, K.M. Effects of cold plasma on food quality: A review. Foods 2018, 7, 4.
[CrossRef] [PubMed]
33. Kamgang-Youbi, G.; Herry, J.M.; Meylheuc, T.; Brisset, J.L.; Bellon-Fontaine, M.N.; Doubla, A.; Naitali, M.
Microbial inactivation using plasma-activated water obtained by gliding electric discharges. Lett. Appl.
Microbiol. 2009, 48, 13–18. [CrossRef] [PubMed]
34. Duan, J.; Lu, X.; He, G. The selective effect of plasma activated medium in an in vitro co-culture of liver
cancer and normal cells. J. Appl. Phys. 2017, 121, 013302. [CrossRef]
35. Yan, D.; Talbot, A.; Nourmohammadi, N.; Cheng, X.; Canady, J.; Sherman, J.; Keidar, M. Principles of using
cold atmospheric plasma stimulated media for cancer treatment. Sci. Rep. 2015, 5, 18339. [CrossRef]
36. Park, J.Y.; Park, S.; Choe, W.; Yong, H.I.; Jo, C.; Kim, K. Plasma-functionalized solution: A potent antimicrobial
agent for biomedical applications from antibacterial therapeutics to biomaterial surface engineering. ACS Appl.
Mater. Interfaces 2017, 9, 43470–43477. [CrossRef]
37. Ikawa, S.; Tani, A.; Nakashima, Y.; Kitano, K. Physicochemical properties of bactericidal plasma-treated
water. J. Phys. D Appl. Phys. 2016, 49, 425401. [CrossRef]
38. Van Boxem, W.; Van der Paal, J.; Gorbanev, Y.; Vanuytsel, S.; Smits, E.; Dewilde, S.; Bogaerts, A. Anti-cancer
capacity of plasma-treated PBS: Effect of chemical composition on cancer cell cytotoxicity. Sci Rep. 2017,
7, 16478. [CrossRef] [PubMed]
39. Liedtke, K.R.; Bekeschus, S.; Kaeding, A.; Hackbarth, C.; Kuehn, J.P.; Heidecke, C.D.; von Bernstorff, W.;
von Woedtke, T.; Partecke, L.I. Non-thermal plasma-treated solution demonstrates antitumor activity against
pancreatic cancer cells in vitro and in vivo. Sci. Rep. 2017, 7, 8319. [CrossRef] [PubMed]
40. Balan, G.G.; Rosca, I.; Ursu, E.L.; Doroftei, F.; Bostanaru, A.C.; Hnatiuc, E.; Nastasa, V.; Sandru, V.;
Stefanescu, G.; Trifan, A.; et al. Plasma-activated water: A new and effective alternative for duodenoscope
reprocessing. Infect. Drug Resist. 2018, 11, 727–733. [CrossRef] [PubMed]
41. Tanaka, H.; Mizuno, M.; Ishikawa, K.; Nakamura, K.; Utsumi, F.; Kajiyama, H.; Kano, H.; Maruyama, S.;
Kikkawa, F.; Hori, M. Cell survival and proliferation signaling pathways are downregulated by
plasma-activated medium in glioblastoma brain tumor cells. Plasma Med. 2012, 2, 207–220. [CrossRef]
42. Sakudo, A.; Toyokawa, Y.; Imanishi, Y.; Murakami, T. Crucial roles of reactive chemical species in modification
of respiratory syncytial virus by nitrogen gas plasma. Mater. Sci. Eng. C Mater. Biol. Appl. 2017, 74, 131–136.
[CrossRef]
43. McDonnell, G.E. Antisepsis, Disinfection, and Sterilization; ASM Press: Washington, DC, USA, 2007.
44. Knipe, D.M.; Howley, P.M.; Griffin, D.E.; Lamb, R.A.; Martin, M.A. Fields Virology, 5th ed.; Lippincott Williams
& Wilkins: Philadelphia, PA, USA, 2006.
45. Yokoyama, T.; Murai, K.; Murozuka, T.; Wakisaka, A.; Tanifuji, M.; Fujii, N.; Tomono, T. Removal of small
non-enveloped viruses by nanofiltration. Vox Sang. 2004, 86, 225–229. [CrossRef]
46. Niemira, B.A. Cold plasma reduction of Salmonella and Escherichia coli O157:H7 on almonds using ambient
pressure gases. J. Food Sci. 2012, 77, M171–M175. [CrossRef]
47. Klampfl, T.G.; Isbary, G.; Shimizu, T.; Li, Y.F.; Zimmermann, J.L.; Stolz, W.; Schlegel, J.; Morfill, G.E.;
Schmidt, H.U. Cold atmospheric air plasma sterilization against spores and other microorganisms of clinical
interest. Appl. Environ. Microbiol. 2012, 78, 5077–5082. [CrossRef]
48. Sung, S.J.; Huh, J.B.; Yun, M.J.; Chang, B.M.; Jeong, C.M.; Jeon, Y.C. Sterilization effect of atmospheric
pressure non-thermal air plasma on dental instruments. J. Adv. Prosthodont 2013, 5, 2–8. [CrossRef]
49. Tian, Y.; Sun, P.; Wu, H.; Bai, N.; Wang, R.; Zhu, W.; Zhang, J.; Liu, F. Inactivation of Staphylococcus aureus and
Enterococcus faecalis by a direct-current, cold atmospheric-pressure air plasma microjet. J. Biomed. Res. 2010,
24, 264–269. [CrossRef]
50. Vandervoort, K.G.; Brelles-Marino, G. Plasma-mediated inactivation of Pseudomonas aeruginosa biofilms
grown on borosilicate surfaces under continuous culture system. PLoS ONE 2014, 9, e108512. [CrossRef]
[PubMed]
Int. J. Mol. Sci. 2019, 20, 5216 14 of 17

51. Niemira, B.A.; Boyd, G.; Sites, J. Cold plasma rapid decontamination of food contact surfaces contaminated
with Salmonella biofilms. J. Food Sci. 2014, 79, M917–M922. [CrossRef] [PubMed]
52. WHO (World Health Organization). Antimicrobial Resistance. Available online: http://www.who.int/
mediacentre/factsheets/fs194/en/ (accessed on 24 June 2019).
53. CDC (The US Centers for Disease Control and Prevention). National Antimicrobial Resistance Monitoring
System for Enteric Bacteria (NARMS). Available online: https://www.cdc.gov/narms/index.html (accessed on
24 June 2019).
54. Krauland, M.G.; Marsh, J.W.; Paterson, D.L.; Harrison, L.H. Integron-mediated multidrug resistance in a
global collection of nontyphoidal Salmonella enterica isolates. Emerg Infect. Dis. 2009, 15, 388–396. [CrossRef]
[PubMed]
55. Maeda, K.; Toyokawa, Y.; Shimizu, N.; Imanishi, Y.; Sakudo, A. Inactivation of Salmonella by nitrogen gas
plasma generated by a static induction thyristor as a pulsed power supply. Food Control. 2015, 52, 54–59.
[CrossRef]
56. Guo, L.; Xu, R.; Zhao, Y.; Liu, D.; Liu, Z.; Wang, X.; Chen, H.; Kong, M.G. Gas Plasma pre-treatment increases
antibiotic sensitivity and persister eradication in methicillin-resistant Staphylococcus aureus. Front. Microbiol.
2018, 9, 537. [CrossRef]
57. Sakudo, A.; Toyokawa, Y.; Misawa, T.; Imanishi, Y. Degradation and detoxification of aflatoxin B1 using
nitrogen gas plasma generated by a static induction thyristor as a pulsed power supply. Food Control. 2017,
73B, 619–626. [CrossRef]
58. Scholtz, V.; Pazlarova, J.; Souskova, H.; Khun, J.; Julak, J. Nonthermal plasma-A tool for decontamination
and disinfection. Biotechnol. Adv. 2015, 33, 1108–1119. [CrossRef]
59. Soušková, H.; Scholtz, V.; Julák, J.; Savická, D. The fungal spores survival under the low-temperature plasma.
In Plasma for Bio-Decontamination, Medicine and Food Security (NATO Science for Peace and Security Series A:
Chemistry and Biology); Machala, Z., Hensel, K., Akishev, Y., Eds.; Springer: New York, NY, USA, 2012;
pp. 57–66.
60. Souskova, H.; Scholtz, V.; Julak, J.; Kommova, L.; Savicka, D.; Pazlarova, J. The survival of micromycetes and
yeasts under the low-temperature plasma generated in electrical discharge. Folia Microbiol. (Praha) 2011,
56, 77–79. [CrossRef]
61. Sakudo, A.; Shimizu, N.; Imanishi, Y.; Ikuta, K. N2 gas plasma inactivates influenza virus by inducing
changes in viral surface morphology, protein, and genomic RNA. Biomed. Res. Int. 2013, 2013, 694269.
[CrossRef]
62. Sakudo, A.; Toyokawa, Y.; Imanishi, Y. Nitrogen gas plasma generated by a static induction thyristor as a
pulsed power supply inactivates adenovirus. PLoS ONE 2016, 11, e0157922. [CrossRef]
63. Alshraiedeh, N.H.; Alkawareek, M.Y.; Gorman, S.P.; Graham, W.G.; Gilmore, B.F. Atmospheric pressure,
nonthermal plasma inactivation of MS2 bacteriophage: Effect of oxygen concentration on virucidal activity.
J. Appl. Microbiol. 2013, 115, 1420–1426. [CrossRef] [PubMed]
64. Guo, L.; Xu, R.; Gou, L.; Liu, Z.; Zhao, Y.; Liu, D.; Zhang, L.; Chen, H.; Kong, M.G. Mechanism of virus
inactivation by cold atmospheric-pressure plasma and plasma-activated water. Appl. Environ. Microbiol.
2018, 84. [CrossRef] [PubMed]
65. Wu, Y.; Liang, Y.; Wei, K.; Li, W.; Yao, M.; Zhang, J.; Grinshpun, S.A. MS2 virus inactivation by
atmospheric-pressure cold plasma using different gas carriers and power levels. Appl. Environ. Microbiol.
2015, 81, 996–1002. [CrossRef] [PubMed]
66. Yamashiro, R.; Misawa, T.; Sakudo, A. Key role of singlet oxygen and peroxynitrite in viral RNA damage
during virucidal effect of plasma torch on feline calicivirus. Sci. Rep. 2018, 8, 17947. [CrossRef] [PubMed]
67. USEPA (US Environmental Protection Agency). Registration Division Office of Pesticide Program, Criteria
and Standards Division Office of Drinking Water, Guide Standard and Protocol for Testing Microbiological
Water Purifiers. Available online: https://nepis.epa.gov/Exe/ZyPDF.cgi/9102362E.PDF?Dockey=9102362E.pdf
(accessed on 24 June 2019).
68. Filipic, A.; Primc, G.; Zaplotnik, R.; Mehle, N.; Gutierrez-Aguirre, I.; Ravnikar, M.; Mozetic, M.; Zel, J.;
Dobnik, D. Cold atmospheric plasma as a novel method for inactivation of potato virus Y in water samples.
Food Environ. Virol. 2019, 11, 220–228. [CrossRef] [PubMed]
Int. J. Mol. Sci. 2019, 20, 5216 15 of 17

69. Tyczkowska-Sieron, E.; Markiewicz, J.; Grzesiak, B.; Krukowski, H.; Glowacka, A.; Tyczkowski, J. Short
communication: Cold atmospheric plasma inactivation of Prototheca zopfii isolated from bovine milk.
J. Dairy Sci. 2018, 101, 118–122. [CrossRef] [PubMed]
70. Wang, X.Q.; Wang, F.P.; Chen, W.; Huang, J.; Bazaka, K.; Ostrikov, K.K. Non-equilibrium plasma prevention
of Schistosoma japonicum transmission. Sci. Rep. 2016, 6, 35353. [CrossRef]
71. Hayes, J.; Kirf, D.; Garvey, M.; Rowan, N. Disinfection and toxicological assessments of pulsed UV
and pulsed-plasma gas-discharge treated-water containing the waterborne protozoan enteroparasite
Cryptosporidium parvum. J. Microbiol. Methods 2013, 94, 325–337. [CrossRef]
72. Baxter, H.C.; Campbell, G.A.; Whittaker, A.G.; Jones, A.C.; Aitken, A.; Simpson, A.H.; Casey, M.;
Bountiff, L.; Gibbard, L.; Baxter, R.L. Elimination of transmissible spongiform encephalopathy infectivity
and decontamination of surgical instruments by using radio-frequency gas-plasma treatment. J. Gen. Virol.
2005, 86, 2393–2399. [CrossRef]
73. von Keudell, A.; Awakowicz, P.; Benedikt, J.; Raballand, V.; Yanguas-Gil, A.; Opretzka, J.; Flötgen, C.;
Reuter, R.; Byelykh, L.; Halfmann, H.; et al. Inactivation of bacteria and biomolecules by low pressure plasma
discharges. Plasma Process. Polym. 2010, 7, 327–352. [CrossRef]
74. Sakudo, A.; Imanishi, Y. Degradation and inactivation of Shiga toxins by nitrogen gas plasma. AMB Express
2017, 7, 77. [CrossRef] [PubMed]
75. McCombs, G.B.; Darby, M.L. New discoveries and directions for medical, dental and dental hygiene research:
Low temperature atmospheric pressure plasma. Int. J. Dent. Hyg. 2010, 8, 10–15. [CrossRef] [PubMed]
76. Shintani, H.; Shimizu, N.; Imanishi, Y.; Sekiya, T.; Tamazawa, K.; Taniguchi, A.; Kido, N. Inactivation of
microorganisms and endotoxins by low temperature nitrogen gas plasma exposure. Biocontrol. Sci. 2007,
12, 131–143. [CrossRef] [PubMed]
77. Liao, X.; Liu, D.; Xiang, Q.; Ahn, J.; Chen, S.; Ye, X.; Ding, T. Inactivation mechanisms of non-thermal plasma
on microbes: A review. Food Control. 2017, 75, 83–91. [CrossRef]
78. Traba, C.; Chen, L.; Liang, J.F. Low power gas discharge plasma mediated inactivation and removal of
biofilms formed on biomaterials. Curr. Appl. Phys. 2013, 13, S12–S18. [CrossRef]
79. Kylian, O.; Sasaki, T.; Rossi, F. Plasma sterilization of Geobacillus stearothermophilus by O2 :N2 RF inductively
coupled plasma. Eur Phys. J. Appl. Phys. 2006, 34, 139–142. [CrossRef]
80. Rossi, F.; Kylian, O.; Hasiwa, M. Decontamination of surfaces by low pressure plasma discharges.
Plasma Process. Polym. 2006, 3, 431–442. [CrossRef]
81. Rossi, F.; Kylian, O.; Hasiwa, M. Mechanisms of sterilization and decontamination of surfaces by low-pressure
plasma. In Advanced Plasma Technology; D’Agostino, R., Favia, P., Kawai, Y., Ikegami, H., Sato, N.,
Arefi-Khonsari, F., Eds.; Wiley-VCH Verlag GmbH &Co.: Weinheim, Germany, 2008; pp. 319–340.
82. Pignata, C.; D’Angelo, D.; Fea, E.; Gilli, G. A review on microbiological decontamination of fresh produce
with nonthermal plasma. J. Appl. Microbiol. 2017, 122, 1438–1455. [CrossRef]
83. Toyokawa, Y.; Yagyu, Y.; Misawa, T.; Sakudo, A. A new roller conveyer system of non-thermal gas plasma as
a potential control measure of plant pathogenic bacteria in primary food production. Food Control. 2017,
72A, 62–72. [CrossRef]
84. European Commission. Plasma gas technique as electronic preservation practice of organic food and feed,
EGTOP/2014, Directorate-General for Agriculture and Rural Development, Directorate, B. Multilateral
relations, quality policy, B.4. Organics, Expert Group for Technical Advice on Organic Production EGTOP,
Final Report on Food (III). Available online: https://ec.europa.eu/agriculture/organic/sites/orgfarming/files/
docs/body/egtop-final-report-food-iii_en.pdf (accessed on 26 April 2018).
85. Naitali, M.; Kamgang-Youbi, G.; Herry, J.M.; Bellon-Fontaine, M.N.; Brisset, J.L. Combined effects
of long-living chemical species during microbial inactivation using atmospheric plasma-treated water.
Appl. Environ. Microbiol. 2010, 76, 7662–7664. [CrossRef]
86. Traylor, M.J.; Pavlovich, M.J.; Karim, S.; Hait, P.; Sakiyama, Y.; Clark, D.S.; Graves, D.B. Long-term antibacterial
efficacy of air plasma-activated water. J. Phys. D Appl. Phys. 2011, 44, 472001. [CrossRef]
87. Ito, M.; Oh, J.S.; Ohta, T.; Shiratani, M.; Hori, M. Current status and future prospects of agricultural
applications using atmospheric-pressure plasma technologies. Plasma Process. Polym. 2017, 15, e1700073.
[CrossRef]
Int. J. Mol. Sci. 2019, 20, 5216 16 of 17

88. Nishimura, J.; Takahashi, K.; Takaki, K.; Koide, S.; Suga, M.; Orikasa, T.; Teramoto, Y.; Uchino, T. Removal of
ethylene and by-products using dielectric barrier discharge with Ag nanoparticle-loaded Zeolite for keeping
freshness of fruits and vegetables. Trans. Mater. Res. Soc. Jpn. 2016, 41, 41–45. [CrossRef]
89. Itarashiki, T.; Ohshiro, S.; Sakudo, A.; Hayashi, N. Current trend of medical sterilization and disinfection
methods using plasmas. J. Plasma Fus. Res. 2015, 91, 505–513.
90. Shiely, F.; Fallon, D.; Casey, C.; Kerins, D.M.; Eustace, J.A. Trial of a novel plasma gas disinfection system
(Radica) to reduce mattress residual bacterial contamination in the acute hospital setting: A preliminary
study. Ir J. Med. Sci. 2017, 186, 17–21. [CrossRef]
91. Šimončicová, J.; Kryštofová, S.; Medvecká, V.; Ďurišová, K.; Kaliňáková, B. Technical applications of plasma
treatments: Current state and perspectives. Appl. Microbiol. Biotechnol. 2019, 103, 5117–5129. [CrossRef]
92. Fridman, A.; Friedman, G. Plasma Medicine; Wiley-Blackwell: Oxford, UK, 2013.
93. Laroussi, M.; Kong, M.G.; Morfill, G.; Stolz, W. Plasma Medicine: Applications of Low-Temperature Gas. Plasmas
in Medicine and Biology; Cambridge University Press: Cambridge, UK, 2012.
94. Wu, A.S.; Kalghatgi, S.; Dobrynin, D.; Sensenig, R.; Cerchar, E.; Podolsky, E.; Dulaimi, E.; Paff, M.; Wasko, K.;
Arjunan, K.P.; et al. Porcine intact and wounded skin responses to atmospheric nonthermal plasma.
J. Surg. Res. 2013, 179, e1–e12. [CrossRef]
95. Shahbazi Rad, Z.; Abbasi Davani, F. Experimental investigation on electrical characteristics and dose
measurement of dielectric barrier discharge plasma device used for therapeutic application. Rev. Sci. Instrum.
2017, 88, 043504. [CrossRef]
96. Ballout, H.; Hertel, M.; Doehring, J.; Kostka, E.; Hartwig, S.; Paris, S.; Preissner, S. Effects of plasma jet,
dielectric barrier discharge, photodynamic therapy and sodium hypochlorite on infected curved root canals.
J. Biophotonics 2018, 11, e201700186. [CrossRef]
97. Gan, L.; Zhang, S.; Poorun, D.; Liu, D.; Lu, X.; He, M.; Duan, X.; Chen, H. Medical applications of nonthermal
atmospheric pressure plasma in dermatology. J. Dtsch. Dermatol. Ges. 2018, 16, 7–13. [CrossRef]
98. Choi, J.S.; Kim, J.; Hong, Y.J.; Bae, W.Y.; Choi, E.H.; Jeong, J.W.; Park, H.K. Evaluation of non-thermal
plasma-induced anticancer effects on human colon cancer cells. Biomed. Opt. Express 2017, 8, 2649–2659.
[CrossRef] [PubMed]
99. Lin, A.; Gorbanev, Y.; De Backer, J.; Van Loenhout, J.; Van Boxem, W.; Lemiere, F.; Cos, P.; Dewilde, S.;
Smits, E.; Bogaerts, A. Non-thermal plasma as a unique delivery system of short-lived reactive oxygen
and nitrogen species for immunogenic cell death in melanoma cells. Adv. Sci. (Weinh) 2019, 6, 1802062.
[CrossRef] [PubMed]
100. Azzariti, A.; Iacobazzi, R.M.; Di Fonte, R.; Porcelli, L.; Gristina, R.; Favia, P.; Fracassi, F.; Trizio, I.; Silvestris, N.;
Guida, G.; et al. Plasma-activated medium triggers cell death and the presentation of immune activating
danger signals in melanoma and pancreatic cancer cells. Sci. Rep. 2019, 9, 4099. [CrossRef] [PubMed]
101. Tanaka, H.; Mizuno, M.; Toyokuni, S.; Maruyama, S. Cancer therapy using non-thermal atmospheric pressure
plasma with ultra-high electron density. Phys. Plasmas 2015, 22, 122004. [CrossRef]
102. Hattori, N.; Yamada, S.; Torii, K.; Takeda, S.; Nakamura, K.; Tanaka, H.; Kajiyama, H.; Kanda, M.; Fujii, T.;
Nakayama, G.; et al. Effectiveness of plasma treatment on pancreatic cancer cells. Int. J. Oncol. 2015,
47, 1655–1662. [CrossRef]
103. Utsumi, F.; Kajiyama, H.; Nakamura, K.; Tanaka, H.; Mizuno, M.; Ishikawa, K.; Kondo, H.; Kano, H.;
Hori, M.; Kikkawa, F. Effect of indirect nonequilibrium atmospheric pressure plasma on anti-proliferative
activity against chronic chemo-resistant ovarian cancer cells in vitro and in vivo. PLoS ONE 2013, 8, e81576.
[CrossRef]
104. Nakamura, K.; Peng, Y.; Utsumi, F.; Tanaka, H.; Mizuno, M.; Toyokuni, S.; Hori, M.; Kikkawa, F.; Kajiyama, H.
Novel Intraperitoneal Treatment With Non-thermal plasma-activated medium inhibits metastatic potential
of ovarian cancer cells. Sci. Rep. 2017, 7, 6085. [CrossRef]
105. Sato, T.; Yokoyama, M.; Johkura, K. A key inactivation factor of HeLa cell viability by a plasma flow. J. Phys.
D Appl. Phys. 2011, 44, 372001. [CrossRef]
106. Machala, Z.; Tarabova, B.; Hensel, K.; Spetlikova, E.; Sikurova, L.; Lukes, P. Formation of ROS and RNS in
water electro-sprayed through transient spark discharge in air and their bactericidal effects. Plasma Process.
Polym. 2013, 2013, 649–659. [CrossRef]
Int. J. Mol. Sci. 2019, 20, 5216 17 of 17

107. Kurake, N.; Tanaka, H.; Ishikawa, K.; Kondo, T.; Sekine, M.; Nakamura, K.; Kajiyama, H.; Kikkawa, F.;
Mizuno, M.; Hori, M. Cell survival of glioblastoma grown in medium containing hydrogen peroxide and/or
nitrite, or in plasma-activated medium. Arch. Biochem. Biophys. 2016, 605, 102–108. [CrossRef]
108. Hashim, S.A.; Samsudin, F.N.; Wong, C.S.; Abu Bakar, K.; Yap, S.L.; Mohd Zin, M.F. Non-thermal plasma for
air and water remediation. Arch. Biochem. Biophys. 2016, 605, 34–40. [CrossRef] [PubMed]
109. Magureanu, M.; Piroi, D.; Mandache, N.B.; David, V.; Medvedovici, A.; Bradu, C.; Parvulescu, V.I. Degradation
of antibiotics in water by non-thermal plasma treatment. Water Res. 2011, 45, 3407–3416. [CrossRef] [PubMed]

© 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/).