Plasma Medicine, 11(4):31 – 40 (2021)

Reactive Radical Study Using Polyvinyl
Alcohol–Potassium Iodide Solution as a New
Chemical Probe
H. Matsuura,a,* B. Ouanthavinsak,a T.N. Tran,a M. Hu,a J. Sakamoto,aY. Takemura,b R. Asada,a & M. FurutaaaOsaka Prefecture University, Gakuen-cho 1-2, Sakai Naka-ku, 599-8570, Japan; bKindai
University, Kowakae 3-4-1, Higashiosaka, 577-8502, Japan*Address all correspondence to: H. Matsuura, Osaka Prefecture University, Gakuen-cho 1-2, Sakai Naka-ku, 599-8570,
Japan; Tel.: +81-72-254-7149; Fax: +81-72-254-9938, E-mail: matsu@me.osakafu-u.ac.jp

ABSTRACT: Polyvinyl alcohol–potassium iodine (PVA-KI) solution as a radiation dose meterhas been proposed because it shows clear color changes in the presence of reactive radicals. Here,
PVA-KI is studied to visualize the production and transport of radicals during the interaction between the atmospheric pressure plasma jet and the solution. First, we focus on the challenges ofstoring the PVA-KI sample solution at different temperatures. The reproducibility of the PVA-KI
color reaction depends on this condition. Additionally, preliminary findings on the radical production mechanism and radical types are provided.
KEY WORDS: atmospheric pressure plasma, reactive oxygen speces, polyvinyl alcohol–potassium iodine, absorbance spectra

I. INTRODUCTION
Atmospheric pressure discharge plasma has been receiving attention in various fields,
in particular for its biomedical and sterilization applications.1–4 Plasma generates many
types of free radical species during its interaction with liquids,5–6 such as reactive oxygen species (ROS) and reactive nitrogen species (RNS), which play important roles inbiomedical applications.7–9 The effects of the gas-liquid interface and gas phase surrounding plasma jets on the produced amount and distribution of radicals have been
investigated.6,10 The production of radicals in the liquid by irradiation with the plasma jet
has also been studied.11–14 One of the important radicals is hydroxyl (OH). The production and distribution of OH radicals in the gas phase have been studied by laser-induced
fluorescence imaging.13,15–19 OH radical concentration in the liquid phase has been measured by fluorescence spectroscopy using terephthalic acid as a probe.20–21 To study the
production and transport of radicals, the chemical reaction of the specific compounds isa useful tool.22 Potassium iodide (KI) starch is the best-known chemical probe. Plasmairradiated solutions containing KI starch show clear color changes, and the distribution
of ROS can be visualized.23–25
A similar color reaction has been reported in radiation chemistry. For example,
polyvinyl alcohol (PVA)-KI has a complex reaction mechanism.26–30 The application of
PVA-KI as a chemical dose meter for X-ray and proton beams has been proposed.31–33
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The PVA-KI dose meter has been demonstrated to be inexpensive, highly sensitive, safe,and reusable. However, its reaction mechanism has not been applied in atmosphericplasma research yet. In this work, a PVA-KI solution reaction is proposed as a newchemical probe to study the production of radicals. Many irradiation conditions havebeen reported to affect the color of PVA-KI. This research focuses on the impact of thesample temperature during storage and irradiation condition.
When a PVA-KI solution stored for a long time is used, the color change upon
plasma irradiation is drastically different from results obtained using a radiation dose
meter. It was found that this difference is not due to the difference in electron energy
required to produce ROS in the solution. The irradiation distance between the plasma
source and the PVA–KI solution affects the ROS production mechanism. This problem
is discussed with comparisons in radiation chemistry. Additionally, a preliminary study
on the type and amount of ROS is also conducted.
II. EXPERIMENTAL PROCEDURES
The configuration of the dielectric barrier discharge (DBD) plasma system is shownin Fig. 1. Helium or argon was used as the working gas. The working gas was guided
to flow through a borosilicate glass tube, which served as the dielectric barrier. The
inner diameter of the tube was 8 mm, reduced to 1.5 mm at the nozzle exit. The inner

FIG. 1: Schematic of low-frequency dielectric barrier discharge plasma system with needle
electrode
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needle electrode was made of Tungsten and its diameter was 0.3 mm. The outer electrode was made of copper and its length was 15 mm. The gas flow rate was constantly
controlled at 2 L/min using a flow meter (Kofloc, Model RK1710). When gas molecules flow through the electric field of an electrode supplied with a high voltage ata low frequency (LF-HV) using a commercial power source (HPS008-100 Optimize,
Chuen Electrics Co. Ltd.), a DBD plasma jet is subsequently generated and emitted
from the nozzle exit. This power source was upgraded from that used in our previous study12 and controlled using a transformer. The applied supply voltages wereapproximately 4 and 7 kV for helium and argon, respectively. The frequency was
10 kHz. The distance of the plasma jet between the glass nozzle exit and the surface
sample was usually set as 20 mm. The length of the helium plasma jet from the outlet
tube was 20 mm.
Table 1 shows the composition of the PVA-KI sample used in this study. Before theplasma irradiation experiments, the sample was mixed using a stirrer; it was colorless,like pure water. As the PVA volume increased (> 50 mL), the sample became a gel.
The radical study of the PVA-KI gel will be reported elsewhere. The reaction between
PVA-KI and ROS has been explained in a previous study26,27 and is described in Fig.
2(a). An iodide ion is oxidized by ROS and creates an iodide molecule, which is insoluble in water and forms a ploy iodide ion, which forms a complex with maldistribution residual acetyl groups in PVA, resulting in a red or dark color. The amount of KI isrelated to the sensitivity of the PVA-KI color reaction. Borax is Na2B4O7·10H2O, the saltof a strong base and a weak acid, hydrolyzed in the water. It contains boron, which caneasily form a bond with oxygen. In water, tetraborate ions combine in a complex shape;
however, when PVA is added, they form a bond with the PVA’s oxygen. As a result, the
PVA and Borax bond with each other to form a large network polymer.
A spectrophotometer (HITACHI U-2001) was used to measure the absorbance
spectra of the plasma-irradiated solution samples. The spectrophotometer was calibrated, and the wavelength was set in the range of 350–650 nm. In a double-beamdesign, the energy of the light source is divided in two with a half mirror so that one
beam passes through the reference side, while the other passes through the sample side.
This phenomenon is unachievable with the single-beam design. The absorbance (ABS)
was in the range of 0–3. The spectrophotometer was connected to a computer to obtain
the absorption data, and the function for automated data storage in a USB drive wasincorporated.
TABLE 1: Composition PVA-KI solution
Composition
Type
Ultra pure water
LiquidPVA
Gel
KI
PowderBorax
PowderVolume 11, Issue 4, 2021

Weight (g)——12.5 & 25
5

Volume (mL)
250
10
—
—

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FIG. 2: (a) PVA structure containing two groups of hydroxyl and acetic acid showing oxidationof KI molecules producing iodine ions. (b) Old sample exhibits yellow color (stored for 3 months
before irradiation); the new sample exhibits red color (stored for 3 days before irradiation).

III. RESULTS
In the initial phase of our research with PVA-KI, the reproducibility of the color reaction
was a challenge. As shown in Fig. 2(b), it was difficult to compare the experimental results obtained on different days. To examine reproducibility, the prepared PVA-KI sample was divided into two parts: one stored at room temperature and the other stored at thecontrolled temperature (3°C) in a refrigerator. After various storage times (3, 6, 9, 12,15, and 18 days), plasma irradiation was conducted for 2 min. Beforehand, the samples
stored at the controlled temperature were left to reach room temperature. Figure 3 showsthat the plasma-irradiated PVA-KI samples exhibited different colors at room temperature—from red or dark to yellow or light, as the storage time increased. Conversely, thesamples stored in the refrigerator for a long time retained their red or dark color after
plasma irradiation. This storage time effect or temperature effect was also found in theabsorbance spectra data. As shown in Fig. 4(a), the absorbance peak of the sample stored
at 3°C was observed at approximately 490 nm. Peak intensity gradually decreased withincreaseing storage time; however, the color change trend was similar for all samples.
For the samples stored at room temperature, the absorbance peak was observed at 490
nm for 3 and 9 days of storage, and shifted to 390 nm for 12–18 days. From this observation, we defined the experimental protocol for the PVA-KI chemical probe and ensured
its experimental reproducibility.
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FIG. 3: PVA-KI samples irradiated for 2 min after 3–18 days at different storage temperatures:room temperature (a) and controlled temperature of 3°C (b)FIG. 4: Absorbance spectra of plasma-irradiated samples after 3–18 days of storage: (a) controlled temperature of 3°C; (b) room temperature

One example of the radical production comparison is shown in Fig. 5. The plasma
source exit was set at distances of 20–50 mm from the PVA-KI sample surface. After 2
min of plasma irradiation, the color change and absorbance spectra were compared. The
length of the plasma jet was approximately 20 mm. Therefore, the plasma jet reacted
directly with the surface of the PVA-KI sample at this distance. The detected ROS camefrom water molecules decomposed by plasma electrons. Contrarily, the plasma jet wascompletely outside of the glass beaker with the PVA-KI sample for the 50-mm distance.
Water vapor or oxygen molecules in the surrounding air was the origin of the detected
ROS. The absorbance peak intensity gradually decreased as the irradiation distance
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FIG. 5: Effect of plasma irradiation distance on ROS production showing PVA-KI samples irradiated by plasma jet for 2 min; distances between device exit and sample surface set to 20, 30,
40, and 50 mm: (a) photo image of plasma irradiated PVA-KI samples; (b) absorbance spectraof the same samples

increased, although the radical production mechanism changed drastically. This point
will be discussed later. For distances less than 20 mm, the discharge inside the glass
tube became unstable and local heating of the interface spot occurred. Consequently,
distances less than 20 mm were excluded in this work.
When the irradiation configuration was fixed, the absorbance peak of the plasmairradiated PVA-KI increased with irradiation time. Figure 6 shows the results for 1–5
min. In this study, the irradiation distance was 20 mm, and two samples with different
KI concentrations were compared. The results were similar to those of our previous
research,31 indicating that the concentration of KI significantly influences the sensitivityof the PVA-KI color reaction. Further, the 490-nm absorbance peak intensity was a goodmeasure of the produced ROS or “dose.” In this experiment, the higher the concentration of KI, the faster the change in the solution color. When the concentration of KI ishigh, iodine ions can be easily oxidized by ROS to produce poly iodine ions, which arecaptured by PVA.
IV. DISCUSSION
When previous works on the PVA-KI chemical dose meter31–33 were reviewed, absorbance spectra similar to those in Fig. 5 were observed, and the peak intensity at
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FIG. 6: Absorbance peak at 490 nm for PVA-KI with different KI concentrations, irradiated withplasma for 1–5 min

approximately 490 nm correlated well with dose rate. In radiation chemistry, it is known
that high-energy secondary electrons produced in solution by X-rays or gamma rays
decompose water and other liquid molecules. Many types of ROS are produced in thesolution which enhance chemical reactions and exert biological effects (the indirect effects of radiation). Since similar PVA-KI reactions have been confirmed for atmosphericpressure plasma jet systems, a similar process occurs in plasma irradiation. However,since plasma electron energy is only a few eVs, this process is limited to a narrow interfacial region, and its contribution decreases with the distance between the plasma source
and the sample surface.
A bright plasma jet contains many electrons around it. Such electrons react withthe water or oxygen molecules around the plasma jet, yielding some ROSs, which aretransported to the solution via gas flow. For the 50-mm distance, as shown in Fig. 5,
this second process was dominant, while the gas-liquid interfacial reaction made small
contributions. A detailed study of PVA-KI gel is ongoing.
Among ROSs, the OH radical has high ionizing power, and ethanol is conventionally employed as the OH scavenger. Figure 7 shows the effect of ethanol addition on the
PVA-KI reaction. The distance between the plasma source exit and the sample surface
was 20 mm. Ethanol suppressed the color reaction of the PVA-KI, and the intensity of theobserved absorbance peak at 490 nm decreased with ethanol concentration. This indicatesthat the OH radicals produced by plasma make the largest contribution to the PVA-KI reaction, and many chemical/biological reactions in atmospheric pressure plasma applications.
In addition to the quantitative calibration of the absorbance spectra peak, the effect of
the sample temperature must be carefully considered. According to a previous study,31 red
PVA-KI samples easily lose their color when heated over 50°C. This was confirmed in ourplasma experiment. Therefore, PVA-KI is beneficial in plasma experiments for biologicaltargets, since a small amount of heat has a large effect on bacteria or cell activity. PVA-KI
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FIG. 7: Effect of OH radical scavenger ethanol; increasing ethanol content in PVA-KI eliminatescolor change by plasma irradiation and corresponding increase in absorbance becomes minimal:(a) photo image of PVA-KI samples with/without ethanol; (b) absorbance peak as the functionof mixed ethanol

can be applied to investigate the behavior of ROS produced by plasma under low-temperature conditions without thermometers. Further research is required to examine the effectof temperature on the PVA-KI reaction under controlled temperature conditions.
V. CONCLUSION
Atmospheric pressure plasma jets react with liquid or water molecules to generate free
radicals, including ROS and RNS. Types and amounts of these radical species are receiving widespread attention in biomedical and sterilization applications. In this work,
PVA-KI is introduced as a new chemical probe to study free radicals. The PVA-KI color
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reaction depends on the sample condition, particularly temperature. Although the detailed
mechanism requires further study, an experimental protocol for applying PVA-KI is established in this work. The color reaction is suppressed or reversed by minimal heating.31
Therefore, measurement of the radical production potential of biological samples byplasma irradiation is a good application target for PVA-KI. In this application, we can exclude the effects of excess heat and observe only the color change or absorbance spectra.
This is one significant merit of PVA-KI over the conventional KI starch chemical probe.
ACKNOWLEDGMENTS
One of authors (H.M.) thanks Prof. Sunagawa of Fukui University for important information on PVA-KI. This work was partially supported by the ZE Research Program, IAE(ZE2021B-27), Kyoto University, and the Joint Usage/Research Program, cLPS(21020),
Nagoya University.
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