Magnetic Resonance Imaging of Electrolysis.

W2013502821 · 수집 2026-10-04 18:12:19

T10409 Fuel Cells and Related Materials

CoT 라벨링

라벨링 전

PDF 파일

자동 다운로드한 PDF PDF 보기

기본 정보

DOI 제목 발행년도 저널 URL 페이지 열기 PDF URL PDF 열기 발행기관 피인용수 라이선스

추가 정보 초록 본문

### OPEN

SUBJECT AREAS:

MAGNETIC RESONANCE

IMAGING

BIOMEDICAL ENGINEERING

Received
14 July 2014

Accepted
6 January 2015

Published
9 February 2015

Correspondence and

requests for materials

should be addressed to

[M.H. (mhjouj@](mailto:mhjouj@hotmail.com)

[hotmail.com; mhjouj@](mailto:mhjouj@hotmail.com)

[hpro.alquds.edu)](mailto:mhjouj@hpro.alquds.edu)

- These authors

contributed equally to

this work.

## Magnetic Resonance Imaging of Electrolysis.

Arie Meir 1 *, Mohammad Hjouj 2 *, Liel Rubinsky 3 & Boris Rubinsky 1,3

1Graduate Program in Biophysics, University of California Berkeley, Berkeley, CA 94720, 2Medical Imaging Department; Faculty of
Health Professions, Al-Quds University/Abu Dies/Jerusalem, 3 Department of Mechanical Engineering, University of California
Berkeley, Berkeley, CA 94720.

This study explores the hypothesis that Magnetic Resonance Imaging (MRI) can image the process of
electrolysis by detecting pH fronts. The study has relevance to real time control of cell ablation with
electrolysis. To investigate the hypothesis we compare the following MR imaging sequences: T1 weighted,
T2 weighted and Proton Density (PD), with optical images acquired using pH-sensitive dyes embedded in a
physiological saline agar solution phantom treated with electrolysis and discrete measurements with a pH
microprobe. We further demonstrate the biological relevance of our work using a bacterial E. Coli model,
grown on the phantom. The results demonstrate the ability of MRI to image electrolysis produced pH
changes in a physiological saline phantom and show that these changes correlate with cell death in the E. Coli
model grown on the phantom. The results are promising and invite further experimental research.

n a biomedical context, electrolysis refers to the passage of low magnitude, direct current between two
electrodes through a living tissue. Electrolytic ablation is the application of electrolysis for inducing irreversible
damage to undesired tissues, and research into the biochemical/biophysical aspects of tissue electrolysis can be
# I
traced to the 19 th century 1 . In modern medicine, the tools available for tissue ablation are based on various
biophysical and biochemical processes. Thermally based approaches include heating, cooling or freezing, while
electroporation, injection of chemical agents, sonoporation and photodynamic effects are additional examples for
more recent developments which were suggested as ways to expand the surgical arsenal. Electrolysis is a part of
this toolkit and the basic underlying mechanism revolves around the ionic species in the tissue, species, which
change into compounds that can ablate cells.

A large body of the modern electrolysis research can be traced back to the work of Nordenstrom and colleagues 2,3 . To understand the effects of electrolysis on tissue, researchers apply tools like histology and mathematical
models of the involved electrochemical processes 4–14 . Accompanying the more fundamental research efforts,
there is a growing body of clinical work, e.g. 15–18 . This work was inspired by several findings and the development
of a few research techniques: it was shown that the electrolysis induced pH changes can be used to reliably monitor
the extent of tissue ablation 19 . These findings have led to several basic studies on quantifying the process of
electrolysis through the use of transparent gels with pH dyes 11,20,21 . Expanding the view beyond electrolysis-driven
pH changes, different approaches have been developed to allow monitoring of pH levels. These include classical
instruments such as the glass electrode 22, electrochemically based tools such as cyclic voltammetry 23,24 or semiconductor based micro-electrode pH sensors 25 . These discrete pH-measuring devices typically have very good pH
resolution. However, they generate discrete data and cannot provide a continuous, spatial image of the pH
distribution. The devices are often sensitive, require recalibration and their response time, even in stirred solutions, is between 5 and 15 seconds. Furthermore, even though the probe tips can be of microscopic size (on the
order of microns), the outer diameter of the device’s body is on the order of a centimeters, which limits the spatial
resolution. To summarize, pH meters are excellent devices for measuring local pH, when the pH does not change
in time. However, in electrolytic ablation the continuous generation of electrolytic products and their diffusion
over time through the treated volume leads to changes in local pH in both time and space. This requires
developing a pH detection system that can provide a continuous temporal and spatial distribution of the pH
changes. Therefore, while pH-measuring devices can generate excellent discrete local data, they cannot be used for
continuous measurements in time (because of the long response time, tissue fouling issues and the need to
recalibrate) or in space (because of their discrete nature and size) of the process of electrolysis. The goal of this
study is to introduce a technology than can detect changes in pH in both time and space, to be used as a tool in the
context of electrolytic treatment of tissues.

Table 1 | MRI parameters used in the study

Sequence Coil TR [ms] TE [ms] FOV [mm] Slice thickness Num. slices # of excitations NSA Recon. Matrix size

T1W TSE SENSE-Pediatric Head coil 500 17 120 2 mm 4 2 512
T2W TSE SENSE-PED-HEAD 1000 100 120 2 mm 4 2 512
PD SENSE-PED-HEAD 5000 30 120 2 mm 4 2 512

Medical imaging has been one of the key innovation drivers in the
minimally invasive surgery field. Minimally invasive procedures are
now part of a common practice for ablating tissue deep inside the
body, but most of these procedures cannot be performed without
medical imaging 26 . Electrolysis is currently limited by the lack of an
effective means to monitor the extent of tissue ablation deep in the
body. During electrolysis, pH fronts caused by evolution of protons
(H 1 ) and hydroxide (OH 2 ) ions at the electrodes diffuse from the
electrodes outward 5,27 . Fundamental studies on tissue ablation by
electrolysis have shown that pH changes are indicative of electrolytic
tissue ablation 19 . It was found that pH dyes marked gels 27–29, could be
used to study, monitor and image electrolysis.

Our work draws from a 1957 paper by Meiboom, Luz and Gill
who studied proton relaxation times in water as a function of pH 30 .
During the last several decades, Magnetic Resonance Imaging (MRI)
was used to study pH changes in many bio-medical settings with
various methods and for various applications. For example, the effect
of intracellular pH, as well as blood and tissue oxygen tension on T1
relaxation in the rat brain were studied in Ref. 31. Measurements of
pH changes due to ischemia in the brain, in relation to amine and
amide protons were reported in Refs 32, 33. Measurements of pH
changes due to kidney failure with an MRI-CEST pH responsive
contrast agent, Iopamidol were presented in Ref. 34. An evaluation
of a range of MRI-active pH indicators for food applications is found
in Refs 35, 36. In Ref. 37, it was shown that calf muscle T2 changes
correlate with pH, PCr recovery and oxidative phosphorylation.
Most relevant to this study is the work of Schilling et al, who have
found that changes in intracellular pH, affect the relaxation time of
T2 in brain tissue 38 . While an abundance of papers exist on the use of
MRI for detecting changes in pH, no publication on the use of MRI
for monitoring the development of pH fronts during electrolysis was
found by our literature search. The primary goal of this study is to
explore the hypothesis that pH fronts, produced by electrolysis, can
be detected with MRI, for possible application in monitoring and
controlling cell ablation with electrolysis. As briefly illustrated by the
above literature citations, there are many methods that could be used
for detecting pH changes in tissue with MRI. Inspired by references 30,38, we chose to explore our hypothesis with basic T1 weighted
and T2 weighted based sequences for water.

To explore the hypothesis we conduct an experimental study using
a pH dye stained physiological saline agar-gel based phantom as a
model for a living tissue, from an electrochemical standpoint. The
pH dye generates a continuous temporal and spatial image of the
effected region, but is restricted to use in transparent systems. In
addition we have collected discrete measurements using a pH probe

- with the limitations of these measurements in mind. In the study
we compare images obtained with MRI to optical images acquired
using pH-sensitive dyes and the discrete pH data. The MRI imaging
sequences used were T1 weighted (T1W), T2 weighted (T2W) and
Proton Density (PD). The optical images were acquired using pHsensitive dyes embedded in the agar phantom that is exposed to
electrolysis. In addition to validating the MRI-based approach using
pH-sensitive dyes, we demonstrate a biological application of our
MRI work by comparing a spatial map of bacterial viability exposed
to electrolysis with the MRI image of the phantom during electrolytic
treatment. Our results are promising, and invite further experimental
explorations

Results
MRI Experiment Results. As described in our methods, the agar
plates were scanned before the administration of electrolytic
treatment with the following sequences: T1W, T2W and PD. Then
the gel plates were electrolytically treated for 15 minutes using three
different voltages: 3 V, 6 V, and 9 V. After administering the
treatment, the plates were immediately positioned in a pediatric
head coil, and inserted into the MR scanner. MR sequences with
the same pre-treatment parameters were then acquired. The MRI
parameters are presented in Table 1. To facilitate comparison of
results, figure 1 brings together images obtained for the three
voltages and the three MRI sequences. The three columns are for
the voltages of 3 V, 6 V and 9 V, from left to right, respectively. The
rows from top to bottom are for the following sequences: T1W, T2W,
and PD, respectively. All the images are for a standard Petri dish with
the same diameter, 8.5 cm. The electrolysis was administered via the
same device, positioned at the same place for all the experiments, as
constrained by the application rig in figure 2a. The position of the
electrodes can be seen in some images as the two black traces (void of
signal) at the centerline of the Petri dish. The distance between the
electrodes was 2 cm. In all the images the anode is on the left and the
cathode is on the right.

The first row of figure 1 shows images taken with the T1W
sequence. The signal from the treated volume is isointense to hypointense. It is isointense for the lower voltage of 3 V and becomes
slightly hypointense with the increasing applied voltage. The pH
change front is barely distinguishable for the 9 V treatment. The
second row shows results obtained with the T2W sequence. The
margin of the electrolysis-affected region is marked with dotted yellow line. Hypointense signal can be seen in the treated region, with
the signal intensity decreasing with increasing voltage. The affected
region near the anode is larger than near the cathode. The altered pH
front appears diffused in the anode-affected region and well-delineated in the cathode affected region. The interface between the
cathode affected region and the anode affected region is distinct
and visible. It is also note-worthy that in the cathode-affected region,
the intensity adjacent to the cathode decreases with increasing voltage. The images produced with PD sequences, presented in the third
row, show a generally similar pattern to that described for the T2W
sequence produced images. Images produced with the PD sequence
show a hypo intense signal with lower intensity relative to the T2W
sequence produced images.

pH Dye Experiment. For the pH dye experiments we have infused
the agar gel phantom described in our methods with two pH sensitive
indicator dyes. Figure 3 shows results obtained from the pH dye
experiments. To facilitate the comparison of results, Figure 3
brings together images obtained for the three voltages and results
from the two-pH dyes infused gels. The three columns are for
voltages of 3 V, 6 V and 9 V, from left to right, respectively. The
first row shows results obtained with phenolphthalein staining. The
phenolphthalein stain produces a distinct pink color in the pH range
from 8.2 to 12. The first row shows, as expected, an impression in
only the cathode region on the right. The margins of the marked
regions indicate a minimal pH of 8.2. For a voltage of 3 V, the change
of pH front takes a circular shape, most likely of a pH of 8.2.
Increasing the voltage increases the size of the change in pHaffected area Similar to the MRI images the cathode affected front

Figure 1 | Comparative MRI imaging results. Each row corresponds to a sequence modality (Top to bottom: T1, T2, PD). Each column corresponds to a
stimulation voltage (Left to right: 3 V, 6 V, 9 V).

collides with the anode produced front at a line between the electrode
and cathode. The outer margin of the lesion that has a circular shape
is most likely at a pH of 8.2, while the central line could be at any pH
in the range of pH 8.2 to pH 12. It is interesting to note that
immediately near the electrode for the 9 V voltage the intensity of
the image is reduced compared to a region further away from the
electrode.

The second row shows the results of pH staining using the Hagen
wide range pH testing kit. The cathodic region on the left is marked
with a distinct blue color which indicates a basic pH in the vicinity of
8.3, while the anodic region on the right is marked with pink color
which corresponds to pH level of 6.4. We have used the color-matching card provided by the manufacturer to establish the pH ranges.
For 3 V, the pH change affected regions have a circular shape.
Increasing the voltage increases the size of the affected region. Just
as for the other pH dye, and MRI images, the larger voltages lead to
colliding pH fronts, which can observed as a straight line. Several
interesting phenomena can be seen in the Hagen stained samples.
First, for voltages of 6 V and 9 V, the areas immediately adjacent to
the electrodes appear discolored relative to the surrounding stained
areas. Furthermore, on the cathode side at 6 V and 9 V there is a
drop of fluid, which was observed on the top of the gel. For 9 V, some
of the dye has leaked into this drop and stained it

pH probe experiments. To correlate the pH data captured by MRI
and the pH sensitive dyes, the pH measurements made by the pH
probe are presented in Table 2 and visualized graphically in the
fifth row of Figure 4. The discrete pH measurements are presented
on a single chart for each voltage. The effect of growing voltage is
clearly seen: the extreme value of pH reached on both fronts is
more deviant from the baseline normal pH. The data is presented
in two forms, with the measurements made from left to right and
from right to left. The precision of the pH reading is 0.1 and the
precision of the data point location is 1/20.5 mm. The difference
between the left to right measurement curve and the right to left
measurement curve is indicative of the systematic error of this
measurement.

Bacterial Viability Experiment. To demonstrate the relevance of our
work to a biological model, electrolytic stimulation was applied to an
agar dish plated with E. Coli bacteria. The third row in figure 3 shows
optical images of a bacterial viability pattern after treatment with 3 V,
6 V and 9 V for 15 minutes, captured using a digital camera after
24 hour growth period. The anodic region on the right is marked
with a clear bactericidal region increasing in area with increasing
voltage. The cathodic region on the left is significantly smaller in
terms of bactericidal area and is barely observable in the 3 V image

Figure 2 | Experimental setup and control images for different experimental modalities used in this work. (a) Experimental setup, (b) T1 control study,
(c) T2 control study, (d) PD control study (e) Phenolphtalein 1%, (f) Hagen wide range pH indicator dye, (g) E. Coli control study.

Discussion
As mentioned in the introduction, our hypothesis that MRI can be
used to image the pH fronts during electrolysis is based on reports in
refs 30 and 38. As can be observed in the T1W images (Figure 1 first
row), the treated volume exhibits hypointense to isointense signal,
which indicates that the effect of electrolysis is minimal on T1W
signal. A T1-weighted sequence produces an image where the signal
contrast is determined by the differences in T1 relaxation times. The
tissue signal in a T1 weighted imaging mode is inversely proportional
to its T1 relaxation time. A short echo time (TE) is used to minimize
T2-weighting together with a short repetition time (TR). A T1weighted image is typically characterized by dark fluid signal due
to the long T1 relaxation time of water. This result is consistent with
the original findings of Meiboom, Luz and Gill who studied proton
relaxation times in water as a function of pH 30 and show that T1 in
water does not change in the range of from pH 2 to pH 12,

Visible changes are produced by electrolysis in T2 weighted
images (Figure 1, second row). In the T2-weighted imaging mode,
the signal contrast is determined by differences in T2 relaxation
times. The tissue signal in a T2 weighted image is proportional to
its T2 relaxation time. A long repetition time (TR) is used to minimize T1-weighting together with a long echo time. The results in
figure 1 are also consistent with the original findings of Meiboom,
Luz and Gill 30 . Their findings show that T2 in water is strongly
affected by changes in pH and it increases symmetrically around
pH 7 with an increase and decrease in pH. Shilling et al. 38 provide
a mechanistic explanation for their observed changes in T2 with
changes in pH in the brain which is consistent with the findings of
Meiboom et. al. by noting that ‘‘The effect of pH on spin-spin relaxation time (T2) might be explained by the fact that at pH 7.0, i.e., in
the neutral environment, water molecules build larger hydrogenbound mediated clusters than in the acid or base ranges. The reduced
mobility leads to a prolonged correlation time for the dipolar interactions, which leads to a shortening of T2’’ 39 . Figure 1 shows that the
electrolysis affected region near the anode is larger than that near the
cathode. This difference makes physical sense and can be attributed
to the relative radii of protons (H 1, 0.88 fm) and hydroxide ions
(OH 2, 110 pm). Due to their relative smaller size, the protons are
more mobile hence contributing to a larger extent to the conductivity

increase around the anode. The increased mobility causes the pH
region around the anode to be larger than around the cathode.

Proton Density (PD) is defined as the number of proton spins per
unit volume of a tissue. Proton density may differ from the true water
content due to short T2 components, which are not seen in MRI. So
PD-weighted imaging where the T1 and T2 effects are minimized
leads to images whose contrast is determined primarily by the spin
(proton) density. This requires a short TE and long TR. In figure 1,
the third row shows the process of electrolysis generated PD MRI
images, which correspond well with the T2W images. This confirms
that the observed images are related to the electrolysis caused diffusion of protons and hydroxide ions.

In figure 3, the first and the second rows show the results of the
process of electrolysis obtained with pH stained dyes. While the
optical pH results cannot be quantitatively compared with the MR
images, because the pH dyes have a restricted range, both the MRI
and pH dyes images show similar phenomena and trends. The
observed affected zone increase in both modalities with an increase
in voltage, which is consistent with the increased production of electrolytic compounds with voltage. The anode and cathode electrolysis
affected regions meet at the same location in both the pH dye images
and the MRI images. The pH dye results, in particular the second row
of figure 3, show some additional interesting physical phenomena.
The effect relates to the observed drops of water on the surface of the
gel, during electrolysis with 6 V and 9 V. It is known that in an
electric field, water moves by electro-osmosis from the anode to
the cathode 6 . Therefore, during electrolysis, the gel near the anode
tends to dehydrate while water accumulates near the cathode. This is
the source of the water observed in the second row of figure 3. We
tentatively suggest that this electro-osmotic migration of water is
responsible for the discoloration adjacent to the cathode observed
with both MRI and pH dyes.

In figure 3, row three shows viability results from electrolysis
treated E. Coli, grown on the surface of the gel. This part of the work
is clinically relevant because electrolysis is becoming an important
method for sterilizing surfaces and wounds, considering the growing
antibiotic resistances of microorganisms 40 . The pattern of cell ablation observed here is consistent with electrolytic ablation and further
supports the idea that the MRI detected changes are relevant to

Figure 3 | Comparative pH dyes and bacterial viability results. Each row corresponds to a control modality (Top to bottom: Phenolphtalein 1% pH
indicator,Hagen pH indicator, E. Coli bacterial viability). Each column corresponds to a stimulation voltage (Left to right: 3 V, 6 V, 9 V).

electrolysis. Figure 3, row three shows that the extent of cell ablation
increases with the voltage and charge delivered as expected from a
pH ablation process driven by electrolysis. It is also well established
that the electrolytic products of the anode are more effective at cell
ablation than the products of the cathode 6 . This is also confirmed in
this study, which shows a much larger ablation zone near the anode
than near the cathode.

In Figure 4, a comparison between MRI images, pH dye based
images, discrete pH measurements and bacterial viability data shows
that all the experiments, produce qualitatively similar results with
respect to the effect of voltage on the affected area and with respect to
the difference between the anodic and cathodic regions. A quantitative comparison is not possible because the pH dyes and the bacterial
viability images represent limited ranges of pH while the discrete pH

measurements suffer from systematic errors. However, the results
from the different imaging techniques show that increasing the voltage (charge delivered) increases the affected area in both the anode
and cathode affected volume, the anodic front advances faster than
the cathodic front, and the anode and cathode affected regions meet
on the line perpendicular to the line connecting the electrodes. Local
pH measurements presented in the fifth row of Figure 4 are consistent qualitatively with the other means of pH monitoring, as they
indicate more extreme pH ranges for higher voltages.

Figure 4 was brought here to summarize the results. The first row
shows the T2W MR image onto which we have superimposed the
outline of the pH dye image (rows two and three) and the outline of
the viability experiment (row four). It is interesting to notice that the
interface between the anode and the cathode affected zones lie on the

Table 2 | pH measurements taken by the micro pH probe. Each voltage contains RL and LR columns: measurements taken from right to left
and left to right respectively

Distance from edge[mm] 3 V RL 3 V LR 6 V RL 6 V LR 9 V RL 9 VLR

0 7.94 7.7 9.05 7.35 9.08 7.15
0.5 7.89 7.55 8.98 7.09 8.98 6.57
1 7.86 7.25 7.92 7.15 8.46 6.7
1.5 7.83 7.32 7.06 3.98 8.06 3.35
2 7.55 7.3 6.3 2.47 7.82 2.53
2.5 6.55 4.6 5.47 1.92 7.8 2.19
3 6.43 3.97 5.29 1.98 8.15 1.87
3.5 8.72 5.87 5.4 1.93 11.45 1.81
4 9.19 6.67 10.6 10.25 12.52 11.53
4.5 9.84 7.59 12.17 11.7 12.45 11.63
5 9.95 9.96 12.15 11.84 12.46 11.63
5.5 8.10 9.02 11.98 11.73 12.31 11.57
6 8.01 7.81 11.15 11.36 11.58 11.29
6.5 7.88 7.63 8.1 11.02 9.91 9.57
7 7.8 7.26 8.07 9.65 9.41 7.95
7.5 7.6 7.1 8.00 9.3 8.7 7.71
8 7.35 7.04 7.97 9.34 n/a n/a

same line in the MRI image and the pH dye image – suggesting that
they both represent the same phenomenon. The overall shape of the
pH dye image is similar to the MRI image. The affected zone
observed with MRI is larger than that observed with dyes, because
the range of changes that can be observed with MRI is not restricted
by a certain pH dye value. The extent of cell ablation is substantially
less than the extent of the region in which MRI detects changes in pH.
In the past, studies on the effect of electrolysis on cell death were
carried out using pH probes or pH dyes. This study suggests that MRI
could become a useful tool in fundamental research on the effect of
electrolysis on cells.

To address the temporal resolution of the MRI based approach
and its relevance in the context of a typical length of an electrolysis
procedure, we’d like to note that the resolution is limited by the time
it takes to run a single scan which is given by N?Np?Tr where N
represents the number of signals averaged (NSA excitations), Np
represents the size of the encoding matrix and Tr represents the time
to repetition. When multiple echoes are acquired after each excitation, this allows for accelerating the scan time and occasionally is
referred to as the Turbo Spin Echo (TSE) or Turbo Factor. In such a
case, the time given above should by divided by the Turbo Factor. For
a typical choice of parameters, e.g. in our case study, the scan time
falls in the range of 2–4 minutes. This is shorter than a typical length
of an electrolytic treatment procedure and is much shorter than the
time it would take to generate a complete scan with discrete pH
measurements. This further suggests that MRI could be a potential
method to study electrolysis in tissue.

Methods
Experimental Outline. The experiment was designed to allow for a comparison
between different images of pH fronts produced by the electrolysis of a physiological
saline solution phantom. The images were generated by various MRI sequences and
compared with: a) optical images acquired using pH-sensitive dyes embedded in a
physiological saline agar solution phantom treated with electrolysis and b) bacterial E.
Coli model, grown on a phantom and treated by applying the same electrolysis
protocol. Each experimental study was done separately. In addition we have also
collected a set of discrete measurements using a micro pH probe.

Tissue model. Our tissue model consists of a physiological saline based agar gel
phantom with electrical conductivity designed to simulate that of a tissue. To
construct the phantom, 1% Bacto-Agar (Fisher Scientific) was mixed with 0.9 g/l
Sodium Chloride (Fisher Scientific) in distilled water. The solution was then brought
to a boil and poured into 85 mm diameter Petri dishes. The same dish dimension was
used in all the studies. The conductivity of the agar phantom was measured to be
approximately 0.14 S/m which is close to the range of hepatic tumor conductivity 41 .

Experimental procedure. The experimental setup is shown in Figure 2.a. We have
used two disposable graphite electrodes made of pencil lead (Pentel super HB

0.7 mm), to avoid contamination with products of electrolysis from the electrodes.
The electrodes, mounted in a horizontal holder were placed perpendicularly to the gel
phantom in the Petri dish. The electrodes were inserted 7 mm deep into the gel at a
distance of 2 cm between their centers. The electrodes are connected to constant
voltage batteries. We used 3 V, 6 V and 9 V batteries. The electrolysis process lasted
15 minutes. While typical electrolysis stimulation is administered using a fixed
current source, we have used a fixed voltage source and have taken current
measurements during the procedure (data not shown) for charge dosage estimation
purposes. The overall delivered charge dosages over the 15 minutes stimulation
period were approximately 0.9 C, 1.8 C and 2.9 C, for 3 V, 6 V and 9 V, respectively.
These charge dosages fall within the range of a typical electrolytic ablation therapy
stimulation 3,42 . Identical experiments were done separately for MRI imaging, pH dyes
based optical imaging, discrete pH measurements and the E. Coli viability model.

MRI Experimental Model. The phantom models were scanned, before and after
electrolysis in a clinical 1.5 T MRI system (Philips Achieva SE) using a SENSE
pediatric coil. The specific MRI parameters of each sequence are listed in Table 1. The
mean acquisition time for each sequence was 3 minutes. Images shown in the paper
were taken in the following order: T1W, T2W and PD. For later comparison with the
MR images produced after electrolysis, figures 2.b, 2.c and 2.d show the baseline
images for the sequences introduced above, respectively. In repeated experiments
(results not shown) we changed the order and the time after the end of electrolysis at
which the various sequences were taken and found no measurable effect of the time at
which the images were acquired on the dimensions of the affected region.

pH-Sensitive Dye Model. As a control study, we used 2 distinct, pH sensitive dyes in
order to estimate the boundary of the region where the pH has changed due to
electrolysis. The dyes employed were 1) Phenolphtalein 1% by Sigma (turns pink in
pH range of 8.2–12) and 2) Nutrafin pH wide range by Hagen (indicates pH by color
in the range of 4.5–9). The dye manufacturer instructions prescribed the
concentration of the last dye. We used a digital camera (Casio Exilim EX-ZR100) to
acquire optical images of the experimental chamber before and after electrolysis and
correlate these images with the images acquired using MRI and bacterial viability. For
later comparison with images produced after electrolysis, figures 2.e and 2.f, show the
baseline images prior to electrolysis for the two dyes used, respectively.

Discrete measurements with micro pH probes. To produce numerical values for pH
we have used a MI-4146 Micro-combination pH probe (Microelectrodes, Inc. 40
Harvey Rd., Bedford, NH) with a response time of 5 seconds to 15 seconds in a stirred
solution and a pH meter pH150 Oakton (625 East Bunker Court., Vernon Hills, IL).
The experimental procedure is similar to that used in the MRI study and the pH
sensitive dye study. We have marked a line through the diameter of the Petri dish and
marked off every 5 mm (see Fig 1.h). The two electrodes were placed at a distance of
2 cm between them. Similarly to the previous experiments we applied voltages of 3 V,
6 V and 9 V for 15 minutes. After the electro-stimulation, the electrodes were
removed, and pH measurements were made, starting with the marking furthest away
from the dish center. To minimize and evaluate the effect of diffusion on the
systematic errors of this method, in one set of the experiments the reading was
performed from left to right, and in another set of experiments the reading was
performed from right to left. The pH probe was placed over the mark and into the gel,
making certain that the tip was covered. We waited until the reading was stabilized,
usually between 15 and 25 seconds. The reading was recorded and the pH probe
flushed and moved over to the next mark until the entire dish was transversed. One
transverse reading took about 20 to 25 minutes. (No new electrolytic products were
produced during the measurement period however the pH changed during the

Figure 4 | Comparative pH dyes and bacterial viability results. Each row corresponds to a control modality (Top to bottom: Phenolphtalein 1% pH
indicator,Hagen pH indicator, E. Coli bacterial viability, Local pH measurements taken by pH probe). Each column corresponds to a stimulation voltage
(Left to right: 3 V, 6 V, 9 V).

measurements because of diffusion). While the actual data recorded is with a
precision of 0.1, the accuracy in relation to the colorimetric dye and MRI
measurements is low. Therefore, the discrete pH measurement should be viewed as a
rough indication of the actual values. We also considered measuring pH during the
electro-stimulation process, however, we found, in other studies, that the insertion of
an active pH probe during electrolysis distorts the reading because of the effect of the
current on the pH probe.

Bacterial Model. Lyophilized E. Coli of HB101 strain (BioRad) were grown in LB
broth overnight and plated on LB broth based agar gel filled petri dishes. The LB broth
for the overnight growth consisted of 1% BactoTryptone (BD), 0.5% Yeast Extract
(BD), 1% NaCl (Sigma Aldrich) and 1.5% Agarose (Sigma Aldrich). 6 mm glass beads
(Sigma Aldrich) were used for plating to ensure uniform coverage. After plating, the
beads were removed and the plates were incubated for 15 minutes at 37u. The
conductivity of the gel was measured around 0.2 S/m. After electrolysis the Petri
dishes were covered and incubated for 24 hours. We used a digital camera (Casio
Exilim EX-ZR100) to acquire optical images of the areas where bacterial growth was
inhibited and correlated these images with the images acquired using MRI and gels
with pH dyes. For later comparison with images produced after electrolysis, figure 2.g
shows the baseline images for an untreated with electrolysis, cell growth plate.

Conclusion
In summary, we report experimental findings that support the hypothesis that electrolysis induced pH-changes can be detected with
MRI. Our results indicate the feasibility of using MRI as a means
to monitor dynamic changes in local pH level of a biological sample
during an electrolysis process. Our work uses an agar-based gel
model with conductivity in the range of a biological tissue, and is
validated vs. optical images utilizing pH indicator dyes. In addition
we demonstrate the relevance of our work in the biological context by
correlating bacterial viability data with MRI measurements. It may
be interesting to work on developing different MRI techniques for
detecting pH, using MRI markers. Reference 30, suggests that it may
be also possible to develop MRI sequences that detect discretely
various ranges of pH, because T2 seems to be very sensitive to pH.
Our results are promising, and invite further experimental explorations on the use of MRI for studying electrolysis and in clinical
practice.

1. Amory, R. A treatise on electrolysis and its therapeutical and surgical treatement
in disease. (William Woof & Co., 1886).
2. Nordenstrom, B. E. W. Preliminary clinical trials of electrophoretic ionization in
the treatement of malignant tumors. IRCS Medical Sc. 6, 537 (1978).
3. Nordenstrom, B. E. Electrochemical treatment of cancer. I: Variable response to
anodic and cathodic fields. Am J Clin Oncol 12, 530–536 (1989).
4. Nilsson, E., Berendson, J. & Fontes, E. Development of a dosage method for
electrochemical treatment of tumours: a simplified mathematical model.
Bioelectroch Bioener 47, 11–18, doi:10.1016/s0302-4598(98)00157-3 (1998).
5. Nilsson, E., Berendson, J. & Fontes, E. Electrochemical treatment of tumours: a
simplified mathematical model. J Electroanal Chem 460, 88–99, doi:10.1016/
s0022-0728(98)00352-0 (1999).
6. Nilsson, E. et al. Electrochemical treatment of tumours. Bioelectrochemistry 51,
1–11, doi:10.1016/s0302-4598(99)00073-2 (2000).
7. von Euler, H., Nilsson, E., Olsson, J. M. & Lagerstedt, A. S. Electrochemical
treatment (EchT) effects in rat mammary and liver tissue. In vivo optimizing of a
dose-planning model for EChT of tumours. Bioelectrochemistry 54, 117–124,
doi:10.1016/s1567-5394(01)00118-9 (2001).
8. von Euler, H., Nilsson, E., Lagerstedt, A. S. & Olsson, J. M. Development of a doseplanning method for electrochemical treatment of tumors: A study of mammary
tissue in healthy female CD rats. Electro Magnetobiol 18, 93–1 doi:10.3109/
15368379909012903 (1999).
9. Bergues Pupo, A. E., Bory Reyes, J., Bergues Cabrales, L. E. & Bergues Cabrales, J.
M. Analytical and numerical solutions of the potential and electric field generated
by different electrode arrays in a tumor tissue under electrotherapy. Biomed Eng
Online 10 doi:10.1186/1475-925x-10-85 (2011).
10. Placeres Jimenez, R. et al. 3D Stationary Electric Current Density in a Spherical
Tumor Treated With Low Direct Current: An Analytical Solution.
Bioelectromagnetics 32, 120–130, doi:10.1002/bem.20611 (2011).
11. Turjanski, P. et al. pH front tracking in the electrochemical treatment (EChT) of
tumors: Experiments and simulations. Electrochimica Acta 54, 6199–6206,
doi:10.1016/j.electacta.2009.05.062 (2009).
12. Camue Ciria, H. M. et al. Antitumor effects of electrochemical treatment. Chn J
Cancer Res 25, 223–234, doi:10.3978/j.issn.1000-9604.2013.03.03 (2013).
13. Yoon, D.-S. et al. Introduction of electrochemical therapy (EChT) and application
of EChT to the breast tumor J Breast Canc 10 162 168 (2007)

14. Czymek, R. et al. Electrochemical Treatment: An Investigation of Dose-Response
Relationships Using an Isolated Liver Perfusion Model. Saudi J Gastro 17,
335–342, doi:10.4103/1319-3767.84491 (2011).
15. Griffin, D., Dodd, N., Zhao, S., Pullan, B. & Moore, J. V. Low-level direct electrical
current therapy for hepatic metastases. I. Preclinical studies on normal liver. Brit J
Cancer 72, 31–34 (1995).
16. Griffin, D., Dodd, N. J., Moore, J. V., Pullan, B. & Taylor, T. The effects of low-level
direct current therapy on a preclinical mammary carcinoma: tumour regression
and systemic biochemical sequelae. Brit J Cancer 69, 875–878 (1994).
17. Miklavcˇicˇ, D. et al. Tumor treatment by direct electric current-tumor temperature
and pH, electrode material and configuration. Bioelectrochem Bioenerget 30,
209–220 (1993).
18. Miklavcˇicˇ, D., Fajgelj, A. & Sersˇa, G. Tumor treatment by direct electric current:
electrode material deposition. Bioelectrochem Bioenerget 35, 93–97 (1994).
19. Finch, J. G. et al. Liver electrolysis: pH can reliably monitor the extent of hepatic
ablation in pigs. Clin Sci 102, 389–395 (2002).
20. Olaiz, N., Suarez, C., Risk, M., Molina, F. & Marshall, G. Tracking protein
electrodenaturation fronts in the electrochemical treatment of tumors.
Electrochem Commun 12, 94–97, doi:10.1016/j.elecom.2009.10.044 (2010).
21. Ivic, M. L. A., Perovic, S. D., Zivkovic, P. M., Nikolic, N. D. & Popov, K. I. An
electrochemical illustration of the mathematical modelling of chlorine impact and
acidification in electrochemical tumour treatment and its application on an agaragar gel system. J Electroanal Chem 549, 129–135, doi:10.1016/s00220728(03)00251-1 (2003).
22. Moore, E. W. Determination of pH by the glass electrode: pH meter calibration for
gastric analysis. Gastroenterology 54, 501–507 (1968).
23. Takmakov, P. et al. Characterization of local pH changes in brain using fast-scan
cyclic voltammetry with carbon microelectrodes. Anal Chem 82, 9892–9900
(2010).
24. Venton, B. J., Michael, D. J. & Wightman, R. M. Correlation of local changes in
extracellular oxygen and pH that accompany dopaminergic terminal activity in
the rat caudate–putamen. J Neurochem 84, 373–381 (2003).
25. De Beer, D., Schramm, A., Santegoeds, C. M. & Kuhl, M. A nitrite microsensor for
profiling environmental biofilms. Appl Environ Microbiol 63, 973–977 (1997).
26. Chu, K. F. & Dupuy, D. E. Thermal ablation of tumours: biological mechanisms
and avdances in therapy. Nat Rev Cancer 14, 199–208 (2014).
27. Turjanski, P. et al. pH front tracking in the electrochemical treatment (EChT) of
tumors: Experiments and simulations. Electrochimica Acta 54, 6199–6206 (2009).
28. Turjanski, P. et al. The role of pH fronts in reversible electroporation. PLoS One 6,
e17303 (2011).
29. Maglietti, F. et al. The Role of Ph Fronts in Tissue Electroporation Based
Treatments. PLoS One 8, e80167 (2013).
30. Meiboom, S., Luz, Z. & Gill, D. Proton Relaxation In Water. J Chem Phys 27,
1411–1412, doi:10.1063/1.1744015 (1957).
31. Kettunen, M. I., Grohn, O. H. J., Silvennoinen, M. J., Penttonen, M. & Kauppinen,
R. A. Effects of intracellular pH, blood, and tissue oxygen tension on T-1 rho
relaxation in rat brain. Magn Reson Med 48, 470–477, doi:10.1002/mrm.10233
(2002).
32. McVicar, N. et al. Quantitative tissue pH measurement during cerebral ischemia
using amine and amide concentration-independent detection (AACID) with
MRI. J Cerebr Blood F Met 34, 690–698, doi:10.1038/jcbfm.2014.12 (2014).
33. Zong, X., Wang, P., Kim, S. G. & Jin, T. Sensitivity and Source of Amine-Proton
Exchange and Amide-Proton Transfer Magnetic Resonance Imaging in Cerebral
Ischemia. Magn Reson Med 71, 118–132 (2014).
34. Longo, D. L., Busato, A., Lanzardo, S., Antico, F. & Aime, S. Imaging the pH
evolution of an acute kidney injury model by means of iopamidol, a MRI-CEST
pH-responsive contrast agent. Magn Reson Med 70, 859–864, doi:10.1002/
mrm.24513 (2013).
35. Evans, S. & Hall, L. Evaluation of a range of MRI-active pH indicators using a
multiple-sample method. Aiche Journal 51, 1541–1547, doi:10.1002/aic.10387
(2005).
36. Evans, S. & Hall, L. Measurement of pH in food systems by magnetic resonance
imaging. Can J Chem Eng 83, 73–77 (2005).
37. Schmid, A. I. et al. Exercising calf muscle T-2 changes correlate with pH, PCr
recovery and maximum oxidative phosphorylation. Nmr Biomed 27, 553–560,
doi:10.1002/nbm.3092 (2014).
38. Schilling, A. M., Blankenburg, F. B., Bernarding, J., Heidenreich, J. O. & Wolf, K. J.
Intracerebral pH affects the T2 relaxation time of brain tissue. Neuroradiology 44,
968–972, doi:10.1007/s00234-002-0873-0 (2002).
39. Schilling, A., Blankenburg, F., Bernarding, J., Heidenreich, J. & Wolf, K.
Intracerebral pH affects the T2 relaxation time of brain tissue. Neuroradiology 44,
968–972 (2002).
40. Piggott, J. M., Berney, H., Hurley, E. & Clair, J. Planar flexible electrode for use in
wound sterilization. Paper presented at 24th Annual International Conference of
the Engineering in Medicine and Biology Society. Annual Fall Meeting of the
Biomedical Engineering Society, 1929–1930 vol. 1923, doi:10.1109/
iembs.2002.1053098 (2002).
41. Laufer, S., Ivorra, A., Reuter, V. E., Rubinsky, B. & Solomon, S. B. Electrical
impedance characterization of normal and cancerous human hepatic tissue.
Physiol Meas 31 995 (2010)

42. Nordenstro¨m, B. Biologically closed electric circuits: Clinical, experimental and
theoretical evidence for an additional circulatory system. (Ursus Medical AB,
1983).

Author contributions
A.M. has performed the bacterial and ph-dye experiments, prepared figures 3–4 and wrote
the paper. M.H. has performed the MRI experiments, prepared table 1, figures 1–2 and
wrote the paper. L.R. prepared table 2 and performed the micro-pH experiments. B.R. has
conceived the experiments and contributed to writing the paper.

Additional information
Competing financial interests: The authors declare no competing financial interests.

How to cite this article: Meir, A., Hjouj, M., Rubinsky, L. & Rubinsky, B. Magnetic
Resonance Imaging of Electrolysis.. Sci. Rep. 5, 8095; DOI:10.1038/srep08095 (2015).

This work is licensed under a Creative Commons Attribution 4.0 International
License. The images or other third party material in this article are included in the
article’s Creative Commons license, unless indicated otherwise in the credit line; if
the material is not included under the Creative Commons license, users will need
to obtain permissionfrom the licenseholderin order toreproduce the material. To
[view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/](http://creativecommons.org/licenses/by/4.0/)

본문 출처 PDF: PDF 보기 · 다른 PDF는 위 [PDF 업로드]로 올릴 수 있습니다.

활용가능성

이력 0건

아직 편집 이력이 없습니다. 저장할 때마다 이전 내용이 여기에 백업됩니다.