Glycative Stress Research
Online edition : ISSN 2188-3610
Print edition : ISSN 2188-3602
Received : March 5, 2022
Accepted : April 24, 2022
Published online : June 30, 2022
doi:10.24659/gsr.9.2_63

Original article

Effect of ceramic-treated water on food functionality of strawberries.
Wakako Takabe 1), Takehiro Nagata 1), Karin Nakamura 1), Mika Asano 2),
Masayuki Yagi 3), Yoshikazu Yonei 3), Yasushi Kondo 4), and Shinichi Sugiura 5)1) Department of Materials and Life Science, Faculty of Science and Technology,
Shizuoka Institute of Science and Technology, Shizuoka, Japan
2) MS Dream Co., ltd., Aichi, Japan3) Anti-Aging Medical Research Center and Glycation Stress Research Center,
Graduate School of Life and Medical Sciences, Doshisha University, Kyoto, Japan4) Ichigono Sato, Aichi Japan
5) Center for Clinical Pharmacy Education and Research, Faculty of Pharmacy,
Doshisha Women's College of Liberal Arts, Kyoto, Japan

Abstract

Aim : Glycation is a non-enzymatic reaction between reducing sugars and proteins. It forms advanced glycation end
products (AGEs) and is involved in multiple diseases. We have reported that strawberries are one of the plants that inhibit
the formation of AGEs. It has been reported that water treated with ceramic chips (ceramic-treated water) enhanced thegrowth of agricultural crops. In this study, we examined the effect of ceramic-treated water on the food functionality ofstrawberries, especially the anti-glycative effect and antioxidant efficacy.
Methods: Fourteen varieties of strawberry plants were cultivated using either normal water or ceramic-treated water, andharvested strawberries were sliced, dried, and ground. Extracts were prepared with hot water and introduced into the humanserum albumin (HSA)-Glucose glycation model. Inhibitory effects on glycation were evaluated by measuring fluorescent
AGEs, and antioxidative efficacy was measured by the DPPH method.
Results: All the strawberry samples showed anti-glycative effects and antioxidant activity. There was no significant
difference between the cultivation water in the paired t-test for either effect. However, one variety showed a significant
increase in inhibition of AGEs formation, and 4 varieties showed a significant increase in antioxidant activity in the ceramictreated water cultivation. No correlation between anti-glycative and antioxidative effects was found.
Conclusion: In this study, we examined the effect of growing water on 14 strawberry varieties. Ceramic-treated waterchanged the feature of strawberries, but the change depends on the varieties. This indicates that the effect of ceramic-treatedwater may be limited to certain varieties. In addition, the lack of correlation between anti-glycative and antioxidative effects
suggested that the substances contributing to each effect were different.

KEY WORDS: special-glaze-applied ceramic chips, anti-glycative effect, antioxidative efficacy, Fragaria × ananassa

Corresponding author: Wakako Takabe, PhD
Department of Materials and Life Science, Faculty of Science and Technology,
Shizuoka Institute of Science and Technology, Shizuoka, Japan
220 0-2, Toyosawa, Fukuroi City, Shizuoka 437-0032, Japan.
TEL & FAX: 0538-45-0164 e-mail: takabe.wakako@sist.ac.jp
Contact for Co-authors; Nagata T, 1714 024.nt@sist.ac.jp;
Nakamura K, 1814038.nk@sist.ac.jp; Asano M, info@msdream.co.jp;
Yagi M, myagi@mail.doshisha.ac.jp; Yonei Y, yyonei@mail.doshisha.ac.jp;
Kondo Y, ichigonosato1027@facebook.com; Sugiura S, ssugiura@dwc.doshisha.ac.jp_ 63 _

Glycative Stress Research 2022; 9 (2) : 63-72(c) Society for Glycative Stress ResearchEffect of Cultivation Water on Strawberries.

water treated with ceramic chips (ceramic-treated water) isused for crops, it has been shown to increase weight, tighten
flesh, and improve taste in peach cultivation; to increase theyield in rice cultivation 15). However, the effect of ceramictreated water on strawberries has not been reported.
In this study, we examined whether the use of ceramictreated water for growing 14 strawberry varieties enhanced
their food functions, specifically their anti-glycative andantioxidative effects.

Introduction

Reducing sugars, such as glucose and fructose, areessential nutrients for biological activities. However, an
excess of reducing sugars in the body reacts with proteins
non-enzymatically, forming advanced glycation end products(AGEs) 1). This reaction is called glycation. AGEs accumulate
in the body as we age and are involved in the development oflifestyle diseases such as cancer 2), diabetes 3), osteoporosis 4),
atherosclerosis 5), and Alzheimer's disease 6, 7). The concept ofglycative stress is a comprehensive view of the stress caused
by glycation and the process leading to lifestyle diseases.
The ways to inhibit glycative stress include preventing
hyperglycemia, inhibiting AGE production, inhibiting AGE
absorption, promoting AGE degradation and excretion, andinhibiting AGE-induced damage to cells and tissues. In
our previous study, we found that strawberry (Fragaria ×ananassa) is one of the plants that has the effect of inhibiting
glycation (anti-glycative effect) 8). Strawberry is a plantnative to the Netherlands, and vitamin C and polyphenols
included in its edible pseudo -fruits have been reported tohave not only antioxidative and anti-glycative effects but
also anti-inflammatory effects 8 - 11). There are many varietiesof strawberries produced by crossbreeding, and Japan is
one of the countries with the largest number of strawberryvarieties, with more than 300 varieties on the market 12).
The consumption of fruits has been reported to reduce therisk of developing type 2 diabetes 13) ; however, the fructose
contained in fruits is often considered to be related to obesity,
hyperlipidemia, and diabetes.
Special-glaze-applied ceramic chips (referred to asceramic chips) developed in 2000 have been reported to
improve water quality by decomposing odor, sterilizing, andreducing biochemical oxygen demand (BOD) 14, 15). When

Material and methods
Materials

Fourteen varieties of strawberry plants were provided
from the Center for Clinical Pharmacy Education andResearch, Faculty of Pharmacy, Doshisha Women's Collegeof Liberal Arts ( Table 1 ).
Strawberry seedlings of each variety were planted
on cultivation devices about 1 m in height in the samearrangement and irrigated with either normal water or
ceramic-treated water, which was prepared by passing
normal water once through a metal pipe (10 cm in diameterand 100 cm in length) filled with special-glaze-applied
ceramic chips. Each plant was drip irrigated four times a
day with about 50 mL per plant ( Fig. 1 ). The irrigation
conditions were controlled to be the same.
Human serum albumin (HSA) was purchased from
Sigma-Aldrich (St. Louis, MO). DPPH, 2, 2- diphenyl-1(2, 4, 6-trinitrophenyl) hydrazyl (DPPH) free radical and2- ( N-morpholino) ethanesulfonic acid (MES) were obtained
from Tokyo Chemical Industry (Tokyo, Japan). All otheranalytical-grade reagents were obtained from Fujifilm Wako
Chemicals (Osaka, Japan).

Table 1. The cross-fertilization of starawberries
variety

the cross-fertilization of strawberry varieties

Ai-berryundisclosed

Asuka-rubyNyohouAkihime
Amaka

KaorinoNyohouAmaou

Kuno-wase

Asuka-wave

Kiyokabreeding lines(0028401)(breeding among Nyohou Ai-berry
Tochiotome Sanchigo)

KiyokaNyohouAi-berry

NyohouHarunokaDana

Koiminori
Benihoppe

Hoshinokirameki

Toukunbreeding lines (03042-08)
AkihimeSatinoka

Reikouundisclosed( Tonehoppe

Red-pearlAi-berry

Tochiotome)

Mie - bohon1gou

Toyonoka

Hinoshizuku

breeding lines (undisclosed)

Yayoihime

Yotsuboshi

breeding lines (0023001)

A8S4 -147

Kurume-IHIgou
Tonehoppe

Toyonoka_ 64 _

Houkou-wase

AkihimeAkasyanomitshuko

Glycative Stress Research

Watersupplytank

Watering tube
Normal water
Ceramic-treated waterSpecial ceramic filter

Fig. 1. Special ceramic chip filtration system.

Water was supplied from a common irrigation tank, one connected to normal water and the other to a
special ceramic chip filtration system. Each plant was irrigated four times a day, about 50 mL each time.

Sample preparation

Evaluation of the antioxidative efficacy

Strawberries were sliced, dried at 65 ℃ for 120 hours,
and ground. To prepare the water extract, 2 g of dried powder
was mixed with 4 0 mL of distilled water and was incubatedat 80 °C for 75 minutes. Extracted samples were centrifuged
at 3,350 × g for 10 minutes and filtered using filter paper.
Five milliliters of the extracts were evaporated to measure
the solid content, and then the extracts were adjusted to theirappropriate concentrations with distilled water.

The antioxidative efficacy of the strawberry extracts
was determined as the free radical scavenging capacity
using DPPH as described by Oki 18) with slight modification.
Specifically, 0. 5 mg/mL of the strawberry extract and 200
mol/L DPPH were reacted in 50 mmol/L 2-(N-morpholino)ethanesulfonic acid (MES) buffer (pH 6.0) for 20 minutesat room temperature. The absorbance of the solution wasmeasured at 520 nm. Trolox, an analog of vitamin E, wasused as a control.

Preparation of Glycated Proteins
An HSA glycation model was used to evaluate the effectof strawberries on glycation as previously described 16). HSA(8 mg/mL) and 0.2 mol/L glucose in 50 mmol/L phosphatebuffer (PB, pH 7.4) were incubated at 60 °C for 40 hours(solution A). Simultaneously, heated proteins without glucose
were also prepared (solution B). To determine the effects ofstrawberry extract, 30 mg/mL of the extract was added toreach 1/10 of volume concentration (final concentration;
3 mg/mL) with or without glucose ( solutions C and D,
respectively).

Measurement of AGE-derived fluorescence
After incubation, the fluorescence of the reaction mixture
was measured as previously described 17). Two hundred μL of
the reaction mixture was used to measure fluorescence at an
excitation wavelength of 370 nm and an emission wavelengthof 440 nm by Infinite M1000 (Tecan Japan, Kanagawa,Japan) microplate reader. The ratio of inhibitory effect on
fluorescent AGE formation (%) was calculated using theequation below.

The ratios of inhibitory effect of fluorescence AGEs (%)= {1 − (C − D) / (A − B) } × 100

Statistics
Data are expressed as mean ± standard deviation (SD)of at least three independent experiments. The statisticalanalyses performed by analysis of variance (ANOVA) were
subjected to Dunnett’s test for multiple comparisons between
each of the samples and control groups. To compare the same
varieties of samples that were cultivated using either theceramic-treated water or normal water, we conducted paired
t-tests. Differences were considered significant at p-values
less than 0.05.

Results

Calculation of solid concentration of hot water
extract of strawberry.
The solids concentration of the hot water extract ofeach strawberry variety is shown in Table 2. There was nodifference in the solid concentration among the varieties or
the water used for cultivation (mean value of 36.3 mg/mL
for ceramic-treated water cultivation, 36.4 mg/mL for normal
water cultivation).

_ 65 _

Effect of Cultivation Water on Strawberries.

Table 2. Solid content of hot water extract of samples
varietysolid content [mg/mL]
Ceramic-treated water

Normal water

38.0

38. 2

35.7

36.1

Kiyoka

36.0

36. 3

Nyohou36.6

36.8

Hoshinokirameki

38. 2

38.9

Yayoihime

36.0

36.6

Red-pearl

35. 5

35.0

average

36. 3

36.4

Ai-berry
AkihimeAsuka-ruby
Amaka

KaorinoKoiminori
Benihoppe
ToukunYotsuboshi

36. 5

34.4

35. 2

35. 5

37.1

36.1

36.9

38.6

36.8

33. 5

31.1

37.8

Effect of cultivation with ceramic-treated water on
the inhibition of fluorescent AGE formation in hot
water extracts of strawberry.
The results of the inhibition of fluorescent AGE
formation by the hot water extract of strawberry at a final
concentration of 3 mg/mL are shown in Table 3. All 14
strawberry varieties significantly suppressed the formationof fluorescent AGEs regardless of the cultivation water.
The highest suppression rate of fluorescent AGE formation
was observed in Koiminori with ceramic-treated water
cultivation (92.3 ± 1.77 %) and the lowest in Benihoppe withnormal water cultivation (69.7 ± 3.80 %). Data were analyzed
by a Student’s t-test in each variety, only Koiminori showeda significant increase in the inhibition of fluorescent AGE
formation by water cultivation with ceramic chips (92.3 ±
1.77% for water cultivation with ceramic chips, 87.8 ± 1.37%
for water cultivation with normal water, p = 0.024). Student’s
paired t-tests showed no significant difference between watertypes (78.9 ± 6.47 % [ceramic - treated water cultivation],
79.8 ± 5.16 % [normal water cultivation], p = 0.430, Fig. 2).
Comparing 14 varieties of samples grown with normal water,
8 of them, namely, Yotsuboshi, Akihime, Ai-berry, Amaka,Nyohou, Toukun, Yayoihime, and Benihoppe, showedsignificantly lower levels of inhibition of fluorescent AGE
formation than Koiminori (87.8 ± 1.37 %, Fig. 3 ). This
indicates that the effect of cultivation water on the inhibitionof fluorescent AGE formation in strawberries is smaller thanthe effect of differences among the varieties.

Effect of cultivation with ceramic-treated water onantioxidant activity of strawberry.
As to antioxidative efficacy, we measured the DPPH

35.7

39. 2

radical scavenging activity of hot water extracts of strawberries
cultivated with ceramic-treated water or normal water. DPPH
radical scavenging activity was observed in all 14 strawberryvarieties regardless of the water used ( Fig. 4 ). The highest
DPPH radical scavenging activity was observed in the normal
water cultivation of Toukun (117.9 ± 0.81 nmol Trolox eq./
mg-solid content), and the lowest was Amaka that cultivatedwith normal water (46.5 ± 1.47 nmol Trolox eq. /mg-solid
content). The Student’s t-test in each variety showed thatcultivation with ceramic-treated water significantly enhanced
the DPPH radical scavenging activity in Amaka, Kiyoka,
Nyohou, and Yotsuboshi, while it significantly decreasedin Hoshinokirameki and Red-pearl. Paired t-test for each
cultivation water showed no significant difference (72.1 ±16.7 nmol Trolox eq./mg-solid content [ceramic-treated water
cultivation], 69.9 ± 20.2 nmol Trolox eq./mg-solid content[normal water cultivation], p = 0.540, Fig. 5 ). Comparing
the 14 samples grown with normal water, Toukun showedsignificantly higher DPPH radical scavenging activity than
all other 13 varieties ( Fig. 6 ). This indicated that the effect of
cultivation water on the antioxidative efficacy of strawberries
was smaller than the effect of differences among thevarieties.

The correlation between anti-glycative andantioxidative effects of strawberries.
Since it is known that oxidation occurs during AGE
production, we hypothesized that strawberries with strong
anti-glycative activity would also have high antioxidant
efficacy. However, as shown in Fig. 7, the R 2 values were
− 0.0273 (ceramic-treated water cultivation) and 0.0278(normal water cultivation), respectively; therefore, nocorrelation was observed.

_ 66 _Glycative Stress Research

Table 3. The ratio of inhibitory effect on fluorescent AGE formation (%)
variety

Ceramic-treated water

Normal waterp valueAi-berry
AkihimeAsuka-ruby
Amaka
KaorinoKiyokaKoiminori
NyohouBenihoppe
Hoshinokirameki
ToukunYayoihime
Yotsuboshi
Red-pearl

76.8 ±
72.4 ±
84.8 ±
81.8 ±
78.8 ±
86.9 ±
92. 3 ±
78. 5 ±
74. 3 ±
74. 4 ±
70.3 ±
70.0 ±
80. 8 ±
82. 4 ±

77.7 ±
79.7 ±
85.1 ±
77.4 ±
80.9 ±
85.8 ±
87.8 ±
76.8 ±
69.7 ±
80.8 ±
76.1 ±
73.1 ±
80.0 ±
85.8 ±

0.782
0.0 71
0.845
0.062
0.396
0.118
0.024
0.664
0.253
0.123
0.255
0.486
0.690
0.156

3.26
2.91
1.69
2. 53
3. 09
0. 80
1.77
3.9 0
4. 56
4.91
6.67
5.74
1. 8 0
1.93

4.10
4. 28
1. 59
1. 56
2.26
0.45
1. 37
4. 97
3. 8 02. 8 4
3. 57
4. 0 42.70
2. 86

The results are expressed as mean ± SD of 3 experiments. AGE, advanced glycation end products; SD, standard deviation

The ratio of inhibitory effecton fluorescent AGE formation (%)

Ceramic- treated water
Normal waterp = 0.430

100

Fig. 2. Effect of ceramic-treated water on inhibitory
effect on fluorescent AGE formation of
strawberry varieties.

80

Fourteen variety of strawberry plants were cultivatedeither ceramic-treated water or normal water. Hot water
extracts of each strawberry variety were prepared andused to determine the inhibitory effect of fluorescent
AGE formation. Paired t-test was conducted toinvestigate the effect of cultivation water on antiglycative effect of strawberries. All data were shown as
the mean ± SD (n = 3). AGE, advanced glycation end
product; SD, standard deviation.

60
40
20
0

Ceramic-treated water

Normal water

The ratio of inhibitory effecton fluorescent AGE formation (%)

100

*

*

80

**

**

**

**

**

**

ﬁgure2

60
40
20

pe

e

op

m
B

en

ih

n

hi

ku

Ya
y

oi

u
ho

To
u

a

yo

ak

N

m

ry

i-b

A

m

A

hi
ki
A

er

e

i
sh

i

bo

ek

su

Yo
t

H

os

hi

no

ki

ra

m

rin

o

y

ao

ub

K

rl

-r

ka

A

su

-p

ea

ok

ed

K

iy

R

Ko

im

in

or

i

a

0

Fig. 3. Differences in inhibitory effect on fluorescent AGE formation between strawberry varieties.

Fourteen variety of strawberry plants were cultivated using normal water. Hot water extracts of each strawberry variety wereprepared and used to determine the inhibitory effect of fluorescent AGE formation. All data were shown as the mean ± SD (n =ﬁgure33). * p < 0.05 and ** p < 0.01 vs. Koiminori. AGE, advanced glycation end product; SD, standard deviation.

_ 67 _

Effect of Cultivation Water on Strawberries.

Ceramic- treated water

140
120

**

80

**

**

*

**

100

**

60
40
20

rl

i

ea
-p

ed
R

su

bo

m
hi

Yo
t

oi

sh

e

n
ku

Ya
y

os

hi

To
u

ek

no

en

ki

ih

ra

m

op

i

pe

u
ho
yo

B

Ko

N

im

in

or

i

a
ok
iy
K

ao

rin

o

a
K

ak
m

-r
ka
su

A

ub

e
m
hi
A

A

ki

er
i-b
A

y

0

ry

DPPH free radical scavenging activity[nmol Trolox eq. / mg-solid content]

Normal waterH** p < 0.01
* p < 0.05

Fig. 4. Differences in antioxidative efficacy between strawberry varieties.

DPPH free radical scavenging activity[nmol Trolox eq. / mg-solid content]

Fourteen variety of strawberry plants were cultivated either ceramic-treated water or normal water. Hot water extracts of eachstrawberry variety were prepared and used to examine the DPPH radical scavenging activity. All data were shown as the mean
± SD (n = 6). * p < 0.05 and ** p < 0.01 vs. normal water in each variety. DPPH, 2, 2-diphenyl-1-(2,4,6-trinitrophenyl)hydrazyl;
SD, standard deviation.

p = 0.540

Ceramic- treated water
Normal water

100
80
60
40
20
0

Ceramic-treated water

Normal water

Fig. 5. Effect of ceramic-treated water on antioxidative efficacy of strawberry varieties.

Fourteen variety of strawberry plants were cultivated either ceramic-treated water or normal water. Hot water extracts of eachstrawberry variety were prepared and used to examine the DPPH radical scavenging activity. Paired t-test was conducted toinvestigate the effect of cultivation water on antioxidative efficacy of strawberries. All data were shown as the mean ± SD (nﬁgure5= 6). DPPH, 2, 2-diphenyl-1- (2,4,6-trinitrophenyl)hydrazyl; SD, standard deviation.

_ 68 _

ﬁgure

Glycative Stress Research

DPPH free radical scavenging activity[nmol Trolox eq. / mg-solid content]

140
120

**

100

**

**

80

**

**

**

60

**

**

**

**

**

**

**

40
20

a
ak
m

A

rin

o

e

ao
K

hi

m

a

oi

ok
iy

Ya
y

K

pe
op

B

en

ih

bo

sh

i

e
m

Yo
t

ki

hi

in

A

su

or

i

ry
Ko

im

i-b

er

u
A

N

yo

ea
-p

ho

rl

i
ek

ed

m
ra

ki
no

H

os

hi

R

ub
-r
ka

A

su

To
u

ku

y

n

0

Fig. 6. Differences in antioxidative efficacy between strawberry varieties.

Fourteen variety of strawberry plants were cultivated using normal water. Hot water extracts of each strawberry variety wereprepared and used to examine the DPPH radical scavenging activity. All data were shown as the mean ± SD (n = 6). ** p < 0.01
vs. Toukun. SD, standard deviation.

Ceramic- treated water

140

DPPH free radical scavenging activity[nmol Trolox eq. / mg-solid content]

ﬁgure6

Normal water

120

100

80

60

40

60

65

70

75

80

85

90

95

The ratio of inhibitory effect on fluorescent AGE formation [%]Fig. 7. Correlation between anti-glycative efficacy and antioxidative activity.

The inhibitory effect of fluorescent AGE formation was plotted in X-axis and DPPH radical scavenging activity was plotted in
Y-axis. Black line was regression line of ceramic-treated water and gray break line was regression line of normal water. AGE,
advanced glycation end product; DPPH, 2, 2-diphenyl-1-(2,4,6-trinitrophenyl) hydrazyl.

_ 69 _

ﬁgure7

Effect of Cultivation Water on Strawberries.

Discussion
AGEs are produced in vivo by glycation, a non-enzymatic
binding of reducing sugars to proteins, and accumulate in the
body with age. Increased AGE production and accumulation
are associated with increased concentrations of glucose andits metabolites in the blood as well as decreased activity of
metabolic enzymes. In fact, accumulation of AGEs has been
observed in the blood and tissues of diabetic patients 19, 20).
We have examined the anti-glycation effects of morethan 500 kinds of plants and foods 8, 17, 21-23). We have also
examined fruits that are generally considered to have highfructose content and reported that various fruits including
strawberries have anti-glycative effects 8). In addition, we
have recently reported that the anti-glycative effects vary
among varieties in the same plants 24). This is thought to be
due to differences in the components contained in different
varieties, but detailed verification has not been done yet.

Effect of strawberry varieties on food functionality.
Based on the results of the anti-glycative and antioxidative
effects of 14 strawberry varieties grown in normal water, thedifferences among varieties were examined. All strawberryvarieties were shown to have antioxidative and anti-glycative
effects ( Table 3, Fig. 4 ); however, focusing on each variety
revealed that both anti-glycative and antioxidative effects
vary depending on varieties. For example, Koiminori (87.8± 1.37 %) had the highest inhibition rate of fluorescent AGE
formation which was significantly higher than that of the 8varieties: Yotsuboshi, Akihime, Ai-berry, Amaka, Nyohou,
Toukun, and Yayoihime and the lowest, Benihoppe (69.7 ±3.80 %, Fig. 3 ). In terms of antioxidant activity, Toukun
showed significantly higher DPPH radical scavenging
activity than the other 13 varieties ( Fig. 6 ). This difference
may be due to differences in the components of each variety
as a result of crossbreeding. Although Nyohou was selected
as one of the hybrids for most of the varieties, there were
no common hybrids among the varieties that showed strong
anti-glycative and antioxidative effects.
Anthocyanins and ascorbic acid are known as antioxidant
compounds in strawberries 8-10). Anthocyanins are glycosides
with an anthocyanidin backbone, and appropriate amounts ofanthocyanins intakes may reduce the risk of cardiovasculardisease and type 2 diabetes, improve weight maintenance andneuroprotection, and have effects on vascular and glucose
regulation 25). Among anthocyanins, pelargonidin, cyanidin,
and malvidin have been reported as aglycones in strawberry 26).
However, anthocyanins are easily denatured by heat. Even
though we measured anthocyanins using a colorimetricmethod based on the previous report 27), we did not detect
anthocyanins in the hot water extracts used in our study(data not shown). Since only monomeric anthocyanins canbe measured by this method, these results indicate that thecomponents which showed anti-glycative and antioxidative
effects in the extracts are considered to be something otherthan monomeric anthocyanins. However, the details are notknown. Further studies will be needed to identify the antiglycative components of strawberries and measure theiramounts in each variety.

Effect of Ceramic-treated Water Cultivation on
Strawberries.
At the time of the development of special-glaze-applied
ceramic chips, it was thought to be an environment in which
cyanobacteria could grow, and it was assumed that the water
purification effect of ceramic-treated water was due to thereactive oxygen species derived from the oxygen produced bycyanobacteria and the plant growth-promoting effect was dueto the adaptive response of the antioxidant system, in which
mild oxidative stress enhances the antioxidant system 15).
However, the redox potential of ceramic-treated water was
reported to be lower than that of normal water, suggestingthat the oxygen concentration was not increased. We now
hypothesize that the lower redox potential is due to anincrease in the concentration of dissolved hydrogen in theceramic-treated water. As for the source of hydrogen, weare considering the possibility of vibration of the ceramic
chips due to water flow and the catalytic effect of theglaze applied to the ceramic chips, but the details are still
unknown. Another possibility is that hydrogen-producing
bacteria may be growing on the special ceramic chips, butsince hydrogen-producing bacteria are generally considered
to be anaerobic 28), further studies are needed to determine
whether they can grow under the conditions of this study.
Hydrogen molecules have a strong reducing ability and not
only directly scavenge hydroxyl radicals, but also induce
antioxidants and antioxidative enzyme activity in cells 29, 30).
It has also been reported that hydrogen gas administration
inhibits the injuries caused by reactive oxygen speciesderived inflammation such as ischemia-reperfusion 31) andcellular senescence 32). As for plant growth, it has beenreported that hydrogen-rich water promotes the growth ofroots of komatsuna (Japanese mustard spinach) 33) as well as
leaf and root growth promotion in cucumber by induction ofplant growth hormones such as gibberellins and auxins 34).
We have already shown that the use of ceramic-treated water
induces growth promotion in peaches and rice as well as
increases the sugar content of fruits 15). In this study, we
focused on the food functionality of strawberries rather
than growth promotion, especially the anti-glycative andantioxidative effects.
When a paired t-test was performed on 14 strawberryvarieties, cultivation with ceramic-treated water did notincrease anti-glycative and antioxidative effects ( Fig. 2, 5 ).
However, when each variety was examined by t-test,
Koiminori (p = 0.024) was the only variety that showed asignificant increase in anti-glycative effect by the ceramictreated water cultivation. Other than that, Amaka showed atrend of an increase (p = 0.062), but Akihime showed higher
effects in the normal water cultivation ( p = 0.071, Table 3 ).
No effect was observed for the other 11 varieties. In terms ofantioxidant activity, four varieties (Amaka, Kiyoka, Nyohou,
and Yotsuboshi) showed a significant enhancement of DPPH
radical scavenging activity when cultivated with ceramictreated water, while Hoshinokirameki and Red-pearl had
significantly lower activity level when grown with theceramic-treated water ( Fig. 4 ). These results indicate that theeffects of ceramic-treated water on the varieties are limited.
We suspected that the difference is due to different levelsof responsiveness against oxidative stress in each variety.

_ 70 _Glycative Stress Research

It is possible that the dissolved hydrogen concentration in theceramic-treated water used in this study was not sufficientfor varieties susceptible to oxidative stress, or the hydrogenconcentration of the water was not sufficient to enhancethe expression of genes involved in growth promotion andantioxidant enzymes. Because of that, the effects of theceramic-treated water were not reflected in the promotionof food functions. In addition, the special ceramic chipfiltration system could not be installed vertically in thisstudy, so some of the water did not fully contact the ceramic
chips, which may have resulted in a lower dissolved hydrogenconcentration than originally expected. In the future, it will
be necessary to verify the differences in the expressionof growth hormones and antioxidant enzymes in different
varieties when the ceramic chip treatment device is fixedvertically, and the plants are grown with ceramic-treatedwater. Moreover, identification and verification of changes in
the amounts of anti-glycative and anti-oxidative components
in strawberries is also a future task. It will also be necessary
to verify whether the continuous use of ceramic-treatedwater enhances the food functions of the varieties that were
not affected in this study.

Reference

Conclusion
Anti-glycative and antioxidative effects were observedin all 14 strawberry varieties used in this study regardless of
the water used. However, the effect of ceramic-treated wateron the anti-glycative effect and antioxidative efficacy ofstrawberries differed depending on the varieties, suggestingthat the effects of ceramic-treated water may be limited tocertain varieties.

Acknowledgment
This work was supported by the Advanced Instrumental
Analysis Center of Shizuoka Institute of Science andTechnology. The publication of this study was supported by
the Isyoku-Dogen Research Foundation.

Conflict of interest
The authors claim no conflict of interest in this study.

10) Matsuzoe N, Kawaqnobu S, Matsumoto S, et al. Effect of night1) Negre-Salvayre A, Salvayre R, Augé N, et al. Hyperglycemia

and glycation in diabetic complications. Antioxid Redox
Signal. 2009; 11: 3071-3109.
2) Kan H, Yamagishi S, Ojima A, et al. Elevation of serum
levels of advanced glycation end products in patients with
non-B or non-C hepatocellular carcinoma. J Clin Lab
Anal. 2015; 29: 480- 484.
3) Vlassara H, Striker GE. Advanced glycation endproducts
in diabetes and diabetic complications. Endocrinol MetabClin North Am. 2013; 42: 697-719.
4) Yamagishi S. Role of advanced glycation end products(AGEs) in osteoporosis in diabetes. Curr Drug Targets.
2011; 12: 2096-2102.
5) Ward MS, Fortheringham AK, Cooper ME, et al. Targeting
advanced glycation endproducts and mitochondrial
dysfunction in cardiovascular disease. Curr Opin Pharmacol.
2013; 13: 654 - 661.
6) Zakaria MN, El-Bassossy HM, Barakat W. Targeting AGEs
signaling ameliorates central nervous system diabetic
complications in rats. Adv Pharmacol Sci. 2015; 2015:
346259.
7) Takeuchi M, Yamagishi S. Possible involvement of advanced
glycation end-products (AGEs) in the pathogenesis of
Alzheimer's disease. Curr Pharm Des. 2008; 14: 973-978.
8) Parengkuan L, Yagi M, Matsushima M, et al. Antiglycation activity of various fruits. Anti-Aging Med. 2013;
10: 70 -76.
9) Stintzing FC, Carle R. Functional properties of anthocyanins
and betalains in plants, food, and in human nutrition. Trends
in Food Science and Technology. 2004; 15: 19-38.

temperature on sugar, amino acid, ascorbic acid, anthocyaninsand ellagic acid in strawberry (Fragaria × ananassa. Duch.)
fruit. Shokubutsu kankyo Kougaku. 2006; 18: 115-122. (inJapanese)
11) Benvenuti S, Pellati F, Melegari M, et al. Polyphenols,
anthocyanins, ascorbic acid and radical scavenging activityof rubus, ribes, and aronia. Journal of Food Science. 2004;
69: FCT164 -169.
12) Plant Variety Protection Database by Ministry ofAgriculture, Forestry and Fisheries in Japan.
http://www.hinshu2.maff.go.jp/, accessed on 26 Jun 2022.
(in Japanese)
13) Bazzano LA, Li TY, Joshipura KJ, et al. Intake of fruit,
vegetables, and fruit juices and risk of diabetes in women.
Diabetes Care. 2008; 31: 1311-1317.
14) Yonei Y, Haasbroek K, Yagi M, et al. Water quality
improvement effect from the installation of special-glazeapplied ceramics: Benten Pond, Ichikawa, Chiba, Japan.
Glycative Stress Res. 2021; 8: 20-28.
15) Hasegawa T, Sugiura S, Asano M, et al. Cyanobacterium
proliferative actions by special-glaze-applied ceramicpieces and their utilization. Glycative stress Res. 2020; 7:
88-104.
16) Takabe W, Yamaguchi T, Hayashi H, et al. Identificationof antiglycative compounds in Japanese red water pepper(red leaf variant of the Persicaria hydropiper sprout).
Molecules. 2018; 23: 2319.
17) Ishioka Y, Yagi M, Ogura M, et al. Antiglycation effect ofvarious vegetables: Inhibition of advanced glycation end
product formation in glucose and human serum albumin
reaction system. Glycative Stress Res. 2015; 2: 22-34_ 71 _

Effect of Cultivation Water on Strawberries.

18) Oki T. DPPH radical scavenging activity assay. Nippon

Shokuhin Kagaku Kogaku Kaishi. 2007; 71-78.
https://www.jiu.ac.jp/files/user/education/books/pdf/84182. pdf (in Japanese)
19) Kilhovd BK, Berg TJ, Birkeland KI, et al. Serum levels ofadvanced glycation end products are increased in patientswith type 2 diabetes and coronary heart disease. Diabetes
Care. 1999; 22: 1543-1548.
20) Saito M, Marumo K. Collagen cross-links as a determinantof bone quality: A possible explanation for bone fragility
in aging, osteoporosis, and diabetes mellitus. OsteoporosInt. 2010; 21: 195-214.
21) Hori M, Yagi M, Nomoto K, et al. Inhibition of advanced
glycation end product formation by herbal teas and its
relation to anti-skin aging. Anti-Aging Med. 2012; 9: 135148.
22) Moniruzzaman M, Parengkuan L, Yagi M, et al. Effect ofproteins, sugars and extraction methods on the antiglycation
activity of spices. Glycative Stress Res. 2015; 2: 129-139.
23) Otake K, Yagi M, Takabe W, et al. Effect of tea (Camelliasinensis) and herbs on advanced glycation endproductformation and the influence of post-fermentation. Glycative
Stress Res. 2015; 2: 156-162.
24) Wickramasinghe UPP, Yagi M, Yonei Y. Anti-glycative
effect and total phenolic content of rice water of different
Japonica and Indica varieties. Glycative Stress Res. 2021;
8: 162-170.
25) Kalt W, Cassidy A, Howard LR, et al. Recent research on
the health benefits of blueberries and their anthocyanins.
Adv Nutr. 2020; 11: 224-236.
26) Lopes da Silva F, Escribano-Bailon MT, Perez-Alonso JJ,
et al. Anthocyanin pigments in strawberry. Food Scienceand Technology. 2007; 40: 374-382.
27) Lee J, Durst RW, Wrolstad RE. Determination of total
monomeric anthocyanin pigment content of fruit juices,beverages, natural colorants, and wines by the pH differential
method: Collaborative study. J AOAC Int. 2005; 88: 12691278.
28) Akutsu Y, Li Y, Harada H. Review on anaerobic hydrogen
fermentation from organic wastewater. Journal of Japan
Biological Society of Water and Waste. 2008; 44: 57-75.
(in Japanese)
29) Dixon BJ, Tang J, Zhang JH. The evolution of molecularhydrogen: A noteworthy potential therapy with clinicalsignificance. Med Gas Res. 2013; 3: 10.
30) Ohsawa I, Ishikawa M, Takahashi K, et al. Hydrogenacts as a therapeutic antioxidant by selectively reducing
cytotoxic oxygen radicals. Nat Med. 2007; 13: 688-694.
31) Watanabe M, Kamimura N, Iuchi K, et al. Protective effectof hydrogen gas inhalation on muscular damage using amouse hindlimb ischemia-reperfusion injury model. Plast
Reconstr Surg. 2017; 140: 1195-1206.
32) Hara F, Tatebe J, Watanabe I, et al. Molecular hydrogen
alleviates cellular senescence in endothelial cells. Circ J.
2016; 80: 2037-2046.
33) Hamauzu Y, Ishikawa K, Morisawa S. Effects of deoxidized
nutrient solution on growth of komatsuna (Brassica rapa
var. perviridis) plants. Environmental Control in Biology.
2014; 52; 107-111.
34) Wu Q, Su N, Huang X, et al. Hydrogen-rich water promotes
elongation of hypocotyls and roots in plants throughmediating the level of endogenous gibberellin and auxin.
Funct Plant Biol. 2020; 47: 771-778.

_ 72 _