Antifungal Activity of Olive Oil and Ozonated Olive Oil Against Candida Spp. and Saprochaete Spp.

Abstract

Ozonated olive oil was investigated for their capacity to inhibit growth of 38 yeast strains of Candida albicans, Candida glabrata, Candida krusei, Candida parapsilosis, and Saprochaete capitata. Two different ozonated olive oil (OZO1, OZO2) and two different olive oil (OL1, OL2) samples having different biochemical parameters were assessed in terms of their antifungal ability and comparison was made. Fluconazole was chosen as control antifungal agent. Each sample’s antifungal activity decreased in the following order: OZO1>OZO2>OL1≥OL2. This study demonstrated that ozonated olive oil may help to control some fluconazole-resistant and dose-dependent sensitive fungal strains.

Full text

Full Terms & Conditions of access and use can be found at
http://www.tandfonline.com/action/journalInformation?journalCode=bose20
Download by: [66.249.93.81] Date: 09 June 2017, At: 02:54
Ozone: Science & Engineering
The Journal of the International Ozone Association
ISSN: 0191-9512 (Print) 1547-6545 (Online) Journal homepage: http://www.tandfonline.com/loi/bose20
Antifungal Activity of Olive Oil and Ozonated Olive
Oil Against Candida Spp. and Saprochaete Spp.
Kemal Varol , Ayse Nedret Koc, Mustafa Altay Atalay & Ihsan Keles
To cite this article: Kemal Varol , Ayse Nedret Koc, Mustafa Altay Atalay & Ihsan Keles (2017):
Antifungal Activity of Olive Oil and Ozonated Olive Oil Against Candida Spp. and Saprochaete
Spp., Ozone: Science & Engineering, DOI: 10.1080/01919512.2017.1322490
To link to this article: http://dx.doi.org/10.1080/01919512.2017.1322490
Accepted author version posted online: 27
Apr 2017.
Published online: 27 Apr 2017.
Submit your article to this journal
Article views: 66
View related articles
View Crossmark data

Antifungal Activity of Olive Oil and Ozonated Olive Oil Against Candida Spp. and
Saprochaete Spp.
Kemal Varol
a
, Ayse Nedret Koc
b
, Mustafa Altay Atalay
b
, and Ihsan Keles
a
a
Department of Internal Medicine, Faculty of Veterinary Medicine, Erciyes University, Kayseri 38100, Turkey;
b
Department of Microbiology,
Medical Faculty, Erciyes University, Kayseri 38100, Turkey
ABSTRACT
Ozonated olive oil was investigated for their capacity to inhibit growth of 38 yeast strains of
Candida albicans, Candida glabrata, Candida krusei, Candida parapsilosis, and Saprochaete capitata.
Two different ozonated olive oil (OZO1, OZO2) and two different olive oil (OL1, OL2) samples
having different biochemical parameters were assessed in terms of their antifungal ability and
comparison was made. Fluconazole was chosen as control antifungal agent. Each sample’s
antifungal activity decreased in the following order: OZO1 > OZO2 > OL1 ≥OL2. This study
demonstrated that ozonated olive oil may help to control some fluconazole-resistant and dose-
dependent sensitive fungal strains.
ARTICLE HISTORY
Received 30 January 2017
Accepted 12 April 2017
KEYWORDS
Ozone; Antifungal Activity;
Candida Spp; Fluconazole
Resistance; Olive Oil;
Ozonated Olive Oil; Ozone
Therapy; Saprochaete spp
Introduction
Fungal infections are important problems and fre-
quently occur worldwide (Koc et al. 2011,2016). In
recent years, presence of opportunistic fungi that
cause common invasive fungal infections in hospitals
have been reported to increase (Garcia-Ruiz et al. 2013;
Ulu Kilic et al. 2016).
Although in invasive fungal infections, Candida (C)
spp. and Aspergillus spp. are more common, yeast spe-
cies, such as Trichosporon, Saccharomyces, Saprochaete
(S), fungal genera; such as Zygomycetes,orFusarium
can also be seen. S. capitata is also the reason of some
fungal infections (Garcia-Ruiz et al. 2013; Koc et al.
2016; Pfaller and Diekema 2010; Richardson and Lass-
Flörl 2008). C. albicans and rarely C. parapsilosisve C.
tropicalis cause cutaneous candidiasis, which is a skin
infection occurring acutely or chronically (Wagner and
Sohnle 1995). Again, S. capitata invade skin and cause
generalized maculopapular erosion (Garcia-Ruiz et al.
2013; Koc et al. 2016).
In the treatment of fungal infections, polyenes and
some imidazole derivates have been used (Fernández
Torres et al. 2006). Long periods of drug usage or short-
term but inadequate treatments results in drug resistance.
Furthermore, long-term usage of ketoconazole, flucona-
zole (FLU), itraconazole and their derivatives cause side
effects. For all these reasons, new treatments options have
been investigated alternatively to the antimicrobial agents
(Garcia-Ruiz et al. 2013;Geweely2006; Koc et al. 2016;
Ouf et al. 2016; Silici and Koc 2006). One of the treatment
choice is ozone therapy (Ouf et al. 2016). Ozone (O
3
)is
the high-energy form of atmospheric oxygen (O
2
) (Bocci
2011). Despite the stable oxygen molecules, ozone is an
unstable molecule. Ozone gas; has antiviral, antibacterial,
antiparasitic and antifungal activity (Lezcano et al. 2000;
Ouf et al. 2016; Rodrigues, Cardoso, and Caputo 2004).
Furthermore, in toxicological studies, ozone also found to
be a reliable molecule and had no genotoxic activity
(Rodrigues, Cardoso, and Caputo 2004).
Olive oil (OL) is an essential oil obtained from olive
fruit that contains a high percentage (65–85%) of oleic
acid and is also comprised of different fatty acids
(Geweely 2006).OL is known to inhibit reproduction
of microorganisms (Cicerale, Lucas, and Keast 2012). In
recent years, ozone in the form of ozonated olive oil
(OZO) or ozonated water have widely been used for
medical indications (Fernández Torres et al. 2006;
Geweely 2006; Ouf et al. 2016; Sakazaki et al. 2007;
Skalska et al. 2009). Nowadays, essential oils and ozo-
nated oils have been used for the treatment of bacterial,
viral and fungal infections of skin, otitis, intraocular
infections, dental disease and vaginitis (Bocci 2006;
Geweely 2006; Helal et al. 2006; Kim et al. 2009,2013;
Kutlubay et al. 2010; Sechi, Lezcano, and Nunez 2001).
Ozone has strong antimicrobial and germicidal
activity against virus, bacteria, parasites, and fungus.
CONTACT Kemal Varol kmlvrl@yahoo.com Department of Internal Medicine, Faculty of Veterinary Medicine, University of Erciyes, Talas, Kayseri
38100, Turkey.
OZONE: SCIENCE & ENGINEERING
https://doi.org/10.1080/01919512.2017.1322490
© 2017 International Ozone Association

The interaction of ozone molecule against cellular com-
ponents, especially having double bonds sulfhydryl
groups and phenolic rings cause an oxidation reaction
that prevents their growth. Therefore, membrane phos-
pholipids, intracellular enzymes and genomic materials
are attacked by ozone. As a result of this, cell damage
and death of microorganisms occur (Arana et al. 1999;
Gonçalves 2009; Ouf et al. 2016; Perez, Poznyak, and
Chairez 2015). Approximately 80% carbon hydrate and
20% of protein and glycoproteins are the components
of cell walls of fungi, which are multi-layered. Because
there are several disulphide bonds in the site, this
promotes oxidative inactivation by ozone. Ozone has
the ability to diffuse through the fungal wall, then
enters into the cytoplasm and disturbs the vital cellular
functions. Ozone also has an inhibitory effect on spore
germination and production (Antony-Babu and
Singleton 2009; Ouf et al. 2016).
The effect of ozone combined with olive oil occurs
with almost all of the carbon-carbon double bonds that
are present in unsaturated fatty acids, which cause dif-
ferent toxic products such as many oxygenated com-
pounds, ozonides, aldehydes and peroxides. These
compounds can also account for the wide antimicrobial
activity of OZO (Pryor and Uppu 1993; Ouf et al. 2016).
The safety of ozonated sunflower oil (OSO) was reported
by Gundarova et al. (1996) and Alvarez et al. (1997).
The aim of the present study was to determine antifungal
activity of OZO and OL in vitro against different strains of
yeast using guidelines of Clinical Laboratory Standards
(CLSI) (2008) document M27-A3. Results presented in
this study will assist in future studies to help focus more
studies on OZO as a potential natural drug to control most
fungal pathogens in medical mycology.
Material methods
Study design
In this study, two different olive oil [Olive oil 1
(OL1) and Olive oil 2 (OL2)] samples having differ-
ent acidity, iodine, peroxide, p-anisidine, pH and
viscosity values and two Ozonated olive oil samples
[Ozonated olive oil 1 (OZO1) and Ozonated olive
oil 2 (OZO2)] obtained after ozonation of the olive
oils (OL1 and OL2) were used. Their antifungal
activities were then compared with the effects
of FLU.
Identification and antifungal susceptibility
Thirty-eight non-repetitive strains isolated from blood cul-
tures at the Medical Microbiology Department of Erciyes
University, Gevher Nesibe Hospital were included in the
study. All strains tested were considered as infection agents.
Again, all strains were collected during a 6-month period in
the Mycology Laboratory. They were comprised of 10
strains of C. albicans, 10 strains of C. glabrata,10strains
of C. parapsilosis,4strainsofC. krusei and 4 strains of S.
capitata. One standard C. albicans ATCC 90028 and one
standard C. parapsilosis ATCC 22019 were also used.
Strains were identified as follows: according to carbon
hydrate assimilation of with API AUX C 20 (bioMérieux,
France) kits, the macroscopic and microscopic morphol-
ogy, capability of growing at 37 ºC on corn meal agar, germ
tube test, sensitivity for cycloheximide and urea hydrolysis.
The isolates used in this study were kept at −20 ºC in tryptic
soy broth having 10% glycerine. Before their examination,
they were melted and sub-cultured on Sabouraud glucose
agar plates at least twice. Quality control was made accord-
ing to suggestions of CLSI document M27-A3 by testing C.
albicans ATCC 90028 and C. parapsilosis ATCC 22019
(CLSI 2008).
FLU was obtained as a powder from the manufacturer
(Fako Co., Istanbul, Turkey). CLSI (2008) (M27-A3),
microdilution broth methods were used to determine
MICs. Antifungal activity of OL, OZO and FLU, in vitro
were examined by the guidelines given (M27-A3). FLU
dissolutions were made in sterile distilled water. Final
drug content in the microdilution plates ranged between
0.125 to 64 μg/mL for FLU, and from 0.1 to 50% (v/v) for
all of the olive oils and ozonated olive oils (OZO1, OZO2,
OL1, and OL2) and its’dilutions were made in RPMI 1640
broth medium (Sigma Chemical Co., Madrid, Spain) with
L-glutamine but without sodium bicarbonate and buffered
at pH 7.0 with 0.165 mol/L morpholinepropansulfonic acid
(Sigma Chemical Co.).
Inoculum suspensions of the yeast were formulated
as explained in the CLSI M27-A3 document using
sterile saline solution (0.85%). Cell density was
adjusted. To do this a spectrophotometer was used. At
a 530-nm wavelength, an adequate saline was added to
match the transmittance generated by a 0.5 McFarland
density standard which resulting in a concentration of
0.5 × 10
3
–2.5 × 10
3
cells/mL. MICs were determined
visually at 24 and 48 h of incubation at 35 ºC. The
plates were then investigated for the presence or
absence of growth at 24 and 48 h.
The MIC for olive oils and ozonated olive oils were
described as the lowest concentration that optically
observed. For the fluconazole, the MIC was expressed
as the lowest concentration in which 50% decrease in
turbidity as observed visually. For all of the OL and
OZO the lowest concentration was 100%. MIC
50
and
MIC
90
, minimal inhibitory concentration at which 50%
and 90%, respectively, of the isolates were inhibited.
2K. VAROL ET AL.

Ozonation procedure
OL and OZO were obtained from a certified commercial
company (Aktifoks, Isık Cosmetics/Turkey) as required
for experimental study. According to commercial firm,
Hansler brands (Ozonosan Alpha Plus, Germany) ozone
device (which produce 95% oxygen and 5% ozone mix-
ture) was used to prepare 10 L of OL; this device report-
edly produced 25 mg/L of ozone per min in at 18–20 °C.
This procedure was applied for 10 days.
Characterization of oil and ozonated olive oil
Acidity value
Acidity value of OL and OZO were determined accord-
ing to American Oil Chemists’Society (AOCS) (1998a).
Acidity index expressed as the quantity of potassium
hydroxide that are necessary to neutralize the free fatty
acid in 1 g. of OZO (Diaz et al. 2005,2006;Geweely2006;
Sechi, Lezcano, and Nunez 2001;Travaglietal.2010).
Iodine value
Iodine values of OL and OZO were determined accord-
ing to AOCS (1998b) defined as the number of grams
of iodine that is a measure of unsaturation rate of OZO
(Diaz et al. 2006; Geweely 2006; Molerio et al. 1999;
Sechi, Lezcano, and Nunez 2001; Travagli et al. 2010).
Peroxide value
Peroxide value (PV) of OL and OZO were determined
according to AOCS (1998c), which suggests the amount
of peroxide within the OZO. It is expressed as the quan-
tity of active oxygen per kilogram of OZO (mmol/kg)
and defined as milliequivalent (Cirlini et al. 2012;Diaz
et al. 2005; Geweely 2006; Molerio et al. 1999; Moureu
et al. 2016, 2006; Sechi, Lezcano, and Nunez 2001;Tellez,
Lozano, and Gomez 2006; Travagli et al. 2010).
P-anisidine value
P-anisidine value of OL and OZO were determined
according to AOCS (2011). It is about the aldehyde
ratio, which is determined by adding free hydroxylamine
to the aldehyde carboxylic group. The result is expressed
as in mmol/g. (Sechi, Lezcano, and Nunez 2001)
pH value
pH values of OL and OZO were determined at room
temperature (24 °C).
Viscosity
Viscosity of OL and OZO were determined by vibrating
viscometer device (AND SV-10 Japan) at 24–40 °C. To
provide a swift quality control assessment during the entire
ozonation process, a typical trend can be a useful tool, and
also deciding on the process time for obtaining the desired
ozonation level for the sample is important (Diaz et al.
2005; Sechi, Lezcano, and Nunez 2001;Travaglietal.2010).
Results
OZO and OL peroxide, acidity, iodine, p-anisidine,
viscosity and pH values that were used in this study
are given in Table 1. Antimicrobial sensitivities are
given in Table 2. According to the data all microorgan-
isms used in the present study OL had no antifungal
activity but FLU, OZO1 and OZO2 had antifungal
activity (Table 2 and Table 3).
Each fungal strains sensitivity against OZO1, OZO2,
OL1 and OL2 were tested. Their MIC values were also
summarized in Table 2. When all strains are taken into
consideration together at 24 h, MICs of geometric mean
values of OZO1, OZO2, OL1, OL2 and FLU were 0.437%
(v/v), 0.678% (v/v), 48.11% (v/v), 50% (v/v) and 1.924 µg/
mL, respectively. When all strains are taken into considera-
tion together at 48 h, MICs of geometric mean values of
OZO1, OZO2, OL1, OL2 and FLU were 1.193% (v/v),
1.493% (v/v), 50% (v/v), 50% (v/v) and 1.771 µg/mL,
respectively (Table 2).
The samples showed antifungal activity in terms of the
geometric mean in all strains as follows: OZO1 > OZO2 >
OL1>OL2at24handasfollows:OZO1>OZO2>OL1≥
OL2at48h(Table 2). All antifungal agents, MIC ranges,
geometric mean, and the MIC
50
,MIC
90
values for Candida
and S. capitata were summarized in Table 3.TheMIC
50
values were lowest for OZO1, OZO2, and FLU.
In this study, at 24 h when all strains were considered
together, MIC range values of OZO1, OZO2, OL1, OL2 and
FLU were between 0.1 to 1.56% (v/v), 0.1 to 6.25% (v/v),
50% (v/v), 50% (v/v), and 0.25 to 32 µg/mL, respectively. At
Table 1. Olive oil, ozonated olive oil analysis results.
Olive oil 1 Olive oil 2 Ozonated olive oil 1 Ozonated olive oil 2 Unit
Peroxide value 392 370 1352 1053 mmol—mEq/kg
Acidity value 0.7281 2.0079 8.9951 8.4948 unit
Iodine value 64.8067 62.8731 2.2040 4.3750 unit
P-Anisidine value 0.155 0.100 0.516 0.738 mmol/g
Viscosity (24-40 °C) 81.7 85.5 cp 1000 1630 centipoise
pH (24°C) 4.7 4.4 2.1 1.3 -
OZONE: SCIENCE & ENGINEERING 3

48 h, when all strains were considered together, MIC range
values of OZO1, OZO2, OL1, OL2 and FLU were between
0.1 to 6.25% (v/v), 0.1 to 12.5% (v/v), 50% (v/v), 50% (v/v),
and 0.25 to 64 µg/mL, respectively (Table 3).
At 24 h, two strains of C. glabrata were resistant
against fluconazole. Nine strains of C. glabrata and
10 strains of C. krusei were dose-dependent sensitive
against fluconazole. At 48 h, 2 strains of C. glabrata
were resistant against fluconazole. Eight strains of C.
glabrata and 10 strains of C. krusei were dose-
dependent sensitive against fluconazole. (Table 2).
At 24 h, C. glabrata and C. krusei had the lowest
MIC values against OZO1 and OZO2. At 48 h, C.
krusei had also the lowest MIC values against OZO1
and OZO2. But, C. glabrata had only the lowest
MIC values against OZO2. Furthermore, S. capitate
had also the lowest MIC values against OZO1
(Table 3).
On the other hand, at 24 h, C. albicans had the
highest MIC values against OZO1 and OZO2. At
48 h, C. albicans had the highest MIC values against
only OZO2. In addition, C. parapsilosis had the highest
MIC values against OZO1 (Table 3).
For the resistant strains against FLU MIC range
values of OZO1, OZO2, OL1 and OL2 were 0.1 and
6.25% (v/v), 0.1and 3.125% (v/v), 50% (v/v) and 50%
(v/v), respectively (Table 3). Results of this study, at 24-
h and 48-h incubation, sensitive strains, dose-depen-
dent sensitive strains and resistant strains for FLU are
shown in Table 4.
Discussion
Mycoses are important problems in our daily lives.
Even though we know they may threaten our life,
research to combat their survival is still neglected
(Ansari et al. 2013). Human and animal populations
have been increasing, mycoses are getting more wide-
spread; and, at the same time, antimicrobial resistance
also develops. Thus, scientists are searching for
Table 2. Minimum inhibitory concentrations of olive oil 1, olive oil 2, ozonated olive oil 1, ozonated olive oil 2 and fluconazole.
Incubation time (24 h) Incubation time (48 h)
In µg/mL In µg/mL
Number Strain Material OL 1% OL 2% OZO 1% OZO 2% FLU OL 1% OL 2% OZO 1% OZO 2% FLU
1C. albicans Blood 50 50 1.56 6.25 0.25 50 50 3.125 12.5 0.25
2C. albicans Blood 50 50 1.56 6.25 0.25 50 50 3.125 6.25 0.25
3C. albicans Blood 50 50 1.56 3.125 0.25 50 50 3.125 6.25 0.25
4C. albicans Blood 50 50 1.56 3.125 0.25 50 50 3.125 6.25 0.25
5C. albicans Blood 50 50 1.56 3.125 0.25 50 50 3.125 6.25 0.25
6C. albicans Blood 50 50 1.56 3.125 0.25 50 50 1.56 3.125 0.25
7C. albicans Blood 50 50 1.56 1.56 0.5 50 50 1.56 1.56 0.5
8C. albicans Blood 50 50 0.4 1.56 0.5 50 50 1.56 1.56 0.5
9C. albicans Blood 50 50 0.4 0.8 0.5 50 50 0.8 1.56 0.5
10 C. albicans Blood 50 50 0.1 0.8 1 50 50 0.4 0.8 1
11 C. glabrata Blood 50 50 1.56 0.4 8 50 50 6.25 3.125 8
12 C. glabrata Blood 50 50 1.56 0.2 8 50 50 6.25 3.125 8
13 C. glabrata Blood 50 50 0.8 0.1 16 50 50 6.25 1.56 16
14 C. glabrata Blood 50 50 0.8 0.1 16 50 50 3.125 0.8 16
15 C. glabrata Blood 50 50 0.8 0.1 16 50 50 3.125 0.4 16
16 C. glabrata Blood 50 50 0.8 0.1 16 50 50 3.125 0.4 16
17 C. glabrata Blood 50 50 0.2 0.1 16 50 50 3.125 0.4 32
18 C. glabrata Blood 50 50 0.1 0.1 16 50 50 1.56 0.2 32
19 C. glabrata Blood 50 50 0.1 0.1 32 50 50 1.56 0.2 64
20 C. glabrata Blood 50 50 0.1 0.1 64 50 50 0.4 0.1 64
21 C. parapsilosis Blood 50 50 1.56 3.125 0.5 50 50 6.25 6.25 0.5
22 C. parapsilosis Blood 50 50 1.56 3.125 0.5 50 50 6.25 3.125 0.5
23 C. parapsilosis Blood 50 50 0.8 1.56 0.5 50 50 6.25 3.125 0.5
24 C. parapsilosis Blood 50 50 0.4 1.56 0.5 50 50 3.125 3.125 0.5
25 C. parapsilosis Blood 50 50 0.4 1.56 0.5 50 50 3.125 3.125 0.5
26 C. parapsilosis Blood 50 50 0.4 1.56 1 50 50 3.125 3.125 1
27 C. parapsilosis Blood 50 50 0.4 0.8 1 50 50 3.125 1.56 1
28 C. parapsilosis Blood 50 50 0.4 0.8 1 50 50 3.125 1.56 1
29 C. parapsilosis Blood 50 50 0.4 0.8 1 50 50 1.56 1.56 1
30 C. parapsilosis Blood 50 50 0.1 0.8 1 50 50 0.4 1.56 1
31 C. krusei Blood 50 50 0.2 0.4 8 50 50 0.4 0.8 8
32 C. krusei Blood 50 50 0.1 0.4 16 50 50 0.2 0.8 16
33 C. krusei Blood 50 50 0.1 0.1 16 50 50 0.1 0.1 16
34 C. krusei Blood 50 50 0.1 0.1 32 50 50 0.1 0.1 32
35 S. capitata Blood - - - - - 50 50 0.2 3.125 0.25
36 S. capitata Blood - - - - - 50 50 0.2 3.125 0.5
37 S. capitata Blood - - - - - 50 50 0.1 1.56 0.5
38 S. capitata Blood - - - - - 50 50 0.1 1.56 0.5
39 C. albicans ATCC 90028 Standart 25 50 0.1 3.125 0.25 50 50 0.1 3.125 0.25
40 C. parapsilosis ATCC 22019 Standart 25 50 0.1 1.56 0.5 50 50 0.1 3.125 1
GM of Total strains 48.11 50 0.437 0.678 1.924 50 50 1.193 1.493 1.771
GM, geometric mean; MIC, minimal inhibitory concentration.
4K. VAROL ET AL.

compounds that are more effective compared to older
ones, have wide spectrum, do not cause resistance, and
are natural products (Arif et al. 2009). These new
compounds are natural products and are tolerated bet-
ter by patients and have several beneficiary effects to
them (Di Santo 2010; Fernández Torres et al. 2006;
Hammer, Carson, and Riley 2002; Lima et al. 1993,
2006). Among the natural products having antimicro-
bial activity; besides wild plants, essential oils, ozone
and ozonated oils formed by ozonation of essential oils
(Arif et al. 2009; Di Santo 2010; Fernández Torres et al.
2006, 2013; Kutlubay et al. 2010).
Ozonated vegetable oils have antibacterial and fun-
gicide activity (Geweely 2006). Wu, Doan, and Cuenca
(2006) also reported that ozone gas has an inactivating
effect on fungi. Furthermore, Menendez et al. (2002)
used OSO to treat onychomycosis and found it effective
without side effects. In addition, Daud et al. (2011)
used OSO to treat dermatomycoses caused by
Microsporidium (M) canis, and is reported to be effec-
tive. Moreover, Leopoldina et al. (1998) stated that
oleozone was effective against Trichophyton (T)
rubrum, T. mentagrophytes, C. albicans and M. canis.
Leopoldina et al. (1998) used OSO to treat superficial
mycoses in 1,000 patients and found 91% success in
these cases. OZO was also found to have germicidal
activity against Tinea pedis (Menendez et al. 2002).
In the present study, OZO was also found to have
effective antifungal capacity. Each yeast pathogen exam-
ined in the present study had different sensitivity
responses. These results were in parallel with the results
reported by other workers which investigated infections
caused by Microsporum, Trichophyton, Epidermophyton
and C. albicans (Gewely 2006). Furthermore, Leopoldina
et al. (1998) also found OZO effective against T. rubrum,
T. Mentagrophytes, C. albicans and M. canis. Additionally,
Geweely (2006) found MIC values of OZO against C.
albicans, M. canis ve T. rubrum as 0.78–0.53 mg/mL,
which is in agreement with our results.
In a study carried out by Tara et al. (2014)on
vulvovaginal candidiasis, using OZO and clotrimazole
for treatment, they found OZO as effective as clotrima-
zole in reducing clinical symptoms of vulvovaginal
candidiasis and also resulted in negative specimen cul-
tures. Similarly, in the present study, OZO reduced C.
albicans cultures in vitro.
Ouf et al. (2016) investigated the five most common
dermatophytes (M. canis, M. gypseum, T. rubrum, T.
mentagrophytes, and T. interdigitales), and the effects of
different concentrations of gas ozone and ozonated oils
Table 3. Susceptibility to olive oil 1, olive oil 2, ozonated olive oil 1, ozonated olive oil 2 and fluconazole as determined by the
Clinical Laboratory Standards Broth Microdilution Methods.
MIC values
Incubation time (24 h) Incubation time (48 h)
Antifungal agent Range GM MIC
50
MIC
90
Range GM MIC
50
MIC
90
Olive oil 1 (%(v/v))
C. albicans (n:10) 50 50 50 50 50 50 50 50
C. glabrata (n:10) 50 50 50 50 50 50 50 50
C. parapsilosis(n:10) 50 50 50 50 50 50 50 50
C. krusei (n:4) 50 50 50 50 50 50 50 50
S. capitata (n:4) -- - 50 505050
Olive Oil 2 (%(v/v))
C. albicans (n:10) 50 50 50 50 50 50 50 50
C. glabrata (n:10) 50 50 50 50 50 50 50 50
C. parapsilosis(n:10) 50 50 50 50 50 50 50 50
C. krusei (n:4) 50 50 50 50 50 50 50 50
S. capitata (n:4) 50 - - - 50 50 50 50
Ozonated olive oil 1 (%(v/v))
C. albicans (n:10) 0.1–1.56 0.90 1.56 1.56 0.4–3.125 1.80 1.56 3.125
C. glabrata (n:10) 0.1–1.56 0.40 0.8 1.56 0.4–6.25 2.72 3.125 6.25
C. parapsilosis(n:10) 0.1–1.56 0.44 0.4 1.56 0.4–6.25 2.9 3.125 6.25
C. krusei (n:4) 0.1–1.56 0.35 0.1 0.1 0.1–0.4 0.17 0.1 0.2
S. capitata (n:4) - - - - 0.1–0.2 0.14 0.1 0.2
Ozonated olive oil 2 (%(v/v))
C. albicans (n:10) 0.8–6.25 2.38 3.125 6.25 0.8–12.5 3.35 3.125 6.25
C. glabrata (n:10) 0.1–0.4 0.12 0.1 0.2 0.1–3.125 0.56 0.4 3.125
C. parapsilosis(n:10) 0.8–3.125 1.37 1.56 3.125 1.56–6.25 2.54 3.125 3.125
C. krusei (n:4) 0.1–0.4 0.2 0.1 0.4 0.1–0.8 0.28 0.1 0.8
S. capitata (n:4) - - - - 1.56–3.125 2.20 1.56 3.125
Fluconazole (µg/mL)
C. albicans (n:10) 0.25–1 0.35 0.25 0.5 0.25–1 0.35 0.5 0.5
C. glabrata (n:10) 8–64 17.15 16 32 8–64 21.11 16 64
C. parapsilosis(n:10) 0.5–1 0.70 0.5 1 0.5–1 0.70 0.5 1
C. krusei (n:4) 8–32 16 16 16 8–32 16 16 16
S. capitata (n:4) -- - - 0.25–0.5 0.42 0.5 0.5
GM, geometric mean; MIC, minimal inhibitory concentration; MIC
50
and MIC
90
, minimal inhibitory concentration at which 50% and 90%, respectively, of the
isolates were inhibited.
OZONE: SCIENCE & ENGINEERING 5

on these dermatophytes’growth and germination were
examined. Furthermore, they pointed out that OZO has
a better fungicidal effect compared to gaseous ozone
and they explained the possible reason for this being a
result of long-term ozonation, gradual decrease of fatty
acid chain unsaturation, formation of ozonide, and
increases in peroxide and acid values.
PV is generally used as an indicator of the progression
or controlling ozonation process due to its simplicity,
rapidity, and low cost. In addition, the PV may be suffi-
cient for the stability evaluation of vegetable oil ozonides,
and it seems to be very considerable for trading distribu-
tion as well as determining better storage modalities.
Differences obtained in the present study with concern
to PV; believed to be due to ozonation time and acid
number which had been reported to be essential for a
validated PV (Sechi, Lezcano, and Nunez 2001;Geweely
2006; Travagli et al. 2010).
Diaz et al. (2006) compared the antibacterial efficacy
of OZO and OSO with different PVs and found that
OZO and OSO with high PVs demonstrated better
antimicrobial and germicidal activity.
Fernández Torres et al. (2006) have used antifungal
activity on ozonated theobroma oil against C. albicans
ATCC 10231 standards strain and used four different PVs
of ozonated theobroma oil. Ozonated The obroma oil
showed inhibitory effects on C. albicans:whenPVwas
1200 mmol-Eq/kg, MIC value was 3.75 mg/mL, when PV
was 1002, MIC value was 5.78 mg/mL, when PV was
572 mmol-Eq/kg, MIC value was 15 mg/mL, when PV
was 260, MIC value was 25 mg/mL. They also determined
that the MIC concentration decreased as PV increased. Our
Table 4. Comparison of inhibitory activity of fluconazole, olive oil 1, olive oil 2, ozonated olive oil 1 and
ozonated olive oil 2 samples according to sensitivity and resistance to fluconazole of the strains.
Sample
concentration %(v/v)
24-h incubation 48-h incubation
Sensitive
strains
Dose-dependent
sensitive strains
Resistant
strains
Sensitive
strains
Dose-dependent
sensitive strains
Resistant
strains
Olive oil 1
≤50 22 14 1 24 12 2
25 - - - - - -
12.5 - - - - - -
6.25 - - - - - -
3.125 - - - - - -
1.56 - - - - - -
0.8 - - - - - -
0.4 - - - - - -
0.2 - - - - - -
0.1 - - - - - -
Olive Oil 2
≤50 22 14 1 24 12 2
25 - - - - - -
12.5 - - - - - -
6.25 - - - - -
3.125 - - - - - -
1.56 - - - - - -
0.8 - - - - - -
0.4 - - - - - -
0.2 - - - - - -
0.1 - - - - - -
Ozonated olive oil 1
≤50 - - - - - -
25 - - - - - -
12.5 - - - - - -
6.25 - - - 3 3 -
3.125 - - - 10 4 -
1.56 9 2 - 4 1 1
0.8 1 4 - 1 - -
0.4 8 - 2 1 1
0.2 - 2 - 2 1 -
0.1 4 5 1 4 2 -
Ozonated olive oil 2
≤50 - - - - - -
25 - - - - - -
12.5 - - - 1 - -
6.25 2 - - 5 - -
3.125 7 - - 10 2 -
1.56 7 - - 9 1 -
0.8 6 - - 1 3 -
0.4 - 3 - - 3 -
0.2 - 1 - - 1 1
0.1 - 9 1 - 2 1
Total strains 22 13 1 26 12 2
FLU sensitivity = resistance was classified as follows: sensitive strain, MIC <8 µg/mL; dose-dependent sensitive strain, MIC
8–32 µg/mL; and resistant strains, MIC >64 µg/mL.
6K. VAROL ET AL.

findings were in parallel with the preceding results (Table 1
and Table 3). Furthermore, C. albicans ATCC 90028 at 24
and 48 h, we found that, when PV was 1352 mmol-Eq/kg,
MIC value was 0.1% (v/v) in OZO1, when PV was 1053;
MIC value was 3.125% (v/v) in OZO2. As the PVs
increased, antifungal activity also increased as reported in
the literature. In addition, it was determined that OZO1
showed a better (with concern to geometric range) anti-
fungal effect against all strains compared to OZO2.
Kawamura et al. (1986) reported that when C. para-
psilosis exposed to 0.23 mg/L ozone and C. tropicalis
exposed to 0.02 mg/L ozone for 20 sec, 2 log reduction
in the number of yeast cells observed. Thomson et al.
(2011) investigated the effect of ozonated oil on derma-
tophytes, and they found the MIC values of OZO on C.
parapsilosis ATCC 22019 as 0.25–2% and they found
MIC
50
and MIC
90
values as 1%. They also investigated
C. krusei ATCC 6258 and found same MIC values as
given previously. In the present study, it can be seen
from Table 3 that MIC values of OZO1 and OZO2 were
in accordance with the literature values, with regard to C.
parapsilosis standard (ATCC 22019) and clinical isolates
and C. krusei clinical isolates. Of these, OZO1 against C.
parapsilosis ATCC 22019 was much more effective com-
pared to Thomson et al.’s(2011) findings, but it was less
effective against the same culprit compared to Thomson
et al.’s(2011) findings when OZO2 was the case.
Essential oil of plants has been known to have anti-
microbial activity against a wide range of bacteria and
fungus. In fact, extra virgin olive oil was reported to
have antibacterial character, thereby inhibiting recruit-
ment of microorganisms. It was reported to be used as
an adjunctive therapeutic agent for some diseases
(Cicerale, Lucas, and Keast 2012; Helal et al. 2006).
Furthermore, Markin, Duek, and Berdicevsky (2003)
examined water extract of olive leaf against some
microorganisms and found that C. albicans was killed
within 24 h. In contrast, in the present study, OL had
no antifungal activity against yeast tested.
According to our findings, in general, strains that were
susceptible against FLU were less susceptible against OZO1
and OZO2 or vice versa, meaning that strains resistant
against FLU were much more susceptible against OZO1
and OZO2. Mcintyre and Galgani (1989) studied the effect
of pH on cilofungin antifungal activity according to broth
dilutionmethodandtheytested3,4,5,6,7.4pHvalues
against C. albicans (C17; ATCC 64546), C. tropicalis (F26),
C. parapsilosis (3288), and C. lusitaniae MIC values. They
found against these fungi that susceptibility was better at
pH 7.4 compared to pH 3. On the other hand, they also
examined C. glabrata (R87) strain at pH 3, 4, 5, 6, 7.4 and
found MIC values as 5 µg/mL, 10 µg/mL, 5 µg/mL, 10 µg/
mL and 5 µg/mL, respectively. According to the preceding
results, the C. glabrata (R87) strain responded to different
pH values differently, which is not easy to explain. In our
study, C. glabrata strainsweremoresensitiveagainst
OZO2 compared to OZO1 that OZO1 had higher PV
and had higher pH values compared to OZO2 as seen in
Table 1, for which we also could not explain the behavior of
C. glabrata strains. For these reasons, this issue should be
investigated separately in future studies.
There is insufficient information about antifungal
effects of S. capitata (Miceli, Diaz, and Lee 2011).
However, in some studies, as in our study, FLU was an
active drug for this fungus. Reduced susceptibility to
caspofungin with an MIC range value of 0.25–8g/mL
in all species given (Koc et al. 2016). In literature studies,
there is insufficient information about susceptibility of S.
capitata against OZO. In our study, OZO1 and OZO2
were found to be effective against four strains of S.
capitata. Additionally, OZO1 was more effective com-
pared to OZO2 against 4 strains of S. capitata.
Furthermore, sensitivity to S. capitata strains could not
be determined at 24 h; yet, sensitivity at 48 h was obvious
in the present study and should be evaluated further
In conclusion, antifungal activity was demonstrated in
OZO products. This study indicates that OZO can help
control some fungal pathogens. In addition, this study
has proved that some yeasts resistant to certain antifungal
agents are susceptible to OZO. Furthermore, PV of OZO
is very important in its antifungal activity and increased
proportionally with the PV apart from C. glabrata.
Especially, when OZO is prepared for use, their PVs
need to be analyzed. Those with low PVs should not be
used for yeasts. It has been concluded that the data should
be supported by further in vitro and in vivo studies.
ORCID
Kemal Varol http://orcid.org/0000-0002-3057-2865
References
Alvarez, R., S. Menendez, M. Peguera, and J. Turrent. 1997.
“Treatment of Primary Pyoderma with Ozonized
Sunflower Oil.”76:24–6. Paper presented at the 2nd
International Symposium on Ozone Applications,
Havana, Cuba, March 24–26.
Ansari, M., A. Anurag, Z. Fatima, and S. Hameed. 2013.
“Natural Phenolic Compounds: A Potential Antifungal
Agent.”Microbial Pathogens and Strategies for Combating
Them: Science, Technology and Education 1:1189–95.
doi:10.1155/2014/895193.
Antony-Babu, S., and I. Singleton. 2009. “Effect of Ozone on
Spore Germination, Spore Production and Biomass
Production in Two Aspergillus Species.”Antonie Van
Leeuwenhoek 96:413–22. doi:10.1007/s10482-009-9355-2.
OZONE: SCIENCE & ENGINEERING 7

AOCS. 1998a. “Acid Value. AOCS Official Test Method Cd
3a-63 For.”In Official Methods and Recommended
Practices of the American Oil Chemists’Society, edited by
D. Firestone5th. Champaign, IL: American Oil Chemists’
Society.
AOCS. 1998b. “Iodine Value of Fats and Oils. WIJS Method.
AOCS Official Method Cd 1-25:.”In Official Methods and
Recommended Practices of the American Oil Chemists’
Society, edited by D. Firestone5th. Champaign, IL:
American Oil Chemists’Society.
AOCS. 1998c. “Peroxide Value. AOCS Official Method Cd
8-53:.”In Official Methods and Recommended Practices of
AOCS, edited by D. Firestone5th. Champaign, IL:
American Oil Chemists’Society.
AOCS. 2011. “P-Anisidine Value. AOCS Method Cd 18-90
(11): Official Method.”In Official Methods and
Recommended Practices of the American Oil Chemists’
Society, edited by D. Firestone6th. Champaign, IL:
American Oil Chemists’Society.
Arana, I., P. Santorum, A. Muela, and I. Barcina. 1999.
“Chlorination Andozonation of Waste-Water: Comparative
Analysis of Efficacy through the Effect on Escherichia Coli
Membranes.”Journal of Applied Microbiology 86:883–8.
doi:10.1046/j.1365-2672.1999.00772.x.
Arif, T., J. D. Bhosale, N. Kumar, T. K. Mandal, R. S. Bendre,
G. S. Lavekar, and R. Dabur. 2009. “Natural Products-
Antifungal Agents Derived from Plant.”Journal of Asian
Natural Products Research 7:621–38. doi:10.1080/
10286020902942350.
Bocci, V. 2006. “Scientific and Medical Aspects of Ozone
Therapy.”State of the Art. Archives of Medical Research
37:425–35. doi:10.1016/j.arcmed.2005.08.006.
Bocci, V. 2011. Ozone. A New Medical Drug, 2nd ed., 1–100.
Dordrecht, The Netherlands: Springer.
Cicerale, S., L. J. Lucas, and R. S. Keast. 2012. “Antimicrobial,
Antioxidant and Anti-Inflammatory Phenolic Activities in
Extra Virgin Olive Oil.”Current Opinion in Biotechnology
23:129–35. doi:10.1016/j.copbio.2011.09.006.
Cirlini, M., A. Caligiani, G. Palla, A. De Ascentiis, and P.
Tortini. 2012. “Stability Studies of Ozonized Sunflower Oil
and Enriched Cosmetics with a Dedicated Peroxide Value
Determination.”Ozone: Science & Engineering 34:293–9.
doi:10.1080/01919512.2012.692992.
CLSI. 2008. Reference Method for Broth Dilution Antifungal
Susceptibility Testing of Yeasts; Approved Standard. 3th ed.,
CLSI Document M27- A3. Wayne, PA: Clinical and
Laboratory Standards Institute.
Daud, F. V., S. M. Y. Ueda, A. Navarini, and L. M. J. Mímica.
2011. “The Use of Ozonized Oil in the Treatment of
Dermatophytosis Caused by Microsporum Canis in
Rabbits.”Brazilian Journal of Microbiology 42:274–81.
doi:10.1590/S1517-83822011000100035.
Di Santo, R. 2010. “Natural Products as Antifungal Agents
against Clinically Relevant Pathogens.”Natural Product
Reports 27:1084–98. doi:10.1039/b914961a.
Diaz, M., N. Nuiez, D. Quincose, W. Diaz, and F. Hernandez.
2005. “Study of Three Systems for Ozonation of Coconut
Oil.”Ozone: Science & Engineering 2:153–7. doi:10.1080/
01919510590925275.
Diaz, M. F., R. Hernandez, G. Martinez, G. Vidal, M. Gómez,
H. Fernández, and R. Garcés. 2006. “Comparative Study of
Ozonized Olive Oil and Ozonized Sunflower Oil.”Journal
of the Brazilian Chemical Society 17:403–7. doi:10.1590/
S0103-50532006000200026.
FernándezTorres,I.,V.CurtiellasPiñol,E.Sánchez
Urrutia, and M. Gómez Regueiferos. 2006. “In Vitro
Antimicrobial Activity of Ozonized Theobroma Oil
against Candida Albicans..”Ozone Science and
Engineering 28:187–90. doi:10.1080/
01919510600689380.
Garcia-Ruiz, J. C., L. Lopez-Soria, I. Olazabal, E. Amutio, I.
Arrieta-Aguirre, V. Velasco-Benito, J. Pontóne, and M. D.
Moragues. 2013. “Invasive Infections Caused by S.
Capitata in Patients with Haematological Malignancies:
Report of Five Cases and Review of the Antifungal
Therapy.”Revista Iberoamericana De Micologia 30:248–
55. doi:10.1016/j.riam.2013.02.004.
Geweely, N. S. 2006. “Antifungal Activity of Ozonized Olive
Oil (Oleozone).”International Journal of Agriculture and
Biology 8:670–5.
Gonçalves, A. A. 2009. “Ozone - an Emerging Technology for
These a Food Industry.”Brazilian Archives of Biology and
Technology 52:1527–39. doi:10.1590/S1516-
89132009000600025.
Gundarova,R.A.,I.P.Khoroshilova-Maslova,G.G.
Bordiugova, L. V. Ilatovaskaia, and I. M. Lapina.
1996. “Experimental Validation of Using Ozonized
Physiological Solutions in Intraocular Infection.”
Vestnikoftalmologii 112:9–11.
Hammer, K. A., C. F. Carson, and T. V. Riley. 2002. “In Vitro
Activity of Melaleuca Alternifolia (Tea Tree) Oil against
Dermatophytes and Other Filamentous Fungi.”Journal of
Antimicrobial Chemotherapy 50:195–9. doi:10.1093/jac/
dkf112.
Helal,G.A.,M.M.Sarhan,A.N.K.AbuShahla,andE.K.Abou
El-Khair. 2006. “Antimicrobial Activity of Some Essential Oils
against Microorganisms Deteriorating Fruit Juices.”
Mycobiology 34:219–29. doi:10.4489/MYCO.2006.34.4.219.
Kawamura, K., M. Kaneko, T. Hirata, and K. Taguchi. 1986.
“Microbial Indicators for the Efficiency of Disinfection
Processes.”Water Science and Technology 18:175–84.
Kim,H.S.,S.U.Noh,Y.W.Han,K.M.Kim,H.Kang,
H. O. Kim, and Y. M. Park. 2009. “Therapeutic Effects
of Topical Application of Ozone on Acute Cutaneous.
Wound Healing.”Journal of Korean Medical Science
24:368–74. doi:10.3346/jkms.2009.24.3.368.
Koc, A. N., M. A. Atalay, D. Timur, G. Demir, and L. Kaynar.
2016. “Molecular Epidemiology and Antifungal Susceptibility
of Saprochaetecapitata (Blastoschizomycescapitatus) Isolates
Causing Nosocomial Infection in Kayseri/Turkey.”The
Journal of Infectious Diseases 1–8. doi:10.1080/
23744235.2016.1176246.
Koc, A. N., S. Silici, F. Kasap, F. Kasap, H. T. Hörmet-Öz,
H. Mavus-Buldu, and B. D. Ercal. 2011. “Antifungal
Activity of the Honeybee Products against Candida
Spp.andTrichosporon Spp..”Journal of Medicinal
Food 14:128–34. doi:10.1089/jmf.2009.0296.
Kutlubay, Z., S. Baglam, B. Engin, and Y. Tuzun. 2013.
“Dermatolojide Ozon Tedavisi.”Turkiye Klinikleri
Journal Dermatol-Special Topics 6 (1):8–12.
Kutlubay, Z., B. Engin, S. Serdaroglu, and Y. Tuzun. 2010.
“Dermatolojide Ozon Tedavisi.”Dermatoz 1:209–16.
Leopoldina, F. L., M. C. Silvia, D. S. Ramón, G. O. Enrique,
M. D. Sonia, and A. G. Marlene. 1998. “Aceite Ozonizado
8K. VAROL ET AL.

En Dermatologia. Experiencia De 9 Anos.”Revista CENIC
Ciencias Biológicas 29:192–5.
Lezcano, I., N. Nuiez, M. Espino, and M. Gomez. 2000.
“Antibacterial Activity of Ozonized Sunflower Oil, against
Staphylococcus Aureus and Staphylococcus Epidermis..”
Ozone: Science & Engineering 22:207–14. doi:10.1080/
01919510008547221.
Lima, E. O., O. Fishman, A. M. Giesbrecht, and M. Q. Paula.
1993. “In Vitro Antifungal Activity of Essential Oils
Obtained from Officinal Plants against Dermatophytes.”
Mycoses 36:333–6. doi:10.1111/j.1439-0507.1993.tb00777.x.
Lima, I. D. O., R. D. A. G. Oliveira, E. D. O. Lima, N. M. P.
Farias, and E. L. D. Souza. 2006. “Antifungal Activity from
Essential Oils on Candida Species.”Revista Brasileira De
Farmacognosia 16 (2):197–201. doi:10.1590/S0102-
695X2006000200011.
Markin, D., L. Duek, and I. Berdicevsky. 2003. “In Vitro
Antimicrobial Activity of Olive Leaves.”Mycoses 46:132–
6. doi:10.1046/j.1439-0507.2003.00859.x.
Mcintyre, K. A., and J. N. Galgiani. 1989. “Ph and Other
Effects on the Antifungal Activity of Cilofungin
(LY121019).”Antimicrobial Agents and Chemotherapy
33:731–5. doi:10.1128/AAC.33.5.731.
Menendez, S., L. Falcon, D. R. Simon, and N. Landa. 2002.
“Efficacy of Ozonized Sunflower Oil in the Treatment of
Tinea Pedis.”Mycoses 45:329–32. doi:10.1046/j.1439-
0507.2002.00780.x.
Miceli, M. H., J. A. Diaz, and S. A. Lee. 2011. “Emerging
Opportunistic Yeast Infections.”The Lancet Infectious
Diseases11 142–51. doi:10.1016/S1473-3099(10)70218-8.
Moureu, S., F. Violleau, D. Ali Haimoud-Lekhal, and A.
Calmon. 2016. “Influence of Storage Temperature on the
Composition and the Antibacterial Activity of Ozonized
Sunflower Oil.”Ozone: Science & Engineering 38:143–9.
doi:10.1080/01919512.2015.1128319.
Ouf, S. A., T. A. Moussa, A. M. Abd-Elmegeed, and S. R.
Eltahlawy. 2016. “Anti-Fungal Potential of Ozone against
Some Dermatophytes.”Brazilian Journal of Microbiology
698–702. doi:10.1016/j.bjm.2016.04.014.
Perez, A., T. Poznyak, and I. Chairez. 2015. “Microorganism
Inactivation by Ozone Dissolved in Aqueous Solution: A
Kinetic Study Based on Bacterial Culture Lipid
Unsaturation.”Ozone: Science & Engineering 37 (2):119–
26. doi:10.1080/01919512.2014.917947.
Pfaller, M. A., and D. J. Diekema. 2010. “Epidemiology of
Invasive Mycoses in North America.”Critical Reviews in
Microbiology 36:1–53. doi:10.3109/10408410903241444.
Pryor, W. A., and R. M. Uppu. 1993. “A Kinetic Model for
the Competitive Reaction of Ozone with Amino Acid
Residues in Proteins in Reverse Micelles.”Journal of
Biological Chemistry 268:3120–6.
Richardson, M., and C. Lass-Flörl. 2008. “Changing
Epidemiology of Systemic Fungal Infections.”Clinical
Microbiology and Infection 14:5–24. doi:10.1111/j.1469-
0691.2008.01978.x.
Rodrigues, K. L., C. C. Cardoso, and L. R. Caputo. 2004.
“Cicatrizing and Antimicrobial Properties of an Ozonised
Oil from Sunflower Seeds.”Inflammopharmacology
12:261–70. doi:10.1163/1568560042342275.
Sakazaki, F., H. Kataoka, T. Okuno, H. Ueno, M. Semma, A.
Ichikawa, and K. Nakamuro. 2007. “Ozonated Olive Oil
Enhances the Growth of Granulation Tissue in a Mouse
Model of Pressure Ulcer.”Ozone: Science & Engineering
29:503–7. doi:10.1080/01919510701618205.
Sechi, L. A., I. Lezcano, and N. Nunez. 2001.
“Antibacterial Activity of Ozonized Sunflower Oil.”
Journal of Applied Microbiology 90:279–84.
doi:10.1046/j.1365-2672.2001.01235.x.
Silici, S., and A. N. Koc. 2006. “Comparative Study of in Vitro
Methods to Analyse the Antifungal Activity of Propolis
against Yeasts Isolated from Patients with Superficial
Mycoses.”Letters in Applied Microbiology 43:318–24.
doi:10.1111/j.1472-765X.2006.01949.x.
Skalska, K., S. Ledakowicz, J. Perkowski, and B. Sencio. 2009.
“Germicidal Properties of Ozonated Sunflower Oil.”
Ozone: Science & Engineering 31:232–7. doi:10.1080/
01919510902838669.
Tara, F., Z. Zand-Kargar, O. Rajabi, F. Berenji, F. Akhlaghi,
M. T. Shakeri, and H. Azizi. 2014. “Comparing the Effect
of Ozonated Olive Oil and Clotrimazole Cream for the
Treatment of Vulvovaginal Candidiasis.”Journal of
Alternative and Complementary Medicine 20:83.
doi:10.1089/acm.2014.5218.
Tellez, G., O. Lozano, and M. Gomez. 2006. “Measurement of
Peroxidic Species in Ozonized Sunflower Oil.”Ozone: Science
& Engineering 28:181–5. doi:10.1080/01919510600689356.
Thomson, M. P., C. S. Anticevic, B. H. Rodríguez, and V. V.
Silva. 2011. “In Vitro Antifungal Susceptibility, in Vivo
Antifungal Activity and Security from A Natural Product
Obtained from Sunflower Oil (AMO3) against
Dermatophytes.”Revistachilena De Infectologia 28:512–19.
Travagli, V., I. Zanardi, G. Valacchi, and V. Bocci. 2010.
“Ozone and Ozonated Oils in Skin Diseases: A Review.”
Mediators of Inflammation 2010:1–9. doi:10.1155/2010/
610418.
Ulu Kilic, A., E. Alp, F. Cevahir, Z. Ture, and N. Yozgat.
2016. “Epidemiology and Cost Implications of
Candidemia, a 6 Year Analysis from a Developing
Country.”Mycoses 1–6. doi:10.1111/myc.12582.
Wagner, D. K., and P. G. Sohnle. 1995. “Cutaneous Defences
against Dermatophytes and Yeasts.”Clinical Microbiology
Reviews 8:317–35.
Wu, J., H. Doan, and M. A. Cuenca. 2006. “Investigation of
Gaseous Ozone as an Anti-Fungal Fumigant for Stored
Wheat.”Journal of Chemical Technology & Biotechnology
81:1288–93. doi:10.1002/jctb.1550.
OZONE: SCIENCE & ENGINEERING 9