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REVIEW PAPER
Ozone Technology in Food Processing: A Review
VITHU PRABHA, RAHUL DEB BARMA*, RANJIT SINGH1, ADITYA MADAN
Indian Institute of Crop Processing Technology
GoI, MoFPI, Pudukkottai Road, Near Air-force Station, Thanjavur-613005, Tamilnadu, India
*email : dbarma.rahul@gmail.com
ABSTRACT
Interest in ozone has expanded in recent years in
response to consumer demands for ‘greener’ food
additives, regulatory approval and the increasing
acceptance that ozone is an environmentally friendly
technology. Ozone, a powerful oxidant, is effective
against various kinds of microorganisms on fruits,
vegetables, meat grains and their products. The multi-
functionality of ozone makes it a promising food
processing agent. Excess ozone auto decomposes
rapidly to produce oxygen and thus leaves no residues
in foods from its decomposition. Ozone as an oxidant
is used in water treatment, sanitising, washing and
disinfection of equipment, odour removal, and fruit,
vegetable, meat and seafood processing. Ozone
treatment assures the retention of sensory,
nutritional and physicochemical characteristics of
food. Treatment conditions should be specifically
determined for all kinds of products for effective and
safe use of ozone.
Key words Ozone, greener, additives, oxidant,
physicochemical.
Minimising pathogenic and spoilage
microorganisms in fruits, vegetables and their
products are a primary food-safety concern.
Traditionally thermal processing methods are used
to inhibit the pathogens. This technology, however,
affects the quality of foods. Non-thermal
processing prevents the food quality losses such
as loss of original flavour, taste, appearance, colour,
nutritional quality etc. In general, current
sanitization technologies are crucial to maintaining
the quality and enhancing the safety of fresh
agricultural commodities but it is required to
minimise the drawbacks and potentially hazards
caused by the treatments to consumers. Promising
results have been revealed in solving the problems
of the food industry like microbes, pests,
mycotoxin and pesticide residues by ozone
application. Spontaneous decomposition without
forming hazardous residues in the treatment
medium makes ozone safe in food applications. If
improperly used, ozone can cause some deleterious
effects on products, such as losses in sensory
quality.
Ozone is a strong oxidant and potent
disinfecting agent. Disinfecting agents have
widespread applications to assure safety and quality
in the food industry. However, some of these
agents, such as chlorine, are inefficient against
some organisms, particularly at high pH or against
spore-forming microbes. Furthermore, chlorine can
react to form trihalomethanes, which are of concern
for both human dietary safety and as environmental
pollutants. Therefore, the food industry is in search
of applications that are:
Effective in inactivation of common and
emerging pathogens, and removing toxic
contaminants.
Leading to less loss in product quality and
ensure ‘freshness’.
Adaptable to food processes and
economically feasible.
Environmental friendly
The bactericidal effects of ozone have been
documented on a wide variety of organisms,
including Gram positive and Gram negative bacteria
as well as spores and vegetative cells. There are
numerous application areas of ozone in the industry
such as food surface hygiene, sanitation of food
plant equipment, reuse of waste water, treatment
and lowering biological oxygen demand (BOD) and
chemical oxygen demand (COD) of food plant
waste.
Properties and Characteristics of Ozone
Ozone was discovered and named by
Schoenbein in 1840, but its applications for food
treatment did not develop until much later. Ozone
(O3) is tri-atomic oxygen formed by addition of a
free radical of oxygen to molecular oxygen. The
three atoms of oxygen in the ozone molecule are
Trends in Biosciences 8(16), Print : ISSN 0974-8, 4031-4047, 2015
4032 Trends in Biosciences 8 (16), 2015
arranged at an obtuse angle, whereby a central
oxygen atom is attached to two equidistant oxygen
atoms; the included angle is approximately116° 492
and the bond length is 1.278 Å. The boiling point
of ozone is “111.9 ± 0.3 °C, the melting point is
“192.5 ± 0.4 °C, the critical temperature is “12.1
°C and the critical pressure is 54.6 atm (Manley
and Niegowski, 1967). Ozone exists in the gaseous
state at room and refrigeration temperature and it
is partially soluble in water. At room temperature,
ozone is an unstable gas. Ozone readily degrades
(Manley and Niegowski, 1967) but has a longer
half-life in the gaseous state than in aqueous solution
(Rice, 1986). Ozone is relatively stable in air but
highly unstable in water, decomposes in a very short
time. It cannot be stored and must be generated
continuously. The only result of ozone, when it
decomposes, is oxygen; so, food products treated
with ozone are free of disinfectant residue. It is
readily detectable at 0.01–0.05 ppm level (Miller et
al., 1978). It has a pungent, characteristic odour
described as similar to “fresh air after a
thunderstorm.”Ozone is a blue gas at ordinary
temperature when generated from dried air, but
colourless when generated from high-purity
oxygen.
Purity of water usually affects ozone stability.
Although ozone in pure water degrades rather
quickly to oxygen, it degrades even more rapidly
in impure solutions. Hill and Rice (1982) reported
that approximately 50% of ozone is destroyed in
20 minutes at 20°C in distilled or tap water, whereas
only 10% of ozone breaks down in85 minutes in
20 °C double-distilled water. Ozone solubility in
water is13 times that of oxygen at 0–30 °C and it
is progressively more soluble in colder water (Rice,
1986). Ozone decomposition is faster in higher
water temperatures. Ozone is a toxic gas; toxicity
is dependent on concentration and length of
exposure (Pascual et al., 2007). At short-term
exposure rates of 0.1–1.0 ppm, symptoms include
headaches, nosebleeds, eye irritation, dry throat and
respiratory irritation. At higher exposure levels (1–
100 ppm), symptoms become more severe and
include asthma-like symptoms, tiredness and loss
of appetite.
Generation of Ozone
Ozone (O3) is formed by a high energy input
that splits the oxygen (O2) molecule in the air into
free radical oxygen. Single oxygen (O) molecules
rapidly combine with available O2 to form ozone (3
O2”!2 O3 +heat and light). In nature, the source of
this high energy is the ultraviolet irradiation from
the sun and also lightning discharge. Since ozone
is unstable, it splits back into oxygen molecule.
Ozone can be generated on-site as required by
several techniques, three of which are available
commercially at the present time – corona
discharge, UV radiation and electrolysis.
Corona discharge or plasma technique
The most commercially significant technique
is by corona discharge (the so-called “silent
electrical discharge” procedure). This is tantamount
to producing synthetic lightning. In a corona
discharge ozone generator, the feed gas (dried air,
oxygen, or mixtures thereof), passes between two
closely spaced electrodes (one of which is coated
with a dielectric material) under a nominal applied
potential of ~10 kV. A silent or barrier discharge
occurs when the gas becomes partially ionized,
resulting in a characteristic violet glow when air is
the feed gas (with high purity oxygen the violet
coloration is seldom observed).
There are two electrodes in corona discharge,
the high tension and low tension (ground)
electrodes, separated by a dielectric medium in a
narrow discharge gap. When electrons have
sufficient energy to dissociate the oxygen molecule,
a certain fraction of these collisions occur and a
molecule of ozone can be formed from each oxygen
atom. A schematic diagram of ozone generation by
corona discharge method is given in Fig. 3.1.
Efficiency of ozone production by corona
discharge depends on the strength of micro-
discharges which are influenced by a number of
factors such as the gap width, gas pressure,
properties of the dielectric and metal electrodes,
power supply, and the presence of moisture. In
weak discharges, a significant fraction of the energy
is consumed by ions, whereas in stronger
discharges, almost all of the discharge energy is
transferred to electrons responsible for the
formation of ozone. The optimum is a compromise
that avoids energy losses to ions but at the same
time obtains a reasonable conversion efficiency of
oxygen atoms to ozone.
If air is used as the feed gas, it must be
scrupulously dried and be free of traces of oils and
greases (oxidized by ozone). Moist air gives rise to
nitrogen oxides in the ozone generator which will
form nitric acid which will corrode the generator,
requiring frequent maintenance and down time. If
air is passed through the generator as a feed gas,
PRABHA et al., Ozone Technology in Food Processing: A Review 4033
Fig. 1. Schematic diagram of ozone generation by corona discharge method
1–3 % ozone is made; using high-purity oxygen
may yield as high as 16 % ozone (Rice et al., 1981).
Physico-chemical or ultraviolet radiation
The mechanism of photochemical production
of ozone is similar to that which occurs in the
stratosphere, that is, oxygen atoms formed by the
photo-dissociation of oxygen by short wavelength
UV radiation react with oxygen molecules to form
ozone. Although the theoretical quantum yield of
ozone by this technique is 2 %, in practice the actual
yield is more on the order of 0.5 %, because the
low pressure mercury lamps produce not only the
185 nm radiation responsible for the production of
ozone, but also the 254 nm radiations that destroy
ozone. Medium pressure UV that produces higher
levels of 185 nm radiation produces more ozone.
An advantage of generating ozone by UV radiation
is that ambient air can be used efficiently as the
feed gas. On the other side, quantities of ozone
generated per 40-W UV bulb are low (0.5 g/h) at
maximum concentrations of 0.25 wt %. However,
these maximum ozone yields and concentrations
cannot be attained simultaneously by the UV method.
The low concentrations of ozone available from
UV generators limit their applicability for water
treatment to special applications. However, their
use to generate ozone for air treatment can be
effective.
Electrolysis
High current density electrolysis of aqueous
phosphate solutions at room temperature produces
ozone and oxygen in the anodic gas. Electrolysis
of 68 wt-% sulfuric acid can produce 18-25 wt-
% ozone in oxygen when a well-cooled cell is used.
Although electrolysis of water can produce high
concentrations of ozone, the output is low, and the
cost is several times more than that of the corona
discharge process. However, small electrolytic units
are being used commercially for treatment of ultra-
high purity waters in pharmaceutical and electronic
industries.
Components of Ozone System
An ozonation system takes up air or feed gas
from the atmosphere and concentrates oxygen for
ozone generation. Ozone is generated, its
concentration is analysed and passed to the
treatment chamber (collector) for microbial
inactivation. The excess ozone from treatment
chamber is released into air as oxygen to prevent
chance of health hazards. A complete ozonation
system (gaseous ozone) for use in food processing
plants consists of the following subunits:
Oxygen concentrator
For food processing plants, oxygen-enriched
air (>90% O2) is provided simply and conveniently
by means of oxygen concentrators. These devices
take in ambient air, automatically filter (remove dust
particles), then separate and remove nitrogen
(thereby leaving air considerably enriched with
oxygen), which is also dried to below the desired
maximum dew point (“4°C), all at the same time in
one small device. These oxygen concentrators
operate on the principle of pressure swing
adsorption (PSA) drying. An air compressor
pressurises the airflow and sends it through a
molecular sieve (microscopic porous bead) bed that
adsorbs or traps nitrogen and moisture, while
providing oxygen-enriched air to the supply output
of the concentrator. As the molecular sieve bed
becomes loaded with nitrogen and moisture, they
4034 Trends in Biosciences 8 (16), 2015
desorb to waste in vapour form to the environment,
recovering the adsorption capacity of the sieve bed.
Today the majority of ozone generating systems
use oxygen as the feed gas because the
concentrations of ozone produced are increased
two to three times for the same energy expenditure.
Most ozone equipment manufacturers have
optimised their equipment for food processing
plants to operate on oxygen feed gas.
Ozone generator
Ozone generators may be UV generators or
Corona discharge generators. The quality of gas
fed to an ozone generator can be a critical factor,
particularly if ozone is produced by CD or plasma
techniques. So CD generators require an air
preparation prior to ozone generation. The ozone
output is only slightly increased by increasing the
oxygen content or drying the air. The two common
types of gas preparation to feed CD ozone
generators are oxygen and dry air. UV generators
of ozone do not require any special air preparation.
If an air dryer is selected to feed a CD ozone
system, make sure that the air preparation
equipment is matched and sized to the ozone. Each
ozone generator is designed to operate at an optimal
flow rate depending upon size of ozone generator,
and such information should be stated in the
supplier’s equipment manual. With moisture present
in the feed gas, the very corrosive nitric acid
(HNO3) is readily formed. Consequently, the gas
feeding an ozone generator must be very dry
(maximum “54 °C dew point), because the
presence of moisture also affects ozone production,
as well as leading to the formation of nitric acid.
This very corrosive acid can destroy the internal
parts of a CD ozone generator, which can cause
premature system failure and will increase the
frequency of required maintenance.
When using oxygen as the feed gas, the
concentration of ozone produced is increased two
to three times over ozone concentrations in dried
air for the same energy expenditure. For example,
dry air-fed CD generators of the size used in food
processing plants produce 1–2 wt% ozone, but
when fed high-purity oxygen, ozone concentrations
of 3–6 wt% are produced for the same energy
expenditure. Most ozone equipment manufacturers
have optimised their equipment for oxygen feed
gas. In some instances, ozone generators can
produce as high as 20 wt% ozone (from oxygen),
but the energy and other requirements are higher
to produce ozone at this concentration. Properly
designed and operated CD-ozone generators
commercially available today are capable of
producing kg/h quantities of ozone in gas phase
concentrations of 1-5% by weight in air and up to
14% by weight in high purity oxygen. The advantage
of a higher weight per cent ozone product is
economics.
The process of electrically rupturing oxygen
atoms to produce oxygen ions or atoms that
combine with more oxygen to produce ozone
liberates considerable heat. This heat generated
during the process must be removed from the
generator to avoid the reverse reaction (ozone
reverting to oxygen) from taking over and
decreasing the efficiency of ozone generation.
Consequently cooling of the corona discharge
ozone generator becomes a critical component in
generator design. Normally cooling is provided by
water, but there are air cooled CD- ozone generators
commercially available, especially on smaller scale,
appropriate for many food applications.
Flow meters
Mass flow meter provides accurate
measurement of total oxygen gas flow from the
oxygen concentrator to ozone generator. Ozone
flow meter use small gas stream (less than 2 LPM)
to measure ozone concentration and for regulation
of gas flow through ozone analyser.
Treatment chamber
It is the collector where ozone enters from
the top and returns though the bottom outlet after
completing the exposure time. The chamber is air
tight and avoids leakages. The dimension and type
will be based on the mode of application of ozone
and type of product being processed. The outlet is
connected to ozone analyser and then to ozone
destructor.
Ozone analyser
This device will be used for measuring the
ozone concentration in per cent by weight, or g/m3
that enters and leaves the treatment chamber.
Ozone destructor
At the outlet of the destructor, excess ozone
is destroyed, and the cleaned and decontaminated
air is re-circulated to its intended enclosure or
discharged to the ambient atmosphere. Excess
ozone can be broken down into oxygen and send
to the atmosphere in order to prevent any harm for
PRABHA et al., Ozone Technology in Food Processing: A Review 4035
the worker. Ozone at high concentration is
corrosive and toxic even at lower ppm. In this
device, the excess ozone is allowed to release into
the air such that the contact of both is controlled.
Since ozone is unstable in air, decomposition of
ozone takes place and oxygen is given out through
the outlet. Destructor is an important part that
assures non-hazardous by-product or waste in
ozone treatment.
Data and analysis
The data such as exposure time, ozone
concentration can be acquired using ozone analyser
and used for analysis of results.
Modes of Application
There are many modes of application of ozone
to the food product. The mode of application may
be selected based on the type of food and the need
of ozone treatment. Ozone generators are available
based on the type of feed gas available using for
ozone generation. The concentration and intensity
varies with different modes used for the same
effect. D.M Graham (2000) explains the mode of
application of ozone as follows.
Application in aqueous phase
Ozone forms true solution with water. The
solubility of ozone in water is based on Henry’s
law, which states that ‘pressure applied to a vapour
in equilibrium with a liquid is inversely proportional
to the temperature’. Half-life of ozone in aqueous
solution is less than that of gaseous ozone; so
decomposes rapidly. The decomposition of ozone
is very rapid in the presence of impure water. Ozone
is only partially soluble in water; efficient transfer
of ozone into solution requires the dispersion of
gaseous ozone into small bubbles. This is
accomplished in various types of positive pressure
ozone contractors such as bubble diffuser/bubble
columns, mechanically agitated vessels, turbine
mixers, tubular reactors, in-line static mixers as
well as negative pressure reactors (venturi) and
injectors. In positive pressure devices, ozone gas
under pressure exiting the ozone generator is forced
through small apertures supported under the water.
With negative pressure devices, ozone gas is drawn
into the flowing water stream to be treated. Under
these mixing conditions, bubbles are sheared and
mixed thoroughly with the aqueous fluid, decreasing
the liquid film thickness but increasing both
interfacial area and contact time. Faster ozone mass
transfer rates result in faster disinfection and
(usually) oxidation rates. However, slow-to-oxidize
organics are unaffected by increased ozone mass
transfer rates, and advanced oxidation techniques
should be considered for these types of materials.
Ozone transfer efficiencies vary with the
number of contacting stages and typically are above
90%. However, since even a 95% ozone absorption
efficiency can result in a contactor off-gas
containing as much as 740 ppm (by wt) of ozone
(based on 1.5 wt-% ozone in air feed gas), treatment
is required to reduce the ozone concentration to an
acceptable maximum level for discharge to the local
environment. This can be accomplished thermally
and/or by catalytic means, and sometimes (for low
concentrations and/or small volumes) by passage
through wet granular activated carbon beds.
Application in the Gas Phase
Reaction rates of contaminants in air are much
lower (slower) in the gas phase than in aqueous
phase. Additionally, the option of adjusting pH is
not open in air spaces, although increasing the
relative humidity is an effective option. Pumping
ozone gas from the generator into an air space to
be treated is the simplest approach, and is most
effective when the air contaminants to be treated
are rapidly reactive with ozone – e.g., many
odoriferous compounds such as hydrogen sulfide,
molds, spores, and some airborne microorganisms.
For those air contaminants that are only slowly
affected by ozone, the accepted procedure is to
draw contaminated air into an enclosed structure
in which ozone is mixed with the contaminated air
for such period of time as is necessary to destroy
(or inactivate) the contaminants.
Application of Adjuncts With Ozone
With the advent of ozone advanced oxidation,
there are recent instances reported of coupling
ozone with either hydrogen peroxide or with
ultraviolet radiation, techniques that are designed
to promote the formation of hydroxyl free radicals
with the stated objective of increasing the amount
of microbiocidal activity above that of ozone itself.
However, all information developed to date indicates
that the half-life of hydroxyl free radicals in water
is only microseconds in length, and that the
maximum concentration of hydroxyl free radicals
that has been measured in aqueous solution is very
low, on the order of 10-12 M. These facts tend to
indicate that there can be no microbiocidal benefit
of hydroxyl free radical over that provided by
4036 Trends in Biosciences 8 (16), 2015
molecular ozone. Nevertheless some recent reports
indicate what appears to be a synergy – or an
increased amount of microbial inactivation by
applying these combinations to certain foodstuffs
over what is obtained by applying ozone or UV
radiation alone. For example, Naitoh, 1992
investigated synergistic sporicidal activities of
gaseous ozone and UV irradiation.
Ozone + Hydrogen Peroxide
With this combination of oxidants in aqueous
solution, both agents destroy each other, and both
agents give rise, eventually, to hydroxyl free
radicals. It is customary procedure to add the
requisite amount of hydrogen peroxide to solution
and then pass that solution through an ozone
contacting apparatus. Ozone reacts immediately
with hydrogen peroxide in solution, and if the
amount of ozone dosed in the contactor is always
greater than the amount of peroxide initially added
to solution, at the outlet of the contact chamber it
will not be possible to measure a level of residual
ozone in solution. In advanced oxidation practice,
it has been learned that for optimum oxidative
performance, each pollutant that needs to be
destroyed requires a specific weight ratio of
peroxide to ozone. It is advisable when evaluating
the use of this advanced oxidation process to first
determine which polluting constituents of the water
or waste water requiring treatment are present, and
then to determine experimentally the optimum range
of peroxide to ozone weight ratio required for their
destruction. Glaze et al., 1987 have shown that if
the weight ratio of peroxide to ozone rises above
1:1, the rates of oxidation of organics in water
actually slowly down. This means that if excess
hydrogen peroxide is present over the amount of
ozone added, at least some of the advantages of
advanced oxidation are lessened. It also means that
there will be no molecular ozone present at any
time during ozone contacting for microbial
disinfection.
Ozone + UV Radiation
With this combination of agents, it is
customary to place a UV bulb (or multiple bulbs)
in the ozone contacting chamber. As water flows
through the chamber, first the UV bulb(s)
is(are)turned on and ozone is added. As long as the
amount of UV radiation dosed is in excess of the
amount of ozone present, all ozone will be converted
instantaneously to decomposition products, ending
rapidly as hydroxyl free radicals.
Despite the theories and the work of Kruith
of and Kamp (1999), several reports have been
made of an apparent increase in antimicrobial
activity in some food applications when ozone is
combined in water with either peroxide or UV
radiation.
Advantages and Limitations
The advantages and limitations of ozone
disinfection methods proposed for fresh-cut organic
vegetables, fruits and meat products applications
(Olmez and Kretzschmar, 2009) are as follows.
Advantages of ozone treatment
o High antimicrobial activity compared to non-
oxidative biocins (chlorine) in terms of
concentration and time.
o Short contact time for disinfection compared
to other disinfection methods
o No residue problem as it is completely utilised
and get reduced
o Non-hazardous at low ppm (lower than 4 ppm)
and effective in bactericidal uses
o No need to store hazardous substance
compared to other sanitation methods
o Lower running costs, cost matters only to
filling of oxygen cylinders and power supply.
o No heat requirement and no heat generation
in treatment(applicable to heat sensitive foods)
&thus saves need of input energy
o Saves transport of disinfectant chemicals&
storing of gas cost
o Eco-friendly and economically feasible
technology
Limitations of ozone treatment
o Ozone is toxic; when inhaled it cause throat
and nasal problems, even lead to asthma
o Ozone is highly unstable gas so controlled
release on requirement is to be established
o Recontamination problems in clean in process
pipes as ozone decomposes completely within
a short duration
o Corrosive at high ppm (higher than 4 ppm),
care should be taken in using ozone and
releasing to treatment chamber
o It requires regular monitoring in indoor
applications for any leakages
o Higher initial investment for the generation
equipment
o Onsite generation is required as it is unstable
PRABHA et al., Ozone Technology in Food Processing: A Review 4037
and not suitable for storage
o Storage of ozone is not possible as it
decomposes quickly
o It can be mostly surface treatment as ozone
decomposes in short time and it is liable to
oxidation with organic matter
Factors Affecting Efficacy of Ozone
Processing
There are different parameters that affect the
disinfection ability of ozone in liquid processing
treatment. Extrinsic and intrinsic parameters that
affect the ozone efficacy include flow rate, ozone
concentration, temperature, pH, and presence of
solid contents (organic matter).
Extrinsic Parameters
Flow Rate
Depending on the gas flow rate applied for
ozone production, different bubble sizes are
produced. Bubble size has been shown to have an
effect on ozone’s solubilisation rate and disinfection
efficacy. Ahmad and Faroq, 1985 reported that
ozone mass transfer and disinfection efficacy
increased as bubble size decreased (ozone bubble
size was varied while all other factors were kept
constant). The higher interfacial area available for
mass transfer at the smaller bubble size may have
been responsible for this effect. Decreasing the
bubble diameter from 1 cm to 0.1 cm increases
the contact area by 32 times (Ogden, 1970). Free
suspended bacteria migrate toward the ozone
bubbles due to their surface active properties and
are preferentially inactivated by comparatively high
ozone concentrations at the gas liquid interface of
the bubble (Hill and Spencer, 1974).
Concentration
Ozone concentration present or available in
the medium is another parameter determining ozone
efficacy. Increased ozone concentration causes
saturation and thus makes addition of further ozone
to the reactor ineffective, resulting in longer times
to achieve the same log-reduction values.
Temperature
Ozone solubility in water is 13 times that of
oxygen at 0-30°C and it is progressively more
soluble in colder water (Rice, 1986). The solubility
ratio for ozone increases as the temperature of
water decreases (Bablon et al., 1991). As
temperature increases, ozone becomes less soluble
and less stable, with an increase in the decomposition
rate (Rice et al., 1981). The mass transfer of ozone
gas into the liquid phase is also influenced by
temperature and pH. The inactivating capabilities
of ozone are in line with the decreasing temperature
(Farooq et al., 1977).
Intrinsic Parameters
pH
The effect of pH on ozone inactivation is
mainly attributed to the fact that ozone
decomposition rate changes substantially with
changes in pH (Farooq et al., 1977b; Roy et al.,
1980). Patil et al., 2010a observed that ozone
inactivation of E. coli was much faster at the lower
pH. The ozone treatment duration required for
achieving a5-log reduction was 4 min at the lowest
pH and 18 min at the highest pH studied.
Organic Matter
Ozone demand can be caused by certain
organics, inorganics, or suspended solids.
Dissolved organic matter reduces the disinfection
activity by consuming ozone to produce compounds
with little or no microbiocidal activity, thereby
reducing the concentration of active species
available to react with microorganisms. Williams
et al., 2005 studied the inactivation of E. coli in
orange juice, and found that the efficacy of
ozonation was reduced in the presence of ascorbic
acid and organic matter. In wastewater, proteins,
carbohydrates, lipids, and organic amines will
elevate the concentration of dissolved organic
carbon. Oxidizing disinfectants like ozone will lose
bacteriocidal strength through reaction with organic
matter. The reaction products will generally have
weak or no bacteriocidal activity.
Microbial Inactivation by Ozone
Microbial load of raw material, improper
handling and storage, use of contaminated wash
water, processing equipment, and transportation
facilities, as well as cross-contamination from other
products contribute to the microbial hazards
associated with meat, fruits and vegetables. Ozone
destroys microorganisms by the progressive
oxidation of vital cellular components. The
antimicrobial activity of ozone is based essentially
on its powerful oxidizing effect, which causes
irreversible damage to the fatty acids in the cell
membrane and to cellular macromolecules, such
as proteins, and DNA (Fettner and Ingols, 1959;
4038 Trends in Biosciences 8 (16), 2015
Hoffman, 1971; Naitoh, 1994). This action is
particularly effective in air at high relative humidity,
the bacteria being killed by ozone more readily in
the swollen state than when dry. The bacterial cell
surface has been suggested as the primary target
of ozonation. Microorganisms are inactivated by
disruption of the cell envelope or disintegration
leading to cell lysis.
Two major mechanisms of ozone destruction
of the target organisms were identified:
(1) Ozone oxidises sulfhydryl groups and amino
acids of enzymes, peptides and proteins to
smaller peptides
(2) Ozone oxidises polyunsaturated fatty acids to
acid peroxides (Victorin 1992).
Some authors concluded that molecular
ozone is the main inactivator of microorganisms
(direct oxidation), while others emphasize the
antimicrobial activity of the reactive by-products
of ozone decomposition (indirect oxidation) such
as OH, O2–, and HO3 (Chang 1971; Harakeh and
Butler 1985; Glaze and Kang 1989; Hunt and
Marinas 1997). Both molecular ozone and the free
radicals produced by ozone breakdown play a part
in this inactivation mechanism but there is no
consensus on which is more decisive. It has not
been well established whether molecular ozone or
the radical species are responsible for inactivation
of microorganisms. Thus there are two main
reactions happening on incidence of ozone on the
microbial surfaces viz. direct and indirect reactions.
Ozone takes any of the pathway or both for the
oxidation of sulfhydryl group of enzymes, amino
acids of peptides, proteins and enzymes and
polyunsaturated fatty acids.
Direct reaction
Direct reaction with molecular ozone, is the
predominant mechanism for inactivation of
microorganisms (Finch et al., 1992; Labatiuk et
al., 1994; Hunt and Mariñas, 1997). It is the direct
oxidation of target groups in microbial cell by
ozone. It is likely that the relative importance of
direct and indirect reactions with ozone in
determining microbial inactivation responses will
vary between microorganisms (Blatchley and Hunt,
2002). Because of the molecular structure of ozone,
it can act as an electrophilic or nucleophilic agent
during reactions (von Gunten, 2003), with these
types of reaction occurring in solutions containing
organic pollutants (with microbes). Generally,
electrophilic reactions will occur with organic water
contaminants with a high electron density and will
act faster in solutions consisting of high levels of
aromatic compounds (Gottschalk et al., 2010).
Nucleophilic reactions take place mainly when there
is a shortage of electrons and particularly at carbon
compounds that contain electron-withdrawing
groups such as –COOH and –NO2. However, for
these groups the reaction speed is much lower.
Overall, the direct oxidation of organic matter by
ozone involves a quite selective reaction mechanism
(von Gunten, 2003). Moreover, it is important to
note how the pH value of the water system can
influence ozone decomposition; with pH > 7 causing
an increase in the rate of ozone decomposition. Also,
in strongly acidic solutions (pH < 3) the OH radicals
do not influence the decomposition of ozone.
Indirect reaction
Indirect reactions with radicals are responsible
for inactivation for some groups of microbes
(Bancroft et al., 1984). Ozone decomposition has
been explained as occurring in three stages;
initiation, promotion and inhibition. During the
initiation step, free radicals are generated, such as
superoxide radical ions and hydroperoxide radicals,
which lead to formation of the highly reactive
hydroxyl radical. These hydroxyl radicals are one
of the factors contributing to ozone decomposition.
The promotion step involves regeneration of the
hydroperoxide and superoxide radicals through
reactions involving participation of promotors such
as formic acid, glyoxylic acid, primary alcohols
and aryl groups. In contrast, in the inhibition step
the consumption of hydroxyl radicals occurs via
ions like bicarbonate, carbonate, tertiary alcohols
and alkyl groups, without regeneration of the
superoxide radical ion (Staehelin and Hoigné, 1985;
Khadre et al., 2001). Bicarbonate ions are generally
present in microbial cells, which could act as
scavengers of radicals otherwise responsible for
inactivation of microorganisms.
Additionally, factors promoting ozone
decomposition in the system can lead to faster
dissipation of ozone, resulting in a requirement for
increased ozone concentration in order to achieve
the desired inactivation level (Zuma et al., 2009).
The resultant disruption or lysis of cell walls
(probably by oxidative destruction) associated with
ozone is a faster inactivation mechanism than that
of other disinfectants, which require the disinfecting
agent to permeate through the cell membrane in
order to be effective (Pascual et al., 2007). Scott
and Lesher (1963) reported that ozone caused
PRABHA et al., Ozone Technology in Food Processing: A Review 4039
leakage of cell contents into the medium and lysis
of some cells. Therefore, ozone-demanding
substances are generated during the ozone
inactivation process. Finch and others (1988) found
that E. coli cells demanded 0.06 mg/L ozone after
lysis and attributed the second phase of inactivation
to this ozone-created demand (Kim and Yousef,
2000). Generally, with regard to the spectrum of
microbial action, each microorganism has an
inherent sensitivity to ozone. Bacteria are more
sensitive than yeasts and fungi. Gram-positive
bacteria are more sensitive to ozone than Gram-
negative organisms, and spores are more resistant
than vegetative cells. Due to the mechanism of
ozone action, which destroys the microorganism
through cell lysis, the development of resistance to
ozone disinfection is not found (Pascual et al.,
2007).
Target sites of ozone activity
Inactivation of bacteria by ozone is a complex
process because ozone attacks numerous cellular
constituents including proteins, unsaturated lipids
and respiratory enzymes in cell membranes,
peptidoglycans in cell envelopes, enzymes and
nucleic acids in the cytoplasm, and proteins and
peptidoglycan in spore coats and virus capsids.
Cell envelope
Ozone may oxidize various components of
cell envelope including polyunsaturated fatty acids,
membrane- bound enzymes, glycoproteins and
glycolipids leading to leakage of cell contents and
eventually causing lysis (Scott and Lesher, 1963;
Murray et al., 1965). When the double bonds of
unsaturated lipids and the sulfhydryl groups of
enzymes are oxidized by ozone, disruption of
normal cellular activity including cell permeability
and rapid death ensues. Dave (1999) found that
treatment of Salmonella enteritidis with aqueous
ozone disrupted the cell membranes as seen in
transmission electron micrographs. However,
Komanapalli and Lau (1996) found that short-term
exposures of E. coli K-12 to ozone gas
compromised the membrane permeability but did
not affect viability, which progressively decreased
with longer exposure.
Bacterial spore coat
Foegeding, 1985 found that bacterial spores
(Bacillus cereus) with coat proteins removed were
rapidly inactivated by ozone, compared to intact
spores. The researcher concluded that the spore
coat is a primary protective barrier against ozone.
Recently, Khadre and Yousef, 2001 found that
spores of Bacillus subtilis treated with aqueous
ozone showed heavily disrupted outer spore coats.
Transmission Bacillus spores treated with ozone
suggest that ozone inactivates spores by degrading
the outer spore component (spore coat layers make
up approximately 50% of the spore volume), thus
exposing the cortex and core to the action of ozone
(Foegeding, 1985; Khadre et al., 2001). Young and
Setlow, 2004 determined that ozone does not kill
spores by DNA damage but rather by damaging
the ability of the spores to germinate. The
researchers hypothesised that damage to the inner
membrane of spores causes defects in spore
germination.
Enzymes
Several authors referred to enzyme
inactivation as an important mechanism by which
ozone kills cells. Sykes, 1965 reported that chlorine
selectively destroyed certain enzymes, whereas
ozone acted as a general protoplasmic oxidant.
Ingram and Haines, 1949, on general destruction
of the dehydrogenating enzyme systems in the cell,
proposed that ozone kills E. coli by interfering with
the respiratory system. Takamoto and others, 1992
observed that ozone decreased enzyme activity in
E. coli at a greater degree in case of cytoplasmic â-
galactosidase than in case of the periplasmic alkaline
phosphatase. Inactivation of enzymes by ozone is
probably due to oxidation of sulfhydryl groups in
Cysteine residues (Chang 1971).
Nucleic material
Reaction of aqueous ozone with nucleic acids
in vitro found that it may damage nucleic material
inside the cell. Ozone modified nucleic acids in vitro,
with thymine being more sensitive than cytosine
and uracil (Scott, 1975; Ishizaki et al., 1981). In
another study, ozone opened the circular plasmid
DNA and reduced its transforming ability, produced
single double-strand breaks in plasmid DNA
(Hamelin, 1985), and decreased transcription
activity (Mura and Chung 1990). Herault and Chung,
1984 found that ozone may induce mutations.
Compared to other known mutagens, ozone was
found to be a weak mutagen on Saccharomyces
cerevisiae (Dubeau and Chung, 1982).
Ozone reactions against virus
Sproul and Kim, 1980 found that aqueous
ozone inactivated bacteriophages (f2 and T4) by
4040 Trends in Biosciences 8 (16), 2015
attacking capsid protein, with liberation and
inactivation of the nucleic acid. The RNA from
bacteriophage (f2) was partially inactivated prior
to release from the capsid. They suggested that
ozone breaks the protein capsid into subunits
liberating RNA and disrupting virus adsorption to
the host pili, and that the RNA may be secondarily
inactivated. The DNA released from bacteriophage
(T4) was rapidly inactivated by ozone at about the
same rate as that in the intact phage. CK Kim et
al., 1984 found that ozone randomly destroyed the
head, collar, contractile sheath, end plate, and tail
fibers and liberated the DNA from the head.
Yoshizaki and others, 1988 and Shriniki and others,
1988 concluded that the major cause of tobacco
mosaic virus (TMV) inactivation by ozone was the
inability of the treated virus to uncoat. Roy and
others, 1981 found that ozone altered two of the
four polypeptide chains in the poliovirus protein
coat. They, however, attributed the inactivation of
the virus to the damage in its RNA by ozone. The
observation by Herbold and others, 1989 confirmed
the hypothesis that damage to viral envelopes is
the main cause of inactivation of viruses by ozone.
Enveloped viruses such as HIV are expected to be
much more resistant to ozone compared to non-
enveloped viruses such as poliomyelitis.
Objectives of Ozone in Food Industries
Ozone is an accepted commercial technology
in many aspects of the foods industry, ranging from
irrigation and soil treatment (Parmenter et al.,
2004), to spraying crops to avoid spraying noxious
chemicals (Steffen and Rice, 2008), odour control
in animal housing (Parmenter et al., 2004) and for
uses in food processing plants such as water and
air treatment, food decontamination and
disinfection, safe packaging and storage etc. In
general, ozone finds wide application in the food
industry, including surface decontamination of
meat, fruits and vegetables, drinking water
disinfection, pesticide removal, safe storage and
wastewater treatment (Guzel-Seydim et al., 2004;
Karaca and Velioglu, 2007).
Ozone is applied in either gaseous or aqueous
form. Ozone is very effective against bacteria
because even concentrations as low as 0.01 ppm
aretoxic to bacteria. Whereas disinfection of
bacteria by chlorine involves the diffusion of HOCl
through the cell membrane, disinfection by ozone
occurs with lysing (i.e., oxidative rupture) ofthe
cell wall. Disinfection rates by ozone, however,
depend on the type of organism and are affected
by ozone concentration, temperature, pH, turbidity,
the presence of ozone-oxidizable materials, the
tendency (or not) for the microorganisms to form
clumps, and the type of ozone contact or employed
(Zhu et al., 1989). The presence of ozone-
oxidizable substances in water exerts an ozone
demand, and this can retard disinfection until the
initial ozone demand has been satisfied, at which
point rapid disinfection is observed. Its efficacy
against a wide range of microorganisms, including
bacteria,fungi, viruses, protozoa and bacterial
fungal spores, has been reported(Restaino et al.,
1995; Khadre et al., 2001; Cullen et al., 2009).Such
advantages make ozone attractive to the food
industry and consequently it has been affirmed as
Generally Recognised as Safe (GRAS) for use in
food processing(Graham, 1997) and was approved
as an antimicrobial food additive in 2001 (FDA,
2001).Several incidents of food borne disease have
been associated with fruit and vegetable products.
Some of the ozone uses in food industries as
given by Bharathraj (2000) are as follows.
Removal of contaminants
Fungicides, pesticides and other chemicals
use during farming practises can contaminate the
surface of food. This can be potentially dangerous
as simple washing cannot remove these
accumulations. Ozone can be used to oxide the
chemicals and remove the contaminate safe for sale
or for further processing.
Cleaning in process (CIP)
Ozone charged water is used for CIP for
cleaning pipes, tanks floors, surface equipment etc.
Use of ozone system to food processing is that it
provides the opportunity to reuse the water that
could bring about a lot of savings in terms of
availability and water cost.CIP washing of plant
processing equipment and drains with ozone-
containing water is now common practice
(Parmenter et al., 2004; Lowe, 2002).
Sanitation
Food for consumption must be free from
pathogenic microorganisms. Contamination can
occur from harvesting stage, during transportation,
from processing water, equipment or from human
interventions or by cross contamination. Usually
non- oxidative biocides such as chlorine were widely
used for 2-log unit reduction of microorganisms.
Chlorine, the most common used disinfecting agent,
selectively destroys certain intracellular enzyme
PRABHA et al., Ozone Technology in Food Processing: A Review 4041
systems, whereas ozone causes widespread
oxidation of internal cellular proteins causing rapid
cell death. Chlorine, however, is not effective for
virus. Also it takes more concentration and exposure
time for microbial reduction as compared to ozone
since the mechanism is by penetration through the
membrane. Ozone is an oxidative biocide and is
better than non- oxidative to avoid undesirable taste
or carcinogenic effects. Ozone is 3000 times
powerful than chlorine and regarded as Generally
Recognised as Safe (GRAS). Commercial
processing applications of ozone expanded in
processing water treatments in the near future
especially in fish hatcheries (Blogoslawski et al.,
1993; Brazil and Summerfelt, 2005), beverage-
producing plants, and wineries (Steffens,
2006).The use of ozone for the sanitation of
equipment and surfaces in the beverage
manufacturing industry has yielded impressive
results in terms of controlling microorganisms and
saving costs due to less chemical handling and less
maintenance (Clear Water Tech, 2002; Hampson,
2000; Tinney, 2002).
Stop ripening and spoilage
Consideration of fresh food requirement
standards for consumers should assure fresh and
safe food products. Ethylene formation on the food
surface is responsible for the food ripening and
spoilage. By virtue of its chemical properties, ozone
prevents ethylene formation and thereby retards
ripening and spoilage by microorganisms. This
property extends the shelf life of food.
Ozone oxidises ethylene completely and
leaves carbon dioxide, water and oxygen. The
reaction is as follows.
C2H4+ 6O3
→
2CO2 + 2H2O + O2
Cold storage
Utilisation of the properties makes ozone
eminently suitable for increasing the storage life of
perishables foods in refrigerated premises. At the
same time, it is economic as the investment and
operational cost of the equipment are on an
acceptable level in relation to the size of refrigeration
rooms. Its application eliminates the risk of
unpleasant odours or other traces of antiseptics used
for preservation of food stuffs. During storage
ozone exerts a threefold effects by destroying the
microorganisms, oxidising odours and affecting the
processes of metabolism. Its primary action is mold
free surface and has only slight depth of penetration.
Increasing the moisture content of the environment
favourably influences the germicidal effect. This
brought about by the swelling of microbes making
them more susceptible to destruction. Experiments
conducted with beef showed that ozone is most
efficient if the surface3 has a definite moisture
content of around 60 %.
Current Research and Applications in
Foods
Consumer preference for minimally processed
foods and foods free of chemical preservatives, as
well as recent outbreaks of food borne pathogens,
identification of new food pathogens have all
stimulated demand for novel food processing and
preservation systems Minimising the occurrence
of pathogenic and spoilage microorganisms in
fruits, vegetables and their products is a primary
food-safety concern. For this purpose, the
incorporation of ozone (by itself or in combination
with other technologies) resulted either in improved
product quality, significant costs savings, or both.
Ozone in sea food and shell fish
processing
Violle, 1929, Salmon and LeGall, 1936, Fauvel
et al., 1979 developed the use of ozone for shell
fish depuration from laboratory stage to full
commercial installations in Southern Europe.
Depuration is a process whereby shellfish, freshly
harvested from their natural environments, are
placed for several days in storage chambers,
through which clean, pathogen-free water is passed.
Ozonated water was used for depuration. Over a
several day time period, the molluscs cleanse
themselves by passing disinfected water through
their systems, thus eliminating pathogenic
microorganisms imbibed from their natural
environments.
Abatement of ethylene by ozone
Potential storage life of citrus fruits with fairly
good appearance and eating quality can be obtained
if fruits are stored under the most optimum
conditions after harvest. Ethylene, also known as
stress hormone, has a special role in fruit humidity,
ripening and senescence, and therefore has its own
importance in postharvest management of citrus.
Citrus fruits have a very low rate of ethylene
evolution, in the amount of 0.1 mL/kg/h. Even this
rate can slowly build up ethylene concentration in
closed chambers (Ladaniya, 2007). Ozone has
shown a potential to meet this criterion and given
4042 Trends in Biosciences 8 (16), 2015
encouraging results. Skog and Chu, 2001 claimed
that ozone could reduce the level of ethylene in the
air in a cold storage room. Indeed, ozone was found
to be effective in removing ethylene from export
containers (Palou et al., 2001).
Many factors such as freezing, drying or high
carbondioxide concentrations can increase ethylene
evolution from agricultural products. Controversial
results are found in the literature about ozone from
this point of view. Actually, ozone is known to
impose oxidative stress and cause many
physiological changes, including ethylene synthesis
in crops (Forney, 2003).
During ozone exposure, plants attempt to
maintain a constant redox potential in their cells. In
many cases, this result in an increase in the
concentrations of antioxidants enzymes and
compounds that play a major role in the defense
system of the plants against oxidative stress.
Ascorbic acid levels in spinach (Luwe et al., 1993),
anthocyanins in blackberries (Barth et al., 1995)
and concentrations of phytoalexins in grapes (Sarig
et al., 1996) were reported to increase after ozone
exposure. More research is needed to identify and
define the physiological responses of each
commodity to ozone.
Microbial reduction and sanitation by
ozone
Ozone destroys microorganisms by the
progressive oxidation of vital cellular ozonation.
Numerous studies were conducted to investigate
the effectiveness of ozone against various kinds of
microorganisms. Studies on microbial inactivation
by ozone in fruits and vegetables are summarized
in Table 1. Besides microbial inactivation, ozonation
has also been applied for the purpose of mycotoxin
degradation in food products.
Ozone in fluid food processing
Microbial studies to date typically show
mandatory 5-log reductions of spoilage and
potentially pathogenic species most commonly
associated with fruit and vegetable juices achieved
using ozone. Applying ozone at doses leading to
effective decontamination may impact the sensory
qualities of food. Ozone is not universally beneficial
and in some cases may promote oxidative spoilage
in foods. Oxidation of undesirable or unwanted
organic and inorganic compounds (iron,
manganese, nitrite, cyanide, hydrogen sulphide) by
application of ozone is rapid (Rakness, 2005). Dock,
1995 reported no detrimental change in quality
attributes of apple cider when it was treated with
ozone. Segovia Bravo et al. (2007) concluded that
ozone treatment (7g /h) for 24 h caused rapid
destruction of most of the polyphenols present in
green table olive solutions and ozone bubbling for
a further 72 h was necessary to reduce the
remaining tyrosol content in the solution.
Ozone in meat processing
Steffen and Rice, 2010 demonstrated the use
of ozone (in gas and aqueous phases) for the
preparation of complete meals in a central food
processing plant; the meals were then packaged
and sealed in a sterilised manner. Ozone is reported
to be effective against contamination of natural
casings, which are generally contaminated with
several microorganisms. Benli et al., 2008
suggested that a combination of treatments, such
as washing with ozonated water to whiten casings
with another treatment, such as irradiation, might
prove effective for enhancing the casing value while
ensuring the destruction of potential food borne
pathogens.
Reagan et al., 1996 evaluated trimming
techniques and beef carcass washing techniques
to improve the microbial quality of meat.
Intervention treatments included knife trimming,
washing with water and rinsing with ozone (0.3–
2.3 ppm) or hydrogen peroxide (5%). Ozone
treatment reduced carcass surface contamination
by 1.30 CFU/cm2, whereas hydrogen peroxide
reduced aerobic plate counts by 1.14 CFU/cm2.
Greer and Jones (1989) evaluated the impact of
gaseous ozone treatment on beef carcass bacterial
spoilage profiles and on meat quality and carcass
shrinkage. They found that psychrotrophic
bacterial growth was retarded on carcass surfaces
while under ozone atmosphere.
Ozone has also been used as a pre-treatment
before cooking to determine any synergistic activity
on reducing microorganisms. Novak and Yuan
(2004a) treated beef surfaces with ozone then
cooked the treated beef at temperatures of 45–75
°C to determine the impact on enterotoxin-
producing strains of Clostridium perfringens. The
authors reported a 1–2 log CFU/g reduction in C.
perfringens as a result of aqueous ozone treatment
and heating at 45–75 °C. Additionally, they reported
a reduction in spore count with the same treatments,
but the magnitude of reduction was very small,
indicating that the spores were much more resistant
PRABHA et al., Ozone Technology in Food Processing: A Review 4043
Table 1. Current researches published on microbial inactivation by ozone
Product Treatment Findings Reference
Orange and
Lemon Gas ozone (0.3ppm) for 4 weeks Spore was reduced of P.
Italicum Paluo et al., 2001
Peach Ozone atmosphere storage (0.3ppm,
5°C) for 4 weeks Aerial mycelial growth and
sporulation were inhibited Paluo et al., 2002
Strawberry Storage for 3 days at 2°C with 1.5ppm
ozone Mycelial growth developed
more slowly Nadas et al., 2003
Apple Ozone bubbling and dipping in pre-
ozonated water, 3min Decrease in counts of E coli (<1
log CFU) Achen et al., 2001
Citrus Basket immersed in ozonated water at
1.5-10ppm, stored for 7-21 days at 10°
C and 20° C
5 ppm for 5 min reduced aerobic
bacteria population Smilanick et al., 2002
Celery Dipping to (0.03,0.08,0.18 ppm)
ozonated water for 5 min and storing at
4°C for 9 days
Populations of total bacteria
reached to 5.72, 5.64 and 5.63
from 5.08 log CFU/g
Zhnag et al., 2005
Lettuce Soaking in ozonated water with 5 ppm
ozone Reduced aerobic bacteria and
yeast within 10 min Koseki et al., 2001
Orange juice Ozone gas pumped into juice E. coli was reduced to 5 log
cylce, Ascorbic acid decreased Angelino et al., 2003,
Williams et al., 2004
Cheese Gaseous ozone at 3 different levels Reduction of fungal counts on
ripening room and on cheese
surface
Serra et al., 2003;
Pinto et al., 2007
Orange and
lemon Ozone gas (0.1-2g/m3) at 4.5-10° C Decrease of P. digitatumand its
sporulation Palou et al., 2001
Barley Gaseous ozone Reduction mycelia grow and
spores germinates Allen et al., 2003
Maize Gaseous ozone Reduction of 63% fungal
growth counts after 3 days Kells et al., 2001
Dried fig 0.01-0.02g/m3 gaseous ozone for 3-4
hours Reduction of fungal counts
(mycoflora) Oztekin et al., 2006
Onion Gaseous ozone Reduction on spore germination,
change of colony colour
(Aspergillus)
Vijayanandraj et al.,
2006
Pea seed 3.85 g/m3 of gaseous ozone for 7.5,15 &
30 minutes Reduction of fungal counts
(Fusarium, Alternatia) Ciccarese et al., 2007
Wheat 1.4 g/m3 gaseous ozone Reduction of fungal counts
(Micromycetes) Raila et al., 2006
Tomato Washing with ozonated water (3.8ppm)
for 10 min Spores of B. cinereaon the
surface were inactivated Ogawa et al., 1990
Blackberry Storage for 12 days at 2° C in an ozone
atmosphere Suppressed fungal development
for 12 days Barth et al., 1995
Date fruit 0.002, 0.006, and 0.01 g/m3 gaseous
ozone for 1 hour Reduction of fungal counts
(total mycoflora) Najafi and
Khodaparast, 2009
to ozone and thermal treatments. The authors
concluded that ozone treatment followed by heat
treatment allowed reductions at cooking
temperatures that normally would not impart the
reduction by themselves.
Ozone in grain processing
The control of pest (insects and
microorganisms including mould, fungi and
bacteria) development in stored grains after harvest
is essential as it currently leads to a grain yield loss
around 3–10% in developed countries, and can
reach 50% in certain countries (Jayas, 1999;
Fleurat-Lessard, 2004; Magan and Aldred, 2007).
Storage grains are very susceptible to a
number of insects, such as Tribolium, Sithophilus
and moths, which cause considerable damage and
4044 Trends in Biosciences 8 (16), 2015
which could potentially develop resistance to the
currently used insecticides. Ozone that can be used
in fumigation is an interesting alternative to applied
chemicals for the control of insect development.
Kells et al. (2001) evaluated the efficiency of ozone
fumigation in a corn grain mass against adult insects,
such as the red flour beetle (Triboliumcastaneum),
maize weevil (Sithophiluszeamais) and larvae from
the Indian meal moth (Plodiainterpunctella). Insects
were put in cages containing maize kernels and
placed into a column filled with grains and
positioned just below the surface. The columns
were treated with or without ozone (50 ppm for 3
days or 25 ppm for 5 days) and the number of
dead insects was determined. Results demonstrated
a significant insect mortality increase (92–100%
compared to 3–10% in the control) when the insect
species in grain samples were treated with 50 ppm
for 3 days. The lower dose was also significantly
efficient but led to a lower insect mortality (77–
99.9%depending on the insect species).Similar
results were obtained by Maier et al., 2006 using
identical conditions but with insects positioned
deeper in corn grain samples(0.6 m below the grain
surface) and in the plenum of silos.
Due to its inactivating action on fungi, ozone
can also be considered as helping to reduce
mycotoxin accumulation during grain storage.
Furthermore, its oxidant properties could also be
used for mycotoxin degradation and detoxification.
Degradation of aflatoxin (McKenzie et al., 1997),
as well as trichothecenes (Young et al., 2006), is
initiated by the attack of a double bond with addition
of two oxygen atoms, which further leads to the
molecule breaking apart. Young et al., 2006
furthermore pointed out that trichothecene
degradation depends on ozone concentration as well
as on pH.
Effect of Ozone on Product Quality and
Nutrition
Microbial studies to date typically show that
the mandatory 5 log reductions of spoilage and
potentially pathogenic species most commonly
associated with fruit and vegetable or juices may
be achieved. A number of studies report the effects
of ozone on quality parameters of treated fruits and
vegetables (Zhang et al., 2005; Fonseca and
Rushing, 2006). The effects of ozone treatment
on quality and physiology of various foods are
reported. Applying ozone at doses that are large
enough for effective decontamination may change
the sensory qualities of food. Ozone is not
universally beneficial and in some cases may
promote oxidative spoilage in foods. Surface
oxidation, discolouration or development of
undesirable odours may occur in substrates from
excessive use of ozone (Khadre et al., 2001). Dock,
1999 reported no detrimental change in the quality
attributes of apple cider when it was treated with
ozone.
Chemical attributes
No change in onion chemical composition and
sensory quality was reported by Song et al., 2000.
Ozone-containing water treatment resulted in no
significant difference in total sugar content of celery
and strawberries (Zhang et al., 2005) during
storage. Ozonation is expected to lead to the loss
of antioxidant constituents, because of its strong
oxidising activity. However, ozone washing
treatment was reported to have no effect on the
final phenolic content of fresh-cut iceberg lettuce
(Beltrán et al., 2005a).Contradictory reports were
found in the literature regarding ascorbic acid.
Decomposition of ascorbic acid in broccoli florets
was reported after ozone treatment by Lewis et
al., 1996, but Zhang et al., 2005 reported no
significant difference in ascorbic acid content
between treated and non-treated at other
concentration.
Visual quality
Gabler et al., 2010 investigated the efficacy
of ozone in controlling post-harvest decay of table
grapes and for the potential replacement of sulfur
dioxide, which is used as a commercial fumigant.
They observed that ozone fumigation with up to
10 000 ìl/L for up to 2 hours helps to control
postharvest grey mould of table grapes caused by
Botrytis cinerea. However, grapes stored in ozone-
rich atmospheres may develop thin longitudinal
darkened lesions. This injury is reported to be
irregular and was not always associated with an
ozone dose or cultivar (Gabler et al., 2010).
Martínez-Sánc et al., 2006 investigated the effect
of several sanitisers on the visual quality and colour
of rocket leaves during storage in air and low O2
(1–3 kPa) + high CO2 (11–13 kPa) for 15 days at 4
°C. They observed that ozone effects were
comparable with other sanitisers, except for lactic
acid treated samples.
Texture
Texture or firmness is an important
rheological property pertinent to fresh fruits and
PRABHA et al., Ozone Technology in Food Processing: A Review 4045
vegetables. Fruits and vegetables with a firm,
crunchy texture are highly desirable because
consumers associate these textural attributes with
freshness and wholesomeness. The appearance of
a soft or limp product may give rise to consumer
rejection prior to consumption (Rico et al. 2007).
Textural changes in fruits and vegetables could be
due to various enzymatic and non-enzymatic
processes. Ozone treatment of fresh fruit and
vegetables either by washing or in storage
consisting of ozone gas is reported to have
significant effects on texture. Firmness of fresh
coriander leaves was reported to decrease through
washing with ozone-containing water compared to
control. Another study conducted by Selma et al.,
2008 reported no significant changes in firmness
of fresh-cut cantaloupe irrespective of gaseous
ozone concentration (5000 or 2000 ppm) for 30
minutes during storage compared to control.
Change in texture during ozonation and subsequent
storage may possibly be due to postharvest changes
in cellulose and hemicellulose contents due to ozone
application during MAP. This could be due to
polymerisation and epimerisation of cellulose and
hemicelluloses contents of cell walls inducing
thickening of the cell walls, causing textural
changes in fresh-cut green asparagus during storage
after ozone treatment (An et al., 2007). An et al.,
2007 reported an increase in cellulose,
hemicelluloses and lignin content during MAP
storage after pre-treatment with aqueous ozone.
Ozonation of fruits has been reported to
enhance firmness of citrus fruits and cucumbers
compared to controls (Skog and Chu, 2001). Ozone
is reported to delay softening in strawberries during
cold-room storage and storage at room temperature
(Nadas et al., 2003). Ozone is a strong oxidising
agent which can cause oxidation of feruloylated
cross linkages or phenolic cross linkages among
cell-wall pectin, structural proteins or other
polymers, and thereby change the firmness of the
product (Heun Hong and Gross, 1998).
Sensory quality
The most notable effect of ozone on the
sensory quality of fruits reported in the literature is
the loss of aroma. Ozone-enriched cold storage of
strawberries resulted in reversible losses of fruit
aroma (Nadas et al., 2003; Perez et al., 1999).
This behaviour is probably due to the oxidation of
volatile compounds. However, Tzortzakis et al.,
2007 did not observe any significant changes in
tomato fruit weight, antioxidant status, CO2/
H2Oexchange, ethylene production or in organic
acid, vitamin C (pulp and seed) or total phenolic
content when exposed to ozone concentrations
ranging between 0.005 and 1.0 ìmol/mol at 13 °C
and 95% RH. Similar results were reported by Kute
et al., 1995 for strawberry exposed to ozone
concentrations between 0.3 and 0.7 ìmol/mol for
up to 1 week. Applying ozone at doses that are
large enough for effective decontamination may
change the sensory qualities of these products.
Future Trends
Ozone application has given promising results
for important problems of food industry, such as
mycotoxin and pesticide residues. Degradation
products, formed after ozonation of these residues,
have not exactly been determined, and this seems
to be the most crucial obstacle on this subject. In
vivo and in vitro toxicological tests are needed to
be conducted to screen the effects of degradation
products on human and animal health. Through
emerging new techniques, as well as improvements
and innovations in ozone generating and application
systems, the subject will be evaluated more
effectively in future.
S. Patil and P. Bourke said that the possibility
that interaction of ozone with the types of organic
material present in the food product results in
differing radical production trends that can lead to
microbial inactivation should be studied. Therefore,
studying the mechanism of the reaction of ozone
with organic materials will contribute to establishing
the impact of specific radical species on target
microorganisms. Further research is required to
ascertain the interaction of food constituents with
ozone and role of resulting compounds in the
inactivation process. Optimization of process
parameters may also follow more specific process-
related studies on mechanisms of action.
Ozone is an effective sanitizer with great
potential applications in the food industry. It
decomposes into simple oxygen with no safety
concerns about consumption of residual by-
product. Due to its high oxidation capacity and
microbial inactivation potential, ozone has prevented
various kinds of microbial spoilages usually
encountered in fruits and vegetables.
Decontamination of products by ozone depends on
number and kind of contaminating microorganisms,
physiology of the product, ozone application
system, temperature, pH, and other factors. If
4046 Trends in Biosciences 8 (16), 2015
improperly used, ozone can cause some deleterious
effects on physiology and quality of products such
as losses in sensory quality. For effective and safe
use in food processing, optimum ozone
concentration, contact time and other treatment
conditions should be defined for all products. Pilot
trials must be conducted before starting commercial
application, because every ozone application is
unique.
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Received on 25-07-2015 Accepted on 31-07-2015
