Investigation of Antimicrobial Activity of Grapefruit Seed Extract and

Aerosol and Air Quality Research, 15: 1035–1044, 2015
Copyright © Taiwan Association for Aerosol Research
ISSN: 1680-8584 print / 2071-1409 online
doi: 10.4209/aaqr.2014.09.0208
Investigation of Antimicrobial Activity of Grapefruit Seed Extract and Its
Application to Air Filters with Comparison to Propolis and Shiitake
Bangwoo Han1,2*, Ji-Soo Kang1,2, Hak-Joon Kim1, Chang-Gyu Woo1, Yong-Jin Kim1
1
Environmental & Energy Systems Research Division, Korea Institute of Machinery and Materials, 156, Gajeongbuk-ro,
Yuseong-gu, Daejeon, 305-343, Korea
2
Environment & Energy Mechanical Engineering, University of Science & Technology, 217 Gajeong-ro, Yuseong-gu,
Daejeon, 305-333, Korea
ABSTRACT
Antimicrobial air filters using a new natural product agent, grapefruit seed extract (GSE) have been investigated for
application in air purifiers or heating, ventilation, and air-conditioning (HVAC) systems. The disk diffusion method was
used to evaluate the antimicrobial activity of GSE, propolis, and shiitake against four bacteria: Staphylococcus aureus,
Micrococcus luteus, Psedomonas aeruginosa, and Escherichia coli. The inactivation of S. aureus was then investigated on
air filters treated with two natural products, GSE and propolis (selected for comparison to GSE) using two test methods
based on different deposition weights.
GSE displayed the highest antimicrobial activity against S. aureus, P. aeruginosa, and E. coli of the three natural
products tested by the disk diffusion method. Similar inactivation rates of 58–67% for GSE and propolis air filters were
observed at a relatively low deposition weight of 94–98 μg/cm2 using the aerosol deposition method. However, the
inactivation rate was considerably superior on the GSE air filter (~98%) compared to the propolis air filter (~75%) at
deposition weights of 5000–8000 μg/cm2 using the film attachment method. The inactivation rate for both GSE and
propolis air filters could be expressed as an exponential function [in the form of 1 – exp(–axb)] of the deposition weight
per unit area of the natural product. The superior inactivation performance of the GSE air filter using the film attachment
method was probably due to the high wettability of GSE for bacteria cultures, with a contact angle less than 20°.
Therefore, GSE air filters will be more effective and economical than propolis air filters due to their high microbial
activity and relatively low price.
Keywords: Natural product; Air filter; Bioaerosol; Antimicrobial; Inactivation.
INTRODUCTION
Bioaerosols are defined as airborne particles of biological
origin consisting of bacteria, fungi, fungal spores, viruses,
yeast, and pollen and their fragments, which include various
antigens (Kalogerakis et al., 2005). Exposure to these
bioaerosols in indoor environments has become a serious
concern due to the wide range of adverse health effects,
including contagious infectious diseases (Dales et al.,
1991), hypersensitivity pneumonitis (Siersted and Gravesen,
1993), and respiratory problems such as asthma and
rhinitis (Beaumont, 1988). Due to their presence in nature,
exposure to bioaerosols is almost inevitable in most enclosed
environments (Jones and Harrison, 2004; Ren et al., 1999).
*
Corresponding author.
Tel.: +82-42-868-7068; Fax: +82-42-868-7284
E-mail address: [email protected]
Various techniques have been used to mitigate the
viability of biological contaminants in indoor environments,
such as ultraviolet (UV) irradiation (Lin and Li, 2002), air
electric ion emission (Huang et al., 2008), ozone (Huang et
al., 2012), and photocatalytic oxidation (Chen et al., 2010),
among others. One promising method is an antimicrobial
filtration system that physically captures bioaerosols on air
filters and then inactivates the airborne microorganisms on
the surface of the filters by treatment with antimicrobial
materials such as silver or titanium-based nanoparticles (Li
et al., 2006; Jin et al., 2007,), iodine powders (Lee et al.,
2008), and carbon nanotubes (Brady-Estévez et al., 2008;
Jung et al., 2011a).
Natural products are promising antimicrobial agents for
indoor air quality improvement because they are typically
less toxic compared to other chemicals or nanomaterials
(Carson et al., 2006; Pibiri et al., 2006; Hwang et al.,
2012). For example, Arabidopsis, rice, corn, soy bean, and
the model legume Medicago truncatula are rich sources of
antimicrobial indole, terpenoid, benzoxazinone, and
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Han et al., Aerosol and Air Quality Research, 15: 1035–1044, 2015
flavonoid/isoflavonoid natural products (Dixon, 2001).
However, only a few trials have been conducted to apply
these natural products as antimicrobial agents in air
filtration systems.
Huang et al. (2010) investigated Melaleuca alternifolia
(tea tree oil) as a disinfecting media for the inactivation of
common environmental fungal spores on a filter surface
and found that 50% of Aspergillus niger and 40% of
Rhizopus stolonifer spores could be inactivated over a
period of 60 min. Jung et al. (2011b) investigated a natural
plant extract from Sophora flavescens for application to
antimicrobial air filters and reported that the prepared
natural product on an antimicrobial filter was effective for
inactivating Gram-positive bacteria such as Staphylococcus
epidermidis and Bacillus subtilis. Lee et al. (2013)
investigated the antimicrobial effects of a natural extract of
Mukdenia rossii and unipolar ion emission on air filters for
use against S. epidermidis and noted that the inactivation
rate was about 70% using natural extract-treated filters and
was enhanced by a further 20% due to unipolar ion emission.
Shiitake, one of the most popular edible mushrooms in
the world, is claimed to have antitumor, antiviral, and
antimicrobial properties, as well as hypocholesterolemic
and hypoglycaemic actions (Hearst et al., 2009). Propolis,
a hard and resinous natural product derived by bees from
plant juices, exhibits antibacterial activity due to its high
flavonoid content (Benhanifia et al., 2013; Burdock, 1998;
Kujumgiev et al., 1999). Grapefruit seed extract (GSE), a
commercial product derived from the seeds and pulp of
grapefruit, also has powerful antimicrobial properties (Lim
et al., 2010) mainly due to the polyphenolic compounds it
contains (Saito et al., 1998; Aloui et al., 2014). However,
to our knowledge, no trial using antimicrobial air filters
prepared with shiitake, propolis, and GSE natural products
has been previously reported.
In this study, for the first time, antimicrobial air filters
using new natural product agents, grapefruit seed extract
(GSE) and propolis have been investigated for application
in air purifiers or HAVC systems. The antimicrobial activities
of three natural products, shiitake, propolis, and GSE were
compared using a disk diffusion method for four test
bacteria. The application to antimicrobial air filters treated
with natural products was then performed using two natural
product agents: GSE and propolis. GSE was selected because
it had the highest antimicrobial activity among the three
natural products and propolis was selected for comparison
to GSE. The inactivation of Staphylococcus aureus on the
antimicrobial air filters treated with natural products was
investigated at a variety of deposition weights using two
test methods with different orders of deposition weight.
EXPERIMENTAL METHODS
Materials
Grapefruit seed extract (GSE, DF-100) prepared by
grinding the grapefruit seed and juiceless pulp, then mixing
with glycerine, was purchased from the Food Additives
Bank Co., Ltd. (Anseong, Korea). Naringin, one of the major
antimicrobial contents of GSE, was 0.51%(w/w) in the as-
purchased sample. Propolis was acquired from Seoul Propolis
Co., Ltd. (Seoul, Korea). CAPE (caffeic acid phenethyl ester)
is known to be one of the main components of propolis. 18
ppm of CAPE was measured in the as-obtained propolis
sample according to the manufacturer. Shiitake mushroom
(L. edodes) extract was obtained from YD Global Life
Science (Seoul, Korea). The mushroom extract was obtained
from dried mushroom caps through ethanol extraction. The
extraction process started with 100.12 g of dried mushroom
caps in the market (Mungyeong, Korea) and the extract
was 9.09 g. Lentinan is one of the most active 1,3/1,6-β-Dglucans and its amount in the dried Japanese L. edodes was
reported as 10~15 mg/g (Minato et al., 1999).
Four bacteria, S. aureus, Micrococcus luteus, Psedomonas
aeruginosa, and Escherichia coli were prepared for
antimicrobial activity tests of natural products using the disk
diffusion method. S. aureus, which causes a diverse array
of life-threatening infections and is capable of adapting to
different environmental conditions (Lowy, 2003), and M.
luteus, which is a nonmotile, spherical, saprotrophic
bacterium that colonizes the human mouth, mucosae,
oropharynx, and upper respiratory tract (Mauclaire and
Egli, 2010), were selected as Gram-positive bacteria. P.
aeruginosa, which is a typical opportunistic pathogen that
colonizes the lungs of cystic fibrosis patients and causes
severe infections in immunocompromised hosts (Köller et al.,
2000), and E. coli, which is the most commonly isolated
microorganism from human primary urinary tract infections
(Hull et al., 1981), were selected as Gram-negative bacteria.
E. coli (ATCC 11775) and P. aeruginosa (ATCC 10145)
were cultured in nutrient broth at 37°C and M. luteus
(ATCC 4698) was cultured in nutrient agar at 30°C for 24
h. They were provided by the Korean Research Institute of
Bioscience and Biotechnology.
S. aureus was selected as a test bacterium for evaluation
of the natural product antimicrobial filters because of its
high persistence despite the use of effective antimicrobials
(Mylotte et al., 1987). S. aureus KCTC 1621 (=ATCC 25923)
was purchased from the Korean Collection for Type Cultures
(KCTC, Korea). S. aureus was cultured in nutrient broth
(DifcoTM Nutrient Broth 234000) or nutrient agar (DifcoTM
Nutrient Agar 213000) at 37°C in an incubator. For
nebulization, the concentration of bacterial suspension was
set to about 1 × 108 CFU/mL by optical density. The sterile
water was prepared by autoclaving 20 mL of triple distilled
water for 15 min at 120°C. For the film attachment method, a
1000-fold dilution was prepared to make the concentration
1 × 105 CFU/mL.
Antimicrobial Tests
Disk Diffusion Method
The disk diffusion method was conducted according to
the standard method reported by Bauer et al. (1966) and was
used to compare the antibacterial activities of three natural
product extracts. Natural product extracts were diluted with
an ethanol–water solution containing 95% ethanol (Samchun
Pure Chemical Ind. Co., Ltd., Pyeoungtaek, Korea) with a
concentration of 25% (w/v). The four bacteria were swabbed
uniformly across petri plates, which were autoclaved at
Han et al., Aerosol and Air Quality Research, 15: 1035–1044, 2015
120°C for 15 min. Three filter paper disks impregnated with
three natural product extracts and one filter paper disk filled
with a 25% ethanol aqueous solution as a control paper
were placed on the surface of the agar. The plates were
incubated at 5°C for 1 h to permit good diffusion and then
transferred to an incubator at 37°C for 24 h. Antimicrobial
activity was recorded by measuring the width of the clear
inhibition zone around the disk using calipers.
Aerosol Deposition Method on Antimicrobial Filters
The aerosol deposition method on antimicrobial filters
has been performed in many other studies (Eninger et al.,
2009; Jung et al., 2011b; Miaśkiewicz-Peska and Łebkowska,
2011). Two natural product extracts, GSE and propolis,
were diluted with 95% ethanol (Samchun Pure Chemical
Ind. Co., Ltd.) with a concentration of 0.6% (w/v). The
diluted natural product extracts were sprayed with a singlejet collision nebulizer (BGI Inc., Waltham, MA, USA)
supplied with an airflow of 1 L/min and then passed through a
diffusion dryer containing activated carbon to remove
ethanol from the sprayed droplets. The aerosol particle
sizes for the GSE and propolis produced via nebulization
were similar, with a mode diameter of 0.63–0.68 μm when
measured with an aerodynamic particle sizer (APS, model
3321; TSI, Shoreview, MN, USA). The dried natural
product particles were deposited on test filters composed
of polyethylene terephthalate (PET) with a diameter of 13
mm (fiber diameter of 1.5 μm, thickness of 0.45 mm) for
60–120 s. S. aureus droplets were generated by a single-jet
collision nebulizer and passed through a diffusion dryer
containing silica gel to remove water vapor. The dried
bioaerosols were then deposited on the natural product
antimicrobial filters for 5 min. The PET filter had a singlepass particle collection efficiency of 98–99% at a face
velocity of 0.2 m/s for the S. aureus bioaerosols. The filters
deposited by the bioaerosols were extracted into a 5 mL
buffer fluid consisting of phosphate-buffered saline (PBS)
with 0.05% (v/v) Tween 80. The extraction fluid was
vortexed for 2 min, agitated in an ultrasonic bath for 10
min, and then vortexed once more for 2 min. The extraction
fluid was diluted with PBS at ratios of 1, 10, and 100 times,
placed on a nutrient agar and then incubated for 24 h at
37°C. The numbers of CFU formed on the nutrient agar
were counted after incubation. The inactivation rate was
defined as
Inactivation rate (%) = (1 – CFU/CFUcontrol) × 100
(1)
where CFUcontrol is the number of colonies recovered from
the control filter on which no antimicrobial treatment using
natural products was performed.
Film Attachment Method on Antimicrobial Filters
The antimicrobial properties of the filters were also
determined by the film attachment method (Kawakami et
al., 2008; Shin et al., 2011), which is a Japanese industrial
standard test (JIS Z 2801:2000). Two natural products,
GSE and propolis, were diluted with 95% ethanol with a
concentration of 0.6% (w/v). Air filters with a size of 40 ×
1037
40 mm2 were uniformly coated with the natural products
with a spray gun at an airflow of 5 L/min. We used two
kinds of air filters (#56 and #90), which are used as pre
filters (non-woven PET, MERV 10 class, fiber diameter of
30 μm, thickness of 0.26 mm) and medium filters (meltblown PP, H13 class, fiber diameter of 30 μm, thickness of
0.55 mm; 3AC ltd Co. Seoul, Korea) in commercial indoor
air purifiers, respectively. They had a single-pass particle
collection efficiency of 56% and 90%, respectively at a
face velocity of 0.2 m/s for the S. aureus bioaerosols. The
pre filter (#56), which is probably little suitable for bioaerosol
filtration owing to its low collection efficiency, was selected
to compare the antimicrobial performances of the filters
which have different particle collection efficiencies.
The test samples were placed in petri dishes and inoculated
with 0.4 mL of a bacterial culture containing 1×105
CFU/mL. The inoculum was covered with a polyester film
(X-131 transparent copier film; Folex Imaging, Solihull,
West Midlands, UK), and the petri dishes were incubated
at 37°C for 24 h in a humid chamber to prevent desiccation.
After the incubation period, 20 mL of extraction solution
[0.1% (v/v) Tween 20, 145 mM sodium chloride, 20.5 mM
sodium phosphate; pH 7.4] was added to the petri dishes,
which were then shaken for 2 min. Subsequently, serial
dilutions of the extraction solution were spread on agar
plates in triplicate and incubated at 37°C overnight. Colonies
were counted visually, and the numbers of CFU per sample
were determined. The activity value was calculated using
Eq. (1) to compare the results between the two different
methods rather than using the original JIS standard tool, in
which it is obtained by subtracting the log value determined
for the test sample from the log value determined for the
control.
RESULTS
Fig. 1 shows the size of the clear zones produced by the
three natural products against the four bacteria. The size of
the clear zone on control filters was approximately10 mm.
GSE had the highest antimicrobial activity for both Grampositive and Gram-negative bacteria, except M. luteus. Of
the three natural products, GSE produced a clear zone with
the largest size for S. aureus, P. aeruginosa, and E. coli,
and had the highest antimicrobial activity against S. aureus.
Shiitake and propolis displayed little antimicrobial activity
against Gram-negative bacteria, whereas they displayed a
similar activity to GSE against Gram-positive bacteria.
Fig. 2 shows the change in the size of the clear zone for
the action of GSE against three bacteria at different GSE
concentrations [from 50 ppm to 250,000 ppm (25%)]. The
size of the clear zone increased proportionally to the GSE
concentration, and was larger in the order of P. aeruginosa
< E. coli < S. aureus at GSE concentrations larger than
5000 ppm (0.5%). Antimicrobial activity against S. aureus
increased continuously with GSE concentration, whereas
activity against E coli and P. aeruginosa was almost saturated
at a GSE concentration of 5000 ppm.
Fig. 3 shows the size of the clear zones observed in this
study compared to previous studies. The sizes of the clear
Han et al., Aerosol and Air Quality Research, 15: 1035–1044, 2015
1038
Bacterial strains
S.aureus(+)
Control
M.luteus(+)
Shiitake mushroom
Propolis
P. aeruginosa(-)
GSE
E.coli(-)
0
6
12
18
24
30
36
42
Clear zone size (mm)
Fig. 1. Comparison of the size of clear zones of the four bacteria for the control paper and three natural products.
35
Clear zone size (mm)
E.coli(-)
28
P. aeruginosa(-)
S.aureus(+)
21
14
7
0
50
500
5000
50000
500000
GSE concentration (ppm)
Fig. 2. The size of the clear zones of three bacteria at different concentrations of the GSE natural product.
zones of propolis against E. coli and S. aureus in the study
of Rahman et al. (2010) were compared to those of GSE in
our study and are shown in Fig. 3(a). Similarly to the result
in Fig. 2, the sizes of the clear zones resulting from the
action of GSE were larger than those for propolis at a given
concentration of natural product, even after considering the
different sizes of clear zones on the control filter papers
(10 mm in this study and 7–8 mm in Rahman et al., 2010).
At concentrations larger than 1000 ppm, the antimicrobial
activities of GSE against E-coli (Gram-negative bacteria)
were superior to those of propolis, which is similar to the
result shown in Fig. 1. The sizes of the clear zones produced
by a honey (Baidhyanath honey) against E. coli and S.
aureus in Chute et al. (2010) were also compared to those
of GSE in our study, as shown in Fig. 3(b). The initial size
of the clear zone on control filter papers in Chute et al.
(2010) was also about 10 mm. GSE at a concentration of
25% had a higher antimicrobial activity against S. aureus
compared to the honey at a concentration of 40%, while it
had a similar activity against E. coli to the honey at a
concentration of 40%.
Fig. 4 shows SEM images of filter #56 and filter #90
with GSE and propolis deposited for film attachment tests.
The liquid phases of GSE and propolis were coated on to
the filters and then dried on the surfaces, producing
different shapes compared to other studies in which natural
products were deposited on the filters in the form of already
dried isolated particles (Jung et al., 2011b; Lee et al., 2013).
As seen from Fig. 4, no significant morphological difference
existed between the filters coated by GSE and propolis.
Figs. 5(a) and 5(b) show the inactivation rates of air
filters containing GSE and propolis deposits against S.
aureus for different colony numbers in the control filters
obtained by the aerosol deposition and film attachment
methods, respectively. In the aerosol deposition method,
the inactivation rates of air filters were similar for both
GSE and propolis at deposition weights of 94–98 μg/cm2,
and they decreased as the colony number increased on the
control filter. However, in the film attachment method, the
inactivation rate of the propolis air filters decreased from
Han et al., Aerosol and Air Quality Research, 15: 1035–1044, 2015
1039
30
GSE_E.coli (This work)
Clear zone size (mm)
GSE_S.aureus (This work)
Propolis_E.coli (Rahman et al.(2010))
20
Propolis_S.aureus (Rahman et al.(2010))
10
0
10
100
1000
10000
Natural product concentration (ppm)
(a)
35
E.coli
Clear zone size (mm)
28
S.aureus
21
14
7
0
Honey 20%
Honey 40%
(Chute et al.(2010)) (Chute et al.(2010))
GSE 25%
(This work)
Natural product
(b)
Fig. 3. Comparison of the size of clear zones of E. coli and S. aureus for the GSE natural product in this study to those of
previous studies: (a) Rahman et al. (2010) and (b) Chute et al. (2010).
78 to 67–69% as the colony number increased on the
control filter from 5000 to 20,000–25,000 at a high deposition
weight of 5000 μg/cm2, whereas the inactivation rate of the
GSE air filters was maintained at more than 95% at a high
deposition weight of 8000 μg/cm2, even when the colony
number on the control filter was increased from 5000 to
20,000–25,000. The deposition quantities of GSE and
propolis in this study were as similar or quite low for use in
air filters as compared to previous researches (75–300 μg/cm2
in Jung et al. (2011b), 0.9 g/cm2 in Huang et al. (2010)).
Little difference was observed in the inactivation rates
between filter #56 and filter #90 coated with GSE and
propolis, respectively.
Fig. 6 shows the inactivation rate of air filters containing
GSE and propolis deposits against S. aureus at different
deposition weights per unit area for both the aerosol
deposition and film attachment methods. Here, PET filters
of 13 mm in diameter and melt-blown PP filters (#90) with
a 40 × 40 mm2 were used in the aerosol deposition method
and film attachment method, respectively. The colony
number in the control filters was fixed to be about 1 × 104.
The inactivation rate of the GSE and propolis air filters
increased with the deposition weight per unit area of natural
products and represented a typical exponential function of
1 – exp(–axb). Similar to the result based on the disk diffusion
method, the inactivation rate of GSE air filters was higher
than for the propolis air filters at the higher deposition
weights per unit area.
1040
Han et al., Aerosol and Air Quality Research, 15: 1035–1044, 2015
Fig. 4. Scanning electron micrographs of (a) #56 and (b) #90 air filters on which GSE was deposited, and (c) #56 and (d)
#90 air filters on which propolis was deposited.
Fig. 7 shows the contact angles of a bacterial culture
droplet on the surface of the control, propolis, and GSE air
filters. The contact angles of the droplet on the control
filter and propolis-deposited filter were 122° and 93°,
respectively. However, the angle was smaller than 20° on
the GSE air filters, on which the bacterial culture quickly
soaked in as it contacted the filter without a film for
attachment. This indicates that a bacteria culture can be
easily contacted to GSE natural product, which probably leads
to an enhancement of the antimicrobial activity of the GSE air
filter because the wettability of the antimicrobial filter
surfaces is known to be highly related to its antimicrobial
activity in previous literatures (Wang et al., 2008; Necula
et al., 2009; Compagnoni et al., 2012; Agrawal et al.,
2013). Therefore, the high wettability of the bacteria culture,
as well as the inherent superior antimicrobial activity of
GSE, ensured that the GSE natural product was an excellent
agent for application to antimicrobial air filters. Furthermore,
GSE is preferable for air filters because the cost is one or
two orders of magnitude lower (several US dollars per 100
mL) than propolis, and it can be deposited on air filters
with higher deposition weights at a set price.
DISCUSSIONS
To investigate the possibility of the application of the
developed natural products on air filters, pressure drop and
particle collection efficiency of the natural product-treated
air filters were compared to the control filters. Pressure
drops across the PET control filter used in the aerosol
deposition method were 56.1 ± 1.7 Pa at a face velocity of
0.2 m/s. Whereas, pressure drops across the prepared GSE
and propolis air filters with a deposition weight of 94–98
μg/cm2 were 56.1 ± 2.0 Pa and 60.7 ± 3.4 Pa, respectively.
Therefore, the increase of the pressure drop was negligible
for the GSE air filters and only 8.2% for the propolis air
filters compared to the control filter. Particle collection
efficiency for the S. aureus bioaerosols was 98.6% for the
control filter and 98.2%, 96.5% for the prepared GSE and
propolis air filter, respectively. There was little change in
the particle collection efficiency of the prepared GSE and
propolis air filters compared to the control filter. Furthermore,
the effect of life-time and clogging for the natural product
antimicrobial filter is very important for application to air
filters in HVAC systems or air cleaners. According to the
previous antimicrobial durability study using Sophora
flavescens (Chong et al., 2013), the antimicrobial activity
remained stable for 90 days and however, rapid degradation
of the activity owing to the noticeable decrease of the
quantities of major antimicrobial compounds. It provides
useful information regarding the average life expectation
of antimicrobial air filters using Sophora flavescens but
does not be generally prescribed for all natural products.
For the case of GSE and propolis, it is necessary to investigate
the effect of life time or clogging on the antimicrobial
performances of the natural product filters, which will be
prepared as our next study.
CONCLUSIONS
The antimicrobial activity of three natural products,
grapefruit seed extract (GSE), propolis, and shiitake against
Han et al., Aerosol and Air Quality Research, 15: 1035–1044, 2015
1041
(a)
(b)
Fig. 5. Inactivation rates of S. aureus for the antimicrobial filters using GSE and propolis in different test methods: (a)
aerosol deposition; (b) film attachment.
four bacteria, S. aureus, M. luteus, P. aeruginosa, and E. coli,
were compared using a disk diffusion method. Inactivation
tests against S. aureus for the antimicrobial air filters treated
with two natural products, GSE and propolis, were then
performed at various deposition weights using two methods
(aerosol deposition and film attachment). Of the three
natural products, GSE displayed the highest antimicrobial
activity against three bacteria, S. aureus, P. aeruginosa,
and E. coli, and among the four bacteria, it had the highest
antimicrobial activity against S. aureus. In the aerosol
deposition method, the inactivation rates against S. aureus
for GSE and propolis air filters were similar at the same
deposition weights (< 100 μg/m2). However, in the film
attachment method, the inactivation rate of the GSE air
filter was higher than that of the propolis air filter at the
same deposition weights (> 1000 μg/m2) due to the higher
wettability of GSE air filters for bacteria culture. Therefore,
GSE air filters will be more effective and economical than
propolis air filters because of their higher microbial
activity and relatively lower price.
ACKNOWLEDGMENTS
This research was mainly supported by the Eco-Innovation
Project, which is run under the auspices of the Korea
Ministry of the Environment, and partially supported by a
General Research Fund (NK182B) of the Korea Institute
of Machinery and Materials.
Han et al., Aerosol and Air Quality Research, 15: 1035–1044, 2015
1042
Fig. 6. Redrawing of the inactivation rates for the antimicrobial filters using GSE and propolis according to deposition
weight per unit area in the two test methods.
(a)
(b)
(c)
Fig. 7. Photographs indicating the contact angle of a bacterial culture droplet on a (a) control air filter, (b) propolis air filter,
and (c) GSE air filter.
REFERENCES
Agrawal, N.K., Agarwal, R., Vijay, Y.K. and Swami, K.C.
(2013). Surface Modification of Ag Nano Composites
Polymer Membranes by Glow Discharge Plasma. J.
Mater. Sci. Surf. Eng. 1: 23–27.
Aloui, H., Khwaldia, K., Sánchez-González, L., Muneret,
L., Jeandel, C., Hamdi, M. and Desobry, S. (2014).
Alginate Coatings Containing Grapefruit Essential Oil
or Grapefruit Seed Extract for Grapes Preservation. Int.
J. Food Sci. Technol. 49:952–959.
Bauer, A.W., Kirby, M., Sherris, J.C. and Turck, M.
(1966). Antibiotic Susceptibility Testing by a Standardized
Single Disk Method. Am. J. Clin. Pathol. 45:493–496.
Beaumont, F. (1988). Clinical Manifestations of Pulmonary
Aspergillus Infections. Mycoses 31:15–20.
Benhanifia, M., Mohamed, W.M., Bellik, Y. and Benbarek,
H. (2013). Antimicrobial and Antioxidant Activities of
Different Propolis Samples from North-western Algeria.
Int. J. Food Sci. Technol. 48: 2521–2527.
Brady-Estévez, A.S., Kang, S. and Elimelech, M. (2008).
Single-walled-carbon-nanotube Filter for Removal of
Viral and Bacterial Pathogens. Small 4: 481–484.
Burdock, G.A. (1998). Review of The biological Properties
and Toxicity of Bee Propolis (Propolis). Food Chem.
Toxicol. 36: 347–363.
Carson, C.F., Hammer, K.A. and Riley, T.V. (2006).
Melaleuca Alternifolia (Tea Tree) Oil: A Review of
Antimicrobial and Other Medicinal Properties. Clin.
Microbiol. Rev. 19: 15–62.
Chen, F., Yang, X., Mak, H.K.C. and Chan, D.W.T. (2010).
Photocatalytic Oxidation for Antimicrobial Control in
Built Environment: A Brief Literature Overview. Build.
Environ. 45:1747–1754.
Chute, R.K., Deogade, N.G. and Kawale, M. (2010).
Antimicrobial Activity of Indian Honey against Clinical
Isolates. Asiatic J. Biotech. Res. 1: 35–38.
Compagnoni, M.A., Pero, A.C., Ramos, S.M.M., Marra, J.,
Paleari, A.G. and Rodriguez, L. (2014). Antimicrobial
Activity and Surface Properties of an Acrylic Resin
Containing a Biocide Polymer. Gerodontology 31: 220–
226.
Han et al., Aerosol and Air Quality Research, 15: 1035–1044, 2015
Dales, R.E., Zwanenburg, H., Burnett, R. and Franklin, C.A.
(1991) Respiratory Health Effects of Home Dampness and
Molds among Children. Am. J. Epidemiol. 134:196–203.
Dixon, R.A. (2001). Natural Products and Plant Disease
Resistance. Nature 411:843–847.
Eninger, R.M., Hogan, C.J., Biswas, P., Adhikari, A.,
Reponen, T. and Grinshpun, S.A. (2009). Electrospray
versus Nebulization for Aerosolization and Filter Testing
with Bacteriophage Particles. Aerosol Sci. Technol. 43:
298–304.
Hearst, R., Nelson, D., McCollum, G., Millar, B.C.,
Maeda, Y., Goldsmith, C.E., Rooney, P.J., Loughrey,
A., Rao, J.R. and Moore, J.E. (2009). An Examination
of Antibacterial and Antifungal Properties of Constituents
of Shiitake (Lentinula edodes) and Oyster (Pleurotus
ostreatus) Mushrooms. Complement. Ther. Clin. Pract.
15: 5–7.
Huang, H.L., Lee, M.G. and Tai, J.H. (2012). Controlling
Indoor Bioaerosols Using a Hybrid System of Ozone
and Catalysts. Aerosol Air Qual. Res. 12: 73–82.
Huang, R., Agranovski, I., Ryankov, O. and Grinshpun, S.
(2008). Removal of Viable Bioaerosol Particles with a
Low-efficiency HVAC Filter Enhanced by Continuous
Emission of Unipolar Air Ions. Indoor Air 18: 106–112.
Huang, R., Pyankov, O.V., Yu, B. and Agranovski I.
(2010). Inactivation of Fungal Spores Collected on
Fibrous Filters by Melaleuca alternifolia (Tea Tree Oil).
Aerosol Sci. Technol. 44: 262–268.
Hull, R.A., Gill, R.E., Hsu, P., Minshew, B.H. and Falkow,
S. (1981). Construction and Expression of Recombinant
Plasmids Encoding Type 1 or D-mannose-resistant Pili
from a Urinary Tract Infection Escherichia coli isolate.
Infect. Immun. 33: 933–938.
Hwang, G.B., Lee, J.E., Nho, C.W., Lee, B.U., Lee, S.J.,
Jung, J.H. and Bae, G.N. (2012). Short-term Effect of
Humid Airflow on Antimicrobial Air Filters Using
Sophora Flavescens Nanoparticles. Sci. Total Environ.
421–422: 273–279.
Jin, W.J., Jeon, H.J., Kim, J.H. and Youk, J.H. (2007). A
Study on the Preparation of Poly(vinyl alcohol) Nanofibers
Containing Silver Nanoparticles. Synth. Met. 157: 454–
459.
Jones, A.M. and Harrison, R.M. (2004). The Effects of
Meteorological Factors on Atmospheric Bioaerosol
Concentrations. Sci. Total Environ. 326: 151–180.
Jung, J.H., Hwang, G.B., Lee, J.E. and Bae, G.N. (2011a).
Preparation of Airborne Ag/CNT Hybrid Nanoparticles
Using an Aerosol Process and Their Application to
Antimicrobial Air Filtration. Langmuir 27: 10256–10264.
Jung, J.H., Hwang, G.B., Park, S.Y., Lee, J.E., Nho, C.W.,
Lee, B.U. and Bae G.N. (2011b). Antimicrobial Air
Filtration Using Airborne Sophora flavescens NaturalProduct Nanoparticles. Aerosol Sci. Technol. 45: 1510–
1518.
Kalogerakis, N., Paschali, D., Lekaditis, V., Pantidou, A.,
Eleftheriadis, K. and Lazaridis, M. (2005). Indoor Air
Quality—bioaerosol Measurements in Domestic and
Office Premises. J. Aerosol Sci. 36: 751–761.
Kawakami, H., Yoshida, K., Nishida, Y., Kikuchi, Y. and
1043
Sato, Y. (2008). Antibacterial Properties of Metallic
Elements for Alloying Evaluated with Application of
JIS Z 2801:2000. ISIJ Int. 48: 1299–1304.
Köller, T., Curty, L.K., Barja, F., van Delden, C. and
Pechère, J.C. (2000). Swarming of Pseudomonaus
aeruginosa is Dependent on Cell-to-cell Signaling and
Requires Flagella and Pili. J. Bacteriol. 182: 5990–5996.
Kujumgiev, A., Tsvetkova, I., Serkedjieva, Y., Bankova,
V., Christov, R. and Popov, S. (1999). Antibacterial,
Antifungal and Antiviral Activity of Propolis at Different
Geographic Origin. J. Ethnopharmacol. 64: 235–240.
Lee, D.H., Jung, J.H. and Lee, B.U. (2013). Effect of
Treatment with a Natural Extract of Mukdenia rossii
(Oliv) Koidz and Unipolar ion Emission on the
Antibacterial Performance of Air Filters. Aerosol Air
Qual. Res. 13: 771–776.
Lee, J.H., Wu, C.Y., Wysocki, K.M., Farrah, S. and Wander,
J. (2008). Efficacy of Iodine-treated Biocidal Filter Media
against Bacterial Spore Aerosols. J. Appl. Microbiol. 105:
1318–1326.
Li, Y., Leung, P., Yao, L., Song, Q.W. and Newton, E.
(2006). Antimicrobial Effect of Surgical Masks Coated
with Nanoparticles. J. Hosp. Infect. 62: 58–63.
Lim, G., Jang, S. and Song, K.B. (2010). Physical and
Antimicrobial Properties of Gelidium corneum/nano-clay
Composite Film Containing Grapefruit Seed Extract or
thymol. J. Food Eng. 98: 415–420.
Lin, C.Y and Li, C.S. (2002). Control Effectiveness of
Ultraviolet Germicidal Irradiation on Bioaerosols. Aerosol
Sci. Technol. 36 :474–478.
Lowy, FD. (2003) Antimicrobial Resistance: The Example
of Staphylococcus aureus. J. Clin. Invest. 111: 1265–
1273.
Mauclaire, L. and Egli, M. (2010). Effect of Simulated
Microgravity on Growth and Production of Exopolymeric
Substances of Micrococcus luteus Space and Earth
Isolates. FEMS Immunol. Med. Microbil. 59: 350–356.
Miaśkiewicz-Peska, E. and Łebkowska, M. (2011). Effect
of Antimicrobial Air Filter Treatment on Bacterial
Survival. Fibres Text. East. Eur. 19: 73–77.
Minato, K., Mizuno, M., Terai, H. and Tsuchida, H. (1999).
Autolysis of Lentinan, an Antitumor Polysaccharide,
during Storage of Lentinus edodes, Shiitake Mushroom.
J. Agric. Food Chem. 47: 1530–1532
Mylotte, J.M., McDermott, C. and Spooner, J.A. (1987).
Prospective Study of 114 Consecutive Episodes of
Staphylococcus aureus Bacteremia. Rev. Infect. Dis. 9:
891–907.
Necula, B.S., Fratila-Apachitei, L.E., Zaat, S.A.J., Apachiteu,
I. and Duszczyk, J. (2009). In Vitro Antibacterial Activity
of Porous TiO2-Ag Composite Layers against Methicillinresistant Staphylococcus aureus. Acta Biomater. 5: 3573–
3580.
Pastuszka, J.S., Kyaw Tha Paw, U., Lis, D.O., Wlazło, A.
and Ulfig, K. (2000). Bacterial and Fungal Aerosol in
Indoor Environment in Upper Silesia, Poland. Atmos.
Environ. 34: 3833–3842.
Pibiri, M.C., Goel, A., Vahekeni, N. and Roulet, C.A.
(2006). Indoor Air Purification and Ventilation Systems
1044
Han et al., Aerosol and Air Quality Research, 15: 1035–1044, 2015
Sanitation with Essential Oils. Int. J. Aromather. 16: 149–
153.
Rahman, M.M., Richardson, A. and Sofian-Azirun, M.
(2010). Antibacterial Activity of Propolis and Honey
against Staphylococcus aureus and Escherichia coli.
Afr. J. Microbiol. Res. 4: 1872–1878.
Ren, P., Jankun, T.M. and Leaderer, B.P. (1999).
Comparisons of Seasonal Fungal Prevalence in Indoor
and Outdoor Air and in House Dusts of Dwellings in
One Northeast American County. J. Exposure Sci. Environ.
Epidemiol. 9: 560–568.
Saito, M., Hosoyama, H., Ariga, T., Kataoka, S. and Yamaji,
N. (1998). Antiulcer Activity of Grape Seed Extract and
Procyanidins. J. Agric. Food. Chem. 46: 1460–1464.
Shin, J.H., Park, J.S. and Bae, D.H. (2011). Fabrication of
Supersaturated Cu–Sn Alloy Sheets and Their Antibacterial
Properties. Met. Mater. Int. 17: 441–444.
Siersted, H.C. and Gravesen, S. (1993). Extrinsic Allergic
Alveolitis after Exposure to the Yeast Ehodotorula rubra.
Allergy 48:298–299.
Wang, Y.Q., Zhang, H.M. and Wang, R.H. (2008).
Investigation of the Interaction between Colloidal TiO2
and Bovine Hemoglobin Using Spectral Methods. Colloids
Surf., B 65:190–196
Received for review, September 17, 2014
Revised, December 11, 2014
Accepted, January 16, 2015