This study assessed the antimicrobial activity and longevity of N-halamine (1-chloro-2,2,5,5-tetramethyl-4-imidazoidinone or MC), a non-bleaching compound, coated on various materials in broiler chicken houses. Results showed that MC solutions with a concentration of 0.04% or higher could kill Salmonella Typhimurium and Campylobacter jejuni within 30 minutes. Materials coated with 1% MC maintained effectiveness against both pathogens for at least 4 weeks, with active chlorine decreasing from an initial 1016 atoms/cm2 to 1015 atoms/cm2 over 3 weeks. Therefore, MC shows potential as a novel antim

1. 1. MC Antimicrobial Activity Test
• An aliquot of 100 µL of bacteria suspension
(4×107 CFU/mL) were added into each 4 mL
MC.
• At each contact time, 100 µL samples were
diluted decimally with buffer to 3 dilutions.
Each dilution was plated out on plate in
triplicates and recorded.
Antimicrobial Acivity of N-Halamine Coated Materials
in Brolier Chicken House
Tian Ren1, Mingyu Qiao1, Lei Zhang1, Tung-Shi Huang1, Buket Demir2
1Department of Poultry Science, Auburn University, Auburn, AL 2Department of Chemistry, Auburn University, Auburn, AL
The objectives of this study were to assess the antimicrobial activity and the antimicrobial
longevity of N-halamine (1-chloro-2,2,5,5-tetramethyl-4-imidazoidinone (MC), a non-
bleaching compound)-coated materials in broiler chicken houses. The antimicrobial test of
MC illustrated that 0.04% MC solution was able to kill Salmonella Typhimurium and
Campylobacter jejuni completely at 106 CFU/mL. Results showed that 1% MC treated
samples were able to kill Salmonella Typhimurium and Campylobacter jejuni completely as
long as four weeks, at the concentration of 106 CFU/cm2, where the active chlorine atoms in
the treated coupons decreased from the initial 1016 atoms/cm2 to 1015 atoms/cm2. Therefore,
MC is assumed to be used as a novel antimicrobial agent in chicken broiler house.
N
N
H3C
H3C
NH2
CH3
H
O
Cl
Structure of MC
• N-halamines are ideal antimicrobial agents due to their
superior antimicrobial activity against a broad spectrum of
microorganisms, nontoxicity, high stability, rechargeability
and low cost. Antimicrobial activities of various materials
that coated with N-halamines have been investigated, such
as fabrics (Demir, et al., 2015) and paints (Kocer, et al.,
2011).
• MC is a type of monochlorinated N-halamine which has
been reported possessing a powerful antimicrobial activity
and high stability.
2. Antimicrobial Longevity Test of MC Coated Materials
---- Sample Preparation
• Stainless steel, galvanized metal, plastic, and pressure
treated woods were selected as tested materials. Slides
of each material were cut into 1 inch2 coupons. Cleaned,
autoclaved before use.
• Spray with 0.1% and 1% MC in 95% EtOH, respectively.
Stored in a dark, enclosed cabinet at room temperature.
3. Antimicrobial Longevity Test of MC Coated
Materials ---- Antimicrobial Activity Test
• An aliquot of 25 µL of bacteria suspension was
placed onto the center of a coupon, and covered with
a second identical coupon with MC coated side face
down. Then, a sterile weight was placed on the top of
the second coupon.
• After 2 h contact time, coupons were transferred
Na2S2O3 solution to terminate the reaction as well as
wash off all the attached bacterial. Dilution and
plating out as previous methods. Tests were run on
day 0, 3, 6, and week 2, 3, 4.
4. Active Chlorine Content Determination
• 0.25 g of KI was dissolved into 5 mL of 0.1 N
acetic acid and 45 mL of absolute ethanol. Then
two coupons were added into the flask.
• They were shaken for 2 min to mix well. Then,
standardized Na2S2O3 was used to titrate. The
content of active chlorine was calculated by
using this following equation.
N and V are the normality (equiv/L) and volume (L) of the sodium
thiosulfate, respectively, and A is the surface area (cm2) of the coupon.
Cl+
(atom/cm2
) =
6.02×1023
× N × V
2× A
5
10
30
60
0
10
20
30
40
50
60
70
80
90
100
0
0.02
0.04
0.06
Bacteriareduction(%)
MC concentration (%)
Salmonella Typhimurium
5
10
30
60
0
10
20
30
40
50
60
70
80
90
100
0
0.02
0.04
0.08
0.16
Bacteriareduction(%)
MC concentration (%)
Campylobacter jejuni
1. Antimicrobial Activity of MC Solution
• Higher MC concentration, or longer contact time, the
antimicrobial activity of MC against S. Typhimurium and C. jejuni
increased.
• Solutions containing 0.04% or higher concentration of MC were
able to completely inactivate both bacteria at 106 CFU/mL within
30 min.
2. Antimicrobial Longevity of MC Coated Materials
• Materials coated with 1% MC were able to inactivate all the
inoculated bacteria at 106 CFU/sample within 2 h contact time,
and this antimicrobial activity lasted for at least four weeks.
• Materials coated with 0.1% MC kill more than 99.8% of the S.
Typhimurium and all of the inoculated C. jejuni on the initial day,
and then their antimicrobial activities decrease afterwards.
3. Active Chlorine Content in the Materials
• It did not decrease in the first two weeks after materials were
treated with MC. After three weeks of treatment, it decreased
significantly, from the initial 1016 atoms/cm2 to 1015 atoms/cm2
over this three-week period, and were still able to inactivate all
the inoculated microbes.
• MC is a promising antimicrobial agent, which can be applied into
the chicken broiler house.
• For the purpose of inactivating both S. Typhimurium and C.
jejuni, we found that 1% MC is suitable to be used to coat on
various materials.
1. Demir et al. ACS Applied Materials
& Interfaces. 2015, 7(3), 1752-1757.
2. Kocer et al. ACS Applied Materials
& Interfaces. 2011, 3(8), 3189-3194.
3. Scharff. Journal of Food Protection,
2011. 75(1), 9.
0
0.5
1
1.5
2
2.5
3
3.5
4
0 D 3 D 6 D 2 W 3 W 4 W
Cl+atom/cm2(×1016)
Storage time
0
0.5
1
1.5
2
2.5
3
3.5
4
4.5
5
0 D 3 D 6 D 2 W 3 W 4 W
Cl+atom/cm2(×1016)
Storage time
0
0.5
1
1.5
2
2.5
3
3.5
0 D 3 D 6 D 2 W 3 W 4 W
Cl+atom/cm2(×1016)
Storage time
0
0.5
1
1.5
2
2.5
3
3.5
0 D 3 D 6 D 2 W 3 W 4 W
Cl+atom/cm2(×1016)
Storage time
2. Antimicrobial longevity of MC coated materials and change of chlorine content
Galvanized metal Plastic Pressure treated woodsStainless steel
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
DISCUSSIONRESULTS
CONCLUSION
1. Antimicrobial activity of MC solution
• The Center for Disease Control and Prevention (CDC) estimated that each year about 48
million people were infected with foodborne diseases in the US (CDC 2011), and it also
led to a significant economic loss (Scharff, 2012). With the ever-growing demand in
poultry products in the US (USDA ERS, 2015), incidences of pathogen contamination in
poultry products can significantly affect human health. Therefore, protecting the safety of
poultry products becomes a critical issue for reducing the risk of foodborne diseases.
REFERENCES
1-chloro-2,2,5,5-tetramethyl-
4-imidazolidinone
ePoster