Triclocarban in Human Urine Presentation

Vinh Tran
Chem 4101 - Fall 2011
December 9, 2011
• Triclocarban (TCC, 3,4,4’trichlorocarbanilide) is one of the
most widely used antimicrobial
agents found in soaps and personal
care products.1
• In the United States, many
antimicrobial bar soaps contain up to
1.5% of TCC. 1
• Due to its widespread use, TCC have
been found in surface waters and in
biological organisms such as algae
and snails. 1
MW: 315.58 g/mol
(possible contaminant)
MW: 289.54 g/mol
• Studies show at high concentrations, TCC may act as an
agonist to testosterone and other steroids also known as
endocrine disrupting substances (EDS). 1,2
• Animal studies show EDS are linked to cancer, reproductive
failure, and developmental anomilies. 1,2
With continued use of antimicrobial agents and soaps
through daily hygienic activities TCC will enter into the human
biological system. By analyzing urine samples, human
exposure to TCC will be detected.
1. Representative sample group chosen from individuals with
similar detergent usage/habits – this minimizes fluctuation
in analysis
2. Divide into two main focus areas:
1. Shower study – personal hygiene
2. Dish study – TCC ingestion through use of utensils, etc.
3. Prior to experiment, participants will be given
shampoo/body wash or dish washing detergent not
containing TCC (sample standard)
4. Analyze urine samples daily for each participant
1. Control (non TCC Dish and Shower)
2. Dish Study (TCC dish and non TCC shower)
3. Shower Study (non TCC Dish and TCC Shower)
4. Dish and Shower Study (TCC Dish and Shower)
UV-Vis Spectrometry
Rapid means of analysis,
high precision
Not selective for TCC
because a contaminating
species (TCS) absorbs at
similar wavelength4
Capillary Electrophoresis
High separation
efficiency (more
theoretical plates),
separates based on
charge and size
With such small sample
size may not detect
material after separation
Allows detection of
possible stable
Time efficiency, would
still require LC to
quantify analytes 5
A HPLC/MS-triple quadrupole analytical method will be used
to quantify and detect trace levels of Triclocarban. This
method was chosen because liquid chromatography will allow
separation of polar contaminating analytes in the matrix and a
triple quadrupole MS will allow the selective identification and
quantification of TCC in the urine samples.
• The samples will be ran in reverse-phase since TCC is a
hydrophobic and polar compound allowing for its rapid
elution time.
• Although GC is a more sensitive technique and would
provide better trace analysis, TCC has low volatility and
thus would not be compatible.
Direct Urine Analysis
Aliquots of urine will be
mixed with an internal
standard (IS) solution 1/1
(v/v), vortexed, followed by
Samples will be directly
injected to the HPLC column
for analysis
Urine Analysis after
HCl added to sample to
produce final acid
concentration of 1M
Mixtures are vortexed and
neutralized with 6M aq NaOH
Aliquots of resulting solution
will be mixed with IS solution
1:4 (v/v) and analyzed
• Isotopically labeled TCC (4’-Chlorophenyl-13C6) will be
used as the internal standard.
• Deuterated TCC will also be used a control
• HPLC: The mobile phase will be 70% acetonitrile/30%
water, 10 mM acetic acid, 0.4 mL/min3
• Column: C18, 2.1 x 50 mm, 1.7 µm
• Mass spectrum of samples will be analyzed in comparison
to the standard and control
Figures of Merit1
Limit of Detection:
Limit of Quantification:
0.05 nM = 15.8 ng/L
0.3 nM = 94.7 ng/L
Agilent 6400 LC/MS-triple quad
Relative Intensity (%)
• The expected mass
spectrums of TCC (A) and
deuterium labeled TCC (B)
are indicated by the peaks
at 315 and 322 m/z.3
• The TCC acetic adduct is
also observed at 375 and
382 m/z. 3
• The internal standard peak
will observed at 328 m/z
(spectrum not shown)
Using HPLC/MS as the analytical technique, this method will
allow adequate detection of trace amounts of Triclocarban in
human urine samples of the two separate studies of hygiene
and dish exposure to the antimicrobial agent.
A study of the rate of disappearance of TCC in the human
biological system may aid in future toxicological studies.
The alternative electrochemical method that was not chosen
because of cost and time efficiency may provide further
evidence for the ingestion of TCC by detecting its two
hydroxylated derivatives and two nitro derivatives.5
1. Schebb, N. H.; Inceoglu, B.; Ahn, K. C.; Morisseau, C.; Gee, S. J.; Hammock, B. D.
Investigation of Human Exposure to Triclocarban after Showering and Preliminary
Evaluation of its Biological Effects. Environ. Sci. Technol. 2011, 45, 3109-3115.
2. ScienceDaily. Antibacterial Chemical Disrupts Hormone Activities, Study Finds. /releases/2007/12/071207150713.htm (accessed
September 19, 2011).
3. Halden, R. U.; Paull, D. H.; Analysis of Triclocarban in Aquatic Samples by Liquid
Chromatography Electrospray Ionization Mass Spectrometry. Environ. Sci.
Technol. 2004, 38, 4849-4855
4. Jungerman, E.; Beck, E.C. Determination of Germicide Mixtures in Soaps and Detergents.
Journal of the American Oil Chemist's Society. 1961, 38, 513-515
5. Sires, I.; Oturan, N.; Oturan, M. A.; Rodriguez, R. M.; Garrido, J. A.; Brillas, E. Electro-Fenton
Degradation of Antimicrobials Triclosan and Triclocarban. Electrochimica Acta. 2007, 52,

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