Identification of by-products issued from the catalytic oxidation of toluene by chemical and biological methods

Article abstract of DOI:10.1016/j.crci.2015.09.001

Benzene, toluene, ethylbenzene and xylenes (BTEX) are substances that are very commonly encountered in almost all sectors of the industry. Consequently, these molecules may be present in large amounts in industrial exhausts loaded with VOCs. An effective method for their elimination is catalytic oxidation, which is an economic and ecologic alternative to thermal oxidation. The aim of this work is to reveal the by-products issued from the total oxidation of toluene by palladium-based catalysts. The identification of these by-products was done using a chemical and toxicological approach. A toxicological validation of the developed catalysts was performed by coupling the catalytic system with an air-liquid interface (aLI) system, called Vitrocell, to expose lung cells to catalytic exhausts.

Full text of DOI:10.1016/j.crci.2015.09.001

G Model
CRAS2C-4137; No. of Pages 10
C. R. Chimie xxx (2015) xxx–xxx
Contents lists available at ScienceDirect
Comptes Rendus Chimie
www.sciencedirect.com
International Symposium on Air & Water Pollution Abatement Catalysis (AWPAC) – Catalytic pollution
control for stationary and mobile sources
Identification of by-products issued from the catalytic
oxidation of toluene by chemical and biological methods
`
Identification de sous-produits de l’oxydation catalytique du toluene par
´
des methodes chimiques et biologiques
Julien Brunet a, Eric Genty a, Yann Landkocz a, Margueritta Al Zallouha a,
a
a
a
b
´
Sylvain Billet , Dominique Courcot , Stephane Siffert , Diane Thomas ,
Guy De Weireld b, Renaud Cousin a,
*
a Unite´ de chimie environnementale et interactions sur le Vivant, MREI1, 145, avenue Maurice-Schumann, 59140 Dunkerque, France
b
´
Faculte polytechnique de Mons, 20, place du Parc, 7000 Mons, Belgium
A R T I C L E I N F O
A B S T R A C T
Article history:
Benzene, toluene, ethylbenzene and xylenes (BTEX) are substances that are very commonly
encountered in almost all sectors of the industry. Consequently, these molecules may be
present in large amounts in industrial exhausts loaded with VOCs. An effective method for
their elimination is catalytic oxidation, which is an economic and ecologic alternative to
thermal oxidation. The aim of this work is to reveal the by-products issued from the total
oxidation of toluene by palladium-based catalysts. The identification of these by-products
was done using a chemical and toxicological approach. A toxicological validation of the
developed catalysts was performed by coupling the catalytic system with an air–liquid
interface (aLI) system, called Vitrocell1, to expose lung cells to catalytic exhausts.
Received 29 November 2014
Accepted after revision 7 September 2015
Available online xxx
Keywords:
Environmental chemistry
Heterogeneous catalysis
Oxidation
Palladium
´
ß 2015 Academie des sciences. Published by Elsevier Masson SAS. All rights reserved.
´
´
R E S U M E
Mots cle´s :
`
`
´
`
`
´
`
Le benzene, le toluene, l’ethylbenzene et les xylenes (BTEX) sont des molecules tres
Chimie environnementale
couramment rencontre´es dans l’industrie, et ceci dans tous les secteurs d’activite´. Ces
´
´
`
Catalyse heterogene
´
ˆ
´
´
molecules peuvent donc etre presentes en quantites importantes dans les effluents
Oxydation
´
´
´
industriels charges en COV. Une methode efficace pour l’elimination de ces COV est
l’oxydation catalytique, qui est une alternative e´conomique et e´cologique a` l’oxydation
thermique. L’objectif de ce travail est d’identifier les sous-produits issus de l’oxydation
Palladium
`
`
totale du toluene par des catalyseurs a base de palladium. L’identification de ces sous-
produits a e´te´ faite en utilisant une approche chimique et toxicologique. Une validation
toxicologique des catalyseurs de´veloppe´s a e´te´ re´alise´e en couplant le syste`me catalytique
a un systeme d’interface air–liquide (ALI), appele Vitrocell1, afin d’exposer des cellules
pulmonaires aux gaz issus du traitement catalytique.
`
`
´
ß 2015 Acade´mie des sciences. Publie´ par Elsevier Masson SAS. Tous droits re´serve´s.
*
Corresponding author.
E-mail address: Renaud.Cousin@univ-littoral.fr (R. Cousin).
http://dx.doi.org/10.1016/j.crci.2015.09.001
1631-0748/ß 2015 Acade´mie des sciences. Published by Elsevier Masson SAS. All rights reserved.
Please cite this article in press as: Brunet J, et al. Identification of by-products issued from the catalytic oxidation of
toluene by chemical and biological methods. C. R. Chimie (2015), http://dx.doi.org/10.1016/j.crci.2015.09.001

G Model
CRAS2C-4137; No. of Pages 10
2
J. Brunet et al. / C. R. Chimie xxx (2015) xxx–xxx
1. Introduction
As all destructive techniques for treating VOCs emis-
sions (thermal oxidation, plasma, biological treatment. . .),
catalytic oxidation can generate some by-products that are
intermediate compounds more difficult to oxidize than the
substrate and can be potentially released into the
atmosphere. Some studies have demonstrated the forma-
tion of its intermediaries, particularly for oxygenates [16–
18] and aromatics [19–21] compounds. Some of these
compounds are known to interact with cell macromole-
cules, like DNA. They can therefore exert their toxicity, and
induce some pathologies [22]. Recent concern has centred
on the effects of continuous exposure to low concen-
trations of VOCs, like toluene and benzene, both occupa-
tionally and environmentally. Some of them have for a long
time been recognised as carcinogenic for humans. For
example, benzene, the major by-product produced by
toluene catalytic degradation, is known to induce acute
myeloid leukaemia (AML). In a previous work, we showed
that A549 cells were a good cell model for the determina-
tion of benzene and toluene effects on human lung cells.
Mutation hotspots in the tumour suppressor gene TP53,
which are the most common genetic alterations involved
in human cancer, were the same in AML and in A549 in
vitro exposed to benzene [23]. Consequently, several
questions can be asked. Are all by-products removed
when the conversion of the initial VOC is total? If not, is it
possible to find a compromise between efficiency and
economic viability? Finally, can they have toxicological or
ecotoxicological impacts? So, an important part of this
work concerns the oxidation of toluene, specially the
identification of toluene by-products. In a first time, light-
off curves were determined by microGC analysis, which are
suitable for fast and sensitive analyses. A coupling with a
quadrupole mass spectrum allowed for a more complete
identification of by-products. In a second time, coupling
the catalytic process with a cell exposure system permitted
us to study the impact of the by-products on human lung
cells by measuring the gene expression of xenobiotic
metabolizing enzymes (XMEs) involved in the biotrans-
formation of the organic compounds. Coupling an air–
liquid interface (ALI) system, called Vitrocell1, to a catalyst
test was performed for the first time during this study.
Volatile organic compounds (VOCs) are known as one of
the major contributors to atmospheric pollution. Their
anthropic releases are particularly important and have
some consequences in health, environment, and construc-
tion materials.
An important part of these VOCs are single-aromatic
molecules, which are principally represented by the BTEX
class (benzene, toluene, ethylbenzene and xylenes).
Indeed, these compounds are widely used in all industrial
sectors: fuel additives, solvents (paints, varnishes, lac-
quers, inks, adhesives and glues), cleaning or extracting
agents, synthesis precursors (production of explosives,
pesticides, drugs and polymers). In addition to their highly
inflammable and explosive character, BTEX compounds
are highly volatile and have a high mobility in air, soil and
water. Being very irritating and highly toxic (disorders of
the digestive, respiratory, nervous and neurological
systems), they can quickly contaminate and poison an
ecosystem.
Therefore, it is necessary to set up techniques for
reducing BTEX emissions. Nowadays, catalytic oxidation
represents an economical and environmental alternative
to the thermal oxidation, the latter being the most
common in the industry. Indeed, the use of catalysts
allows a drastic reduction of the process temperature
(200–500 8C), which prevents the formation of toxic by-
products (NOx, dioxins) and reduces the energetic cost of
the process.
In order to treat gaseous effluents loaded in BTEX from
industry, the catalytic total oxidation of toluene was here
studied using several Pd-based materials. These materials
have been selected by taking into account their cost,
activity, stability (thermal, mechanical and chemical), and
it can be easy to produce this material at a large scale.
Concerning supports, titanium oxide and HY Faujasite
show interesting performances in the catalytic oxidation
reactions of BTEX [1–4]. Then, cerium oxide is known for
the treatment of automobile exhausts, but this material
also shows good performances as a support for BTEX
oxidation [5,6]. Finally, aluminium oxides were the most
common support in catalytic process, especially
and -Al2O3. They present some interesting properties, like
a high thermal stability for -Al2O3 or a high specific
surface area with acidic properties for -Al2O3, and also
a-Al2O3
g
a
2. Materials and method
g
present good performances as a support for BTEX oxidation
[7–10]. Palladium was chosen as the active phase seeing
that it is the most suitable noble metal in an oxidizing
environment. In fact, palladium is more resistant than
platinum to sintering, vaporizing metal species and
poisoning [11–13]. These properties can be partly explai-
ned by the reversible transition between the two phases of
palladium, Pd0/PdIIO, which has been demonstrated by
Cordi et al. [11]. Moreover, palladium and platinum show
very close performances for VOCs catalytic oxidation
2.1. Preparation of supports
The ceria support was prepared by a precipitation way. A
solution of Ce(NO3)3ꢀ6H2O was added dropwise to a NaOH
solution with a molar ratio of 5 under stirring during 3 h.
The suspension was left under agitation during 2 h at
ambient temperature. Then, the suspension was filtered
and washed 6 times with 200 mL of hot deionized water
(ꢁ 60 8C). The solid was dried 24 h at 100 8C and calcined for
4 h at 500 8C (heating at 1 8C/min) under an air flow (2 L/h).
The titania support was prepared by a sol–gel way. A
15-wt% cetyltrimethylammonium bromide (CTMABr) mi-
cellar solution was added dropwise to a solution of sulfuric
acid (pH = 2) under magnetic agitation during 3 h at
40 8C. Then, a solution of titanium (IV) tetraethoxide
[14,15]. Therefore, palladium is
between performance and cost. Thus, palladium was
impregnated to 0.5 wt% on -Al2O3, TiO2, HY Faujasite
a good compromise
a
and CeO2. A commercial catalyst (0.5 wt% Pd/g-Al2O3) was
selected as a reference.
Please cite this article in press as: Brunet J, et al. Identification of by-products issued from the catalytic oxidation of
toluene by chemical and biological methods. C. R. Chimie (2015), http://dx.doi.org/10.1016/j.crci.2015.09.001

G Model
CRAS2C-4137; No. of Pages 10
J. Brunet et al. / C. R. Chimie xxx (2015) xxx–xxx
3
was added dropwise with a molar ratio CTMABr/Ti4+ of
0.33. The agitation was maintained for 1 h at 40 8C before
the addition. Afterwards, the solution was placed in a
Teflon sheath that was inserted in a stainless steel
autoclave. This one was placed during 48 h in an oven at
60 8C for hydrothermal treatment. The surfactant was
eliminated by Soxhlet extraction with ethanol. The solid
was dried 48 h at 60 8C and calcinated 4 h at 400 8C
(heating at 1 8C/min) under an air flow (2 L/h).
an air flow (2 L/h). For the impregnated catalysts, they
were reduced for 2 h at 200 8C (heating at 1 8C/min) under a
dihydrogen (5.0) flow (2 L/h). The organic compounds
were analysed by gas chromatography with a CP-4900
microGC (Agilent) coupled with a Pfeiffer-Vacuum Omnis-
tar Quadrupole Mass Spectrometer (QMS-200). The mass
spectrometer is configured with a measurement mode MID
(Multiple Ion detection) that is adapted for the detection of
compounds trace. CO and CO2 were quantified with an
infrared analyser (ADEV 4410 IR).
HY Faujasite zeolite was prepared by ionic exchange. A
commercial NaY Faujasite zeolite (Si/Al: 2.7; Sigma–
Aldrich) was slurried in a solution of ammonium nitrate
Toluene conversion was calculated considering pro-
ducts and by-products and as a function of the number of
carbons for each compound:
2.0 mol/L with
a
molar ration NH4+/Na+ of 20. The
suspension was maintained 18 h at 80 8C under magnetic
agitation. Then, the suspension was filtered and washed
with hot deionized water (ꢁ 60 8C) to a neutral pH. These
operations were repeated three times. Then, the solid was
dried one night at 100 8C and calcined for 4 h at 400 8C
(heating at 1 8C/min) under an air flow (2 L/h).
ꢀ
ꢁ
½CO2ꢂT þ ½COꢂT þ ½C6H6ꢂ ꢃ 6
½CO2ꢂ þ ½COꢂ þ ½C6H6ꢂ ꢃ 6 þT½C7H8ꢂ ꢃ 7
XT
¼
ꢃ 100
T
T
T
T
where XT is the toluene conversion at temperature T (%),
[I]T is the concentration of the compound I at temperature
T (ppm).
The catalytic activity was calculated at a toluene
conversion of 20% and considering a plug-flow reactor:
For the
a-Al2O3, a commercial powder was chosen
(Acros Organics).
2.2. Palladium impregnation
Q
273:15 ½C7H8ꢂ0
X
1
Ai ¼
ꢀ
ꢀ
ꢀ ꢀ
m SBET
106
VM
T20
Palladium was deposed on the support (
CeO2 and HY Faujasite) by an aqueous impregnation
method (0.5 wt% Pd). The solid was suspended in an
a
-Al2O3, TiO2,
where Ai is the catalytic activity mol/(m2ꢀh), Q is the
volume flow (L/h), VM is the molar volume (L/mol), T20 is
appropriate volume of
a
palladium nitrate solution
the temperature of the catalyst for a 20% toluene
(0.25 g/L). The suspension was maintained during 18 h
at 60 8C. Then, water was removed by a rotary evaporator.
The solid was dried one night at 100 8C before calcination
at 400 8C (heating at 1 8C/min) under an air flow (2 L/h).
conversion (K), [C7H8]0 is the toluene initial concentration
(ppm), X is the toluene conversion (%); m is the mass of the
catalyst (g), SBET is the BET surface area of the catalyst (m2/
g).
A
commercial 0.5 wt% Pd/
g-Al2O3 catalyst (Acros
Catalytic performances were compared by considering
Organics) was chosen to complete the study.
T50 and T100, which respectively correspond to the
temperatures at which 50 and 100% of toluene were
converted. The value of T100 was chosen in order to achieve
total oxidation for a catalytic industrial application.
2.3. Characterization of the catalysts
The BET surface area was measured by nitrogen
adsorption at –196 8C in a Thermo-Electron QSurf series
Surface Area Analyzer apparatus. The sample was degassed
before measurement at 130 8C under a helium flow.
Adsorption was made with a gas mixture of 30% N2–70%
He. The desorption of gaseous N2 was quantified with a
thermal conductivity detector.
An amount of 50.0 mg of powder were dissolved in 5 mL
of aqua regia (HNO3/HCl 1:2) under microwave irradiation
during 30 min (CEM, model MARSXpress). Then, the
solution was extended to 50.0 mL with ultrapure water
2.5. Cells, culture conditions and coupling of the exposure
system of the cells
Toxicological studies were made by coupling the
catalytic process with an air–liquid interface (ALI) expo-
sure system Vitrocell1 in order to expose human lung
epithelial adenocarcinoma cells A549. In vitro toxicological
studies involve the direct or indirect exposure of human
cells to xenobiotics, and should be carried out under
conditions resembling the in vivo situation as far as
possible. Among the cell exposure systems, the ALI system
appears as the best compromise to expose cells to gaseous
mixtures, because it allows the control of the level of
exposure during all the exposure period. A549 cells were
cultured in sterile plastic flasks (Corning1, Fisher Scientif-
ic), in a minimum essential medium (MEM) supplemented
and filtered with a 0.45-mm cellulosic micro-filter. The
analysis was performed with an ICP-OES (Thermo, model
ICAP 6300 Duo).
2.4. Catalytic test
The total oxidation of toluene was studied in
a
with 5% (v/v) of fetal bovine serum, 1% (v/v) L-glutamin and
conventional fixed-bed reactor loaded with 100 mg of
catalyst. A toluene/air mixture was generated with a
saturator in order to obtain 1000 ppm in a flow of 100 mL/
min. Tests were made between 50 and 400 8C with a
temperature ramp of 1.5 8C/min. Catalysts were pre-
treated at 200 8C (heating at 1 8C/min) during 2 h under
1% (v/v) penicillin and streptomycin (Invitrogen-Life
Technologies). These cells are known to activate VOCs
using XMEs [22]. Cells were incubated at 37 8C, in a 5% CO2
and 100% humidity atmosphere until 80% of confluency.
Then, cells were washed twice with PBS and dissociated
with trypsin + EDTA (Gibco Invitrogen) at 37 8C during
Please cite this article in press as: Brunet J, et al. Identification of by-products issued from the catalytic oxidation of
toluene by chemical and biological methods. C. R. Chimie (2015), http://dx.doi.org/10.1016/j.crci.2015.09.001

G Model
CRAS2C-4137; No. of Pages 10
4
J. Brunet et al. / C. R. Chimie xxx (2015) xxx–xxx
3 min. The cells were collected by centrifugation and
DCt = Cttarget gene – Ct18S). RQ corresponds to the compari-
counted with
a
Cellometer1
.
Four hours before ALI
son of exposed cells versus unexposed cells (i.e. purified
air). The results are expressed as median values of
triplicates.
exposure experiments, 5 ꢃ 105 cells were seeded in
Transwell insert with a surface area of 4.7 cm2 in MEM
supplemented with 1% (v/v) of L-glutamin.
Three culture modules, each one containing three
Transwell inserts with A549 cells maintained at 37 8C,
were exposed in parallel to different gases:
3. Results
3.1. Catalytic results
ꢄ
ꢄ
gas from the catalytic exhaust without dilution;
gas from the catalytic exhaust with a dilution of 10% in
purified air;
Table 1 displays the specific surface areas and the
palladium content of the catalysts. These different catalytic
materials have been studied for the catalytic total
oxidation of toluene. Light-off curves of supports have
been determined and are reported in Fig. 1. It could be
observed (Fig. 1, Table 2) that these materials have a
catalytic activity without palladium. HY Faujasite and TiO2
have similar activity values with T50 (temperatures at
which 50% of toluene was converted) of 302 and 310 8C,
respectively. With a T50 of 233 8C, CeO2 shows the better
performance when the conversion is above 80%. On the
contrary, alumina shows the worst performance; this
support is only active over 350 8C and conversion does not
exceed 75% at 400 8C.
The toluene conversion curves for the impregnated
supports have been determined, as well as for the
commercial catalyst (Fig. 2). Table 2 displays the T50 and
T100 of unimpregnated and impregnated supports. For all
samples, Pd impregnation on supports increases the
performance of materials, as expected. The commercial
ꢄ
purified air (control).
On the one hand, the 10% dilution exposure mimics a
quite realistic dilution in the atmosphere. On the other
hand, this helped us to evaluate the dose–response
relationship of catalytic exhausts. Exposures are perfor-
med at fixed catalytic conditions during 1 h. These
conditions correspond to the beginning of 100% toluene
total conversion (T100). After 1 h of exposure to catalytic
exhausts, the Transwell inserts containing cells were
rinsed with PBS and supernatants were recovered. Both
were frozen at –80 8C until use.
2.6. Toxicological analysis
The cell membrane integrity of A549 cells was
evaluated by measurement of extracellular released
lactate deshydrogenase (LDH) in the culture medium,
according to the manufacturer’s instructions (Roche).
catalyst, Pd/g-Al2O3, shows the best performance in terms
of T50 and T100 at 179 8C and 200 8C, respectively. However,
as regards the activity value (Table 2) obtained for the
catalysts at a toluene conversion of 20%, it could be
Briefly, 100
mL of each cell-free culture medium were
transferred in triplicate into a 96-well plate. Then, 100
m
L
observed that the catalyst Pd/
activity.
a-Al2O3 possesses the best
of LDH-assay reaction mixture were added to each well.
After 30 min of incubation at 37 8C in the dark, the optical
density at 490 nm was measured using a microplate
reader.
3.2. Identification of by-products
Total RNA extraction from exposed or unexposed cells
and cDNA synthesis from mRNA were carried out using
TaqMan1 Gene expression Cell-to-Ct kit (Ambion, Invi-
trogen-Life Technologies), following the manufacturer’s
instructions. Gene expression of xenobiotic metabolizing
enzymes (XMEs) (i.e. CYP2E1, CYP2S1, CYP1A1, CYP1B1,
EPHX1, NQO1) and 18S rRNA as housekeeping gene
were quantified using TaqMan1 Gene Expression Assay
(Applied Biosystems) combined with TaqMan1 specific
primers sets (CYP2E1; Hs00559368_m1, CYP2S1;
Hs00258076_m1, CYP1A1; Hs01054797_g1, CYP1B1;
Hs02382916_s1, EPHX1; Hs01116806_m1, NQO1;
Hs02512143_s1 and 18S rRNA; Hs99999901_s1) on a
7500 Fast Real-Time PCR System. These XMEs play a major
role in the biotransformation of organic compounds, like
VOCs as expected, but also of heavier chemicals, like
polycyclic aromatic hydrocarbons (PAHs).
3.2.1. MicroGC analysis
Lars and Andersson [19] have studied the formation of
the by-products generated in the catalytic oxidation of
toluene. With the 23 products observed by GLC–MS, they
proposed three reaction mechanisms describing the
catalytic oxidation of toluene as a function of the initiation
mode. However, the study had been made under static
conditions and promoting partial oxidation. In dynamic
conditions, the knowledge of the generated by-products is
limited for VOCs total catalytic oxidation. In order to define
the nature of by-products, this study was performed.
Analysis by microGC, which is adapted for quick (less
than 4 min) and sensitive analysis, has allowed the
Table 1
Characteristics of catalysts used.
After 40 cycles of amplification, each gene expression
was determined using cycle threshold (Ct) measured by
Sequence Detection Software v2.0.3 and relative quantifi-
cation (RQ) between exposed and untreated cells was
achieved for each exposure using the following calculation:
Pd/
1
a
-Al2O3 Pd/TiO2 Pd/HY Pd/CeO2 Pd/
g-Al2O3
BET surface
area (m2/g)
Palladium
252 900 93 252
0.48
0.41
0.46 0.40
0.40
content (wt%)
DD
Ct
RQ = 2ꢅ
(where DDCt = DCtexposed – DCtunexposed and
Please cite this article in press as: Brunet J, et al. Identification of by-products issued from the catalytic oxidation of
toluene by chemical and biological methods. C. R. Chimie (2015), http://dx.doi.org/10.1016/j.crci.2015.09.001

G Model
CRAS2C-4137; No. of Pages 10
J. Brunet et al. / C. R. Chimie xxx (2015) xxx–xxx
5
Fig. 1. (Colour online.) Light-off curves of the supports.
Table 2
many years. Consequently, it is important to know under
which conditions and in which amounts this compound
may be issued by the catalytic process. Benzene could be
quantified by microGC, emission profiles have been
determined according to toluene conversion and working
temperature for each catalyst. Fig. 3 shows a general profile
T50, T100a, and activity of catalysts for the catalytic oxidation of toluene.
Support
Impregnated support
T50
T100
T50
T100
Ai
(8C)
(8C)
(8C)
(8C)
(mol/(m2ꢀh))
a-Al2O3
392
310
302
233
–
> 400
> 400
334
218
259
222
197
179
233
285
242
235
200
2.84 ꢃ 10ꢅ7
1.29 ꢃ 10ꢅ9
3.18 ꢃ 10ꢅ10
3.34 ꢃ 10ꢅ9
1.27 ꢃ 10ꢅ9
of the benzene-produced amount as
a function of
TiO2
HY
temperature. Benzene production begins at the same
temperature as toluene conversion. The issued quantity
increases rapidly until it reaches a maximum emission,
after which the issued amount of benzene decreases
slowly. The benzene emission profile may be characterized
by four parameters, which are also illustrated in Fig. 3:
CeO2
> 400
–
Pd/g-Al2O3
a
Temperatures at which 50 and 100% of toluene, respectively, were
converted.
detection of the majority of the oxidation intermediates.
Benzene was identified as the main by-product resulting
from toluene total oxidation. Its toxic properties, particu-
larly its carcinogenic properties, have been known for
ꢄ
ꢄ
Ti: the temperature at which a by-product is detected (8C);
Tmax: the temperature at which the amount of the by-
product is the largest (8C);
Fig. 2. (Colour online.) Light-off curves of the catalysts loaded with 0.5 wt% of palladium.
Please cite this article in press as: Brunet J, et al. Identification of by-products issued from the catalytic oxidation of
toluene by chemical and biological methods. C. R. Chimie (2015), http://dx.doi.org/10.1016/j.crci.2015.09.001

G Model
CRAS2C-4137; No. of Pages 10
6
J. Brunet et al. / C. R. Chimie xxx (2015) xxx–xxx
Fig. 3. (Colour online.) Typical profile of benzene emission and its characteristic parameters.
ꢄ
ꢄ
Qmax: the maximum observed amount of the considered
by-product (ppm);
Tf: the temperature at which the by-product is totally
oxidized (8C).
peak is more significant with a Qmax value of 43 ppm. As
regarding the values of Ti, Tmax, and Tf, the results are quite
different for each catalyst and no trend can be evidenced.
For a better understanding of the by-product generated
profile, the values of the range emission of benzene (noted
R) and its persistence (noted P) were evaluated then. The
first term, R, can be calculated as the difference Tf – Ti. The
second term, P, can be considered the temperature that
must be added to the T100 of toluene to have a total
degradation of toluene and benzene. The persistence can
be calculated by the difference Tf – T100. The value of
toluene conversion X corresponding to the maximum
emission of benzene was also determined. The values of R,
P, and X are reported in Table 3. Regarding the emission
range, this value is relatively large, with an average of
126 8C. Pd/HY is especially noteworthy, with a R value of
67 8C. Concerning the persistence, P values clearly show
In Fig. 4, the dashed lines correspond to benzene
production as a function of the toluene light-off curve and
temperature for the four impregnated catalysts and the
commercial one. The results show quite similar benzene
emission profiles for all the catalysts, and this despite the
nature of the support. Only Pd/TiO2 shows a different
profile, with a particularly intense emission peak. Table 3
reports the values of the four parameters established to
describe the benzene profiles: Ti, Tmax, Qmax and Tf. The data
show a similar magnitude of Qmax for the catalysts Pd/
Al2O3, Pd/HY, Pd/CeO2 and Pd/ -Al2O3 with values ranging
between 7 and 14 ppm. However for Pd/TiO2, the emission
a-
g
Fig. 4. (Colour online.) Toluene conversion and production of benzene versus temperature for the impregnated catalysts.
Please cite this article in press as: Brunet J, et al. Identification of by-products issued from the catalytic oxidation of
toluene by chemical and biological methods. C. R. Chimie (2015), http://dx.doi.org/10.1016/j.crci.2015.09.001

G Model
CRAS2C-4137; No. of Pages 10
J. Brunet et al. / C. R. Chimie xxx (2015) xxx–xxx
7
Table 3
Table 4
Parameters characterising the emission profiles of benzene.
Parameters characterising emission profiles of carbon monoxide.
Ti
Tmax
Qmax
Tf
R
P
X
Ti
Tmax
Qmax
Tf
R
P
X
(8C)
(8C)
(ppm)
(8C)
(8C)
(8C)
(%)
(8C)
(8C)
(ppm)
(8C)
(8C)
(8C)
(%)
Pd/
a
-Al2O3
176
150
182
142
129
220
285
222
212
193
8
43
10
7
300
325
249
298
244
124
175
67
67
40
7
86
Pd/
a
-Al2O3
198
222
125
–
213
271
222
–
5
220
284
229
–
22
62
104
–
–13
–1
–13
–
33
73
47
–
Pd/TiO2
Pd/HY
100
47
Pd/TiO2
Pd/HY
117
71
0
Pd/CeO2
156
115
63
44
83
Pd/CeO2
Pd/
g-Al2O3
14
95
Pd/
g-Al2O3
–
–
0
–
–
–
–
that benzene is not removed when toluene conversion is
complete. So, it is necessary to increase the working
temperature for all catalysts in order to completely remove
toluene and benzene. This point is particularly important
because an increase in the working temperature has a
direct and non-negligible impact on the operating cost of
the treatment unit on an industrial scale. The catalyst Pd/
HY stands out again with a P value of only 7 8C. In view of
these two points, the catalyst Pd/HY, which is not the most
active, is the most selective catalyst concerning benzene
emissions. Finally, if we consider the X values, the data
show that benzene is mainly issued for high conversions in
toluene.
A second major by-product evidenced in this study is
carbon monoxide. It was also detected during the catalytic
oxidation of toluene. It has been quantified and emission
profiles of carbon monoxide have been traced. These
profiles can be described in the same manner as those of
benzene. Fig. 5 shows the toluene light-off curve and
carbon monoxide production as a function of temperature
for the five catalysts. Previously, it could be observed that
the emission profiles of benzene are fairly homogeneous.
Concerning the profiles obtained for CO, heterogeneity is
revealed. Indeed, the values of Qmax, given in Table 4, are
relatively high for the Pd/TiO2 and Pd/HY catalysts with
71 and 117 ppm, respectively. On the contrary, the catalyst
5 ppm. As for catalysts Pd/CeO2 and Pd/g-Al2O3, they show
no CO emission. Regarding the range emission, the R values
are generally lower than for benzene. Regarding persis-
tence, P values indicate that carbon monoxide is complete-
ly oxidized before the total conversion of toluene is
reached. So, it can be concluded that carbon monoxide has
no impact on the working temperature for
a total
conversion of toluene. These two points may be explained
by the fact that palladium is a suitable metal for the
oxidation of carbon monoxide [11,13].
3.2.2. Mass spectrometry analysis
MicroGC analysis has helped highlight the role of
benzene, which is considered the main by-product of the
catalytic oxidation of toluene. But this device does not
allow the detection of molecules below the ppm level.
Thus, molecules emitted into much lower concentrations
can be detected. For this reason, mass spectrometer was
coupled with microGC, allowing analysis of by-product
traces. The apparatus, configured for dynamic analysis, can
follow the evolution of the characteristic fragments of
molecules that interest us as a function of temperature.
These fragments were chosen primarily based by-products
observed in the work of Lars and Andersson [19]. Charac-
teristic fragments of C1–C6 paraffins and some common
families of polycyclic aromatic hydrocarbons (C10H8,
Pd/
a-Al2O3 has a very low emission peak with a Qmax of
C14H10, C18H12, C22H14) were also selected.
Fig. 5. (Colour online.) Toluene conversion and production of carbon monoxide versus temperature for the impregnated catalysts.
Please cite this article in press as: Brunet J, et al. Identification of by-products issued from the catalytic oxidation of
toluene by chemical and biological methods. C. R. Chimie (2015), http://dx.doi.org/10.1016/j.crci.2015.09.001

G Model
CRAS2C-4137; No. of Pages 10
8
J. Brunet et al. / C. R. Chimie xxx (2015) xxx–xxx
Table 5
Traces of by-products observed by mass spectrometry.
By-products observed
Pd/a
-Al2O3
Pd/TiO2
Pd/HY
Pd/CeO2
Pd/g-Al2O3
Hydrogen
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
Methane
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
Furan
Maleic anhydride
2-Methylmaleic anhydride
Benzene
Benzaldehyde
Benzyl alcohol
Benzoquinone
2-Benzofurane-1,3-dione
Dihydroanthracene
Methyldiphenylmethane
Naphthacene
H
H
Table 5 lists the molecules that were identified by mass
spectrometry. In general, the same compounds are observed
for all the catalysts. On the one hand, benzyl alcohol,
benzaldehyde, benzene, 1,4-benzoquinone and maleic
anhydride were identified. These compounds are derived
from the main way toluene oxidation proposed by Lars and
Andersson[19]andgiveninFig. 6. Phenolandhydroquinone
were not detected by mass spectrometry; this may be
explained by a high reactivity of these molecules. As for
benzoic acid, it cannot be identified with certainty, as its
characteristic fragments are common with those of the
previous molecules. On the other hand, 2-methylmaleic
anhydride, 2-benzofuran-1,3-dione (phtalic anhydride),
methyldiphenylmethane and dihydroanthracene were
identified. These molecules are characteristics of the second
way of toluene oxidation, which comes from an oxidative
coupling of two molecules of toluene [19]. This results in the
formation of methyldiphenylmethane and diphenylethane.
Moreover, many compounds from this way are isomers of
formula or position. Thus, all these molecules display the
same fragmentation, which is also common with the
monoaromatics of the main way. This is why so few by-
products of this pathway can be identified. These observa-
tions, and the fact that benzene is the only by-product
observed bymicroGC, confirm that the predominantway for
the catalytic oxidation of toluene is the first way described
by Lars and Andersson [19], shown in Fig. 6.
benzene. The formation of hydrogen can be explained by
the oxidation of benzyl alcohol in benzaldehyde. On the
other hand, a response was observed concerning the
fragments m/z 43, 57, 71 and 85. These are characteristic of
C3–C6 paraffins. Their presence can be explained by the
opening and catalytic cracking of the aromatic ring. Finally,
a response was observed concerning the fragments m/z
113 and 114, which are characteristic of C18H12 PAHs, such
as naphthacene, chrysene, benzanthracene or benzo(c)-
phenanthrene.
Nevertheless, it appears that these compounds, which
are more reactive than benzene and emitted in smaller
amounts, are totally eliminated before benzene. Thus, only
benzene appears to be a limiting factor to the catalysts’
performance.
3.3. Toxicological results
With the aim of evaluating the toxicological impact of
the by-products, a coupling of the catalytic process with a
cell exposure system is performed. Regarding the catalytic
results in terms of activity and formation of by-products,
the investigation of these toxicological effects using Pd/
a-
Al2O3 catalyst has been carried out.
In Fig. 7 are reported the profiles of catalyst tempera-
ture, toluene conversion and benzene production versus
time. After achieving toluene total conversion (T100) at
230 8C, the catalyst’s temperature decreases slightly to
stabilize at 227 8C. Consequently, the amount of benzene
emitted becomes relatively constant, with a value between
4 and 5 ppm during cell exposure.
Mass spectrometry has also allowed the identification
of other molecules. On the one hand, methane and the
hydrogen could be identified. The first may be explained by
the cracking of the methyl function of toluene, leading to
Fig. 6. Main way of toluene oxidation proposed by Lars and Andersson [19].
Please cite this article in press as: Brunet J, et al. Identification of by-products issued from the catalytic oxidation of
toluene by chemical and biological methods. C. R. Chimie (2015), http://dx.doi.org/10.1016/j.crci.2015.09.001

G Model
CRAS2C-4137; No. of Pages 10
J. Brunet et al. / C. R. Chimie xxx (2015) xxx–xxx
9
Fig. 7. (Colour online.) Temperature of the catalyst and benzene emissions as a function of time during the exposition with the Vitrocell1 system for the Pd/
a-Al2O3 catalyst.
The study of cytotoxicity did not show any significant
increase of the extracellular released LDH level after
exposure, despite the presence of carbon monoxide,
known for its toxicity. This absence of important cellular
death allows us to consider the early underlying physio-
pathological mechanisms of action occurring before
phenotypical cell damages. The mRNA level of the six
genes coding for XMEs was checked by qPCR. Their
expression was studied in cells exposed to:
When compared to control cells, five genes were
significantly upregulated (CYP2S1, CYP1B1, CYP1A1, NQO1
and EPHX1) for the highest dose of exposure (exhaust gas
without dilution), while three were significantly upregu-
lated (CYP1B1, NQO1 and EPHX1) for the catalytic exhaust
with a 10% dilution in purified air (Fig. 8).
Firstly, NQO1 and EPHX1 can participate in benzene
metabolization. On the one hand, this could be linked to
the presence of benzene when the toluene total conversion
is achieved and under static conditions. On the other hand,
even with a dilution effect when the effluents are out of the
process (10% in air), the results indicate a toxic response to
benzene.
ꢄ
ꢄ
gas from the catalytic exhaust without dilution;
gas from the catalytic exhaust with a 10% dilution in
purified air;
ꢄ
purified air (control).
Secondly, CYP2S1, CYP1B1 and CYP1A1 are mediated by
the aryl hydrocarbon receptor (AhR) [24]. This receptor is a
ligand-activated transcription factor involved in the
regulation of biological responses to planar aromatic
hydrocarbons [25]. This receptor, implied in the regulation
of XMEs, such as cytochrome P450, has been shown to be
induced by PAHs [26,27]. Consequently, the increase in
XMEs gene expression confirms the presence of PAHs in
the catalytic exhausts in these conditions. However, this
does not exclude the presence of other heavier PAHs than
C18H12, which has not been detected by chemical methods.
In addition to the necessary compromise to find
between performance and cost, toxicological studies show
that it is also fundamental to consider the health impacts of
catalytic processing units. This increases the relevance of
performing a toxicological validation of these systems.
4. Conclusion
Fig. 8. (Colour online.) Gene Expression of xenobiotic metabolizing
enzymes in A549 cells exposed for 1 h to purified air (control), or to 10% or
100% of the exhaust gases from the catalytic degradation of toluene by Pd/
This work deals with the identification of the by-
products of the catalytic oxidation of toluene. Firstly,
microGC analysis coupled with a mass spectrometer
a-Al2O3 (RQ: relative quantification of the XME mRNA in exposed cells vs
unexposed cells).
Please cite this article in press as: Brunet J, et al. Identification of by-products issued from the catalytic oxidation of
toluene by chemical and biological methods. C. R. Chimie (2015), http://dx.doi.org/10.1016/j.crci.2015.09.001

G Model
CRAS2C-4137; No. of Pages 10
10
J. Brunet et al. / C. R. Chimie xxx (2015) xxx–xxx
allowed detection and identification of many organic
compounds resulting from toluene oxidation. However,
the main by-product of toluene oxidation is benzene,
which can be explained by a low chemical reactivity of this
aromatic compound. A typical profile of benzene emission
has been described for each one of the catalysts studied,
despite the persistence and the temperature range.
Moreover, benzene seems to be a factor limiting the
performance of catalysts, because it is not completely
eliminated when the toluene is totally oxidized (T100).
Thus, it is necessary to operate the process at a working
temperature higher than T100 in order to obtain a total
oxidation of toluene and benzene. Therefore, for an
industrial processing unit, it is necessary to select a
catalyst that is efficient for the oxidation of VOCs, but also
for the oxidation of their by-products, especially when
these by-products exhibit toxic properties.
for the funding of the work and the ‘‘Agence universitaire
de la Francophonie’’ for Al Zallouha’s master fellowship.
The authors also acknowledge the ‘‘Centre commun de
´
ˆ
Mesures’’ of the ‘‘Universite du Littoral Cote d’Opale’’.
References
[1] K. Everaert, J. Baeyens, J. Hazard. Mater. 109 (2004) 113.
[2] M. Hosseini, S. Siffert, HꢀL. Tidahy, R. Cousin, J.-F. Lamonier, A. Aboukais,
A. Vantomme, M. Roussel, B.-L. Su, Catal. Today 122 (2007) 391.
[3] J. Tsou, L. Pinard, P. Magnoux, JꢀL. Figueiredo, M. Guisnet, Appl. Catal. B
Environ. 46 (2003) 371.
[4] T. Barakat, J. Rooke, HꢀL. Tidahy, M. Hosseini, R. Cousin, J.-F. Lamonier,
J.-M. Giraudon, G. De Weireld, B.-L. Su, S. Siffert, ChemSusChem 4
(2011) 1420.
`
[5] S. Scire, Appl. Catal. B Environ. 45 (2003) 117.
[6] J. Li, W. Li, J. Rare Earths 28 (2010) 547.
[7] P. Papaefthimiou, T. Ioannides, X. Verykios, Appl. Catal. B Environ. 13
(1997) 175.
[8] N. Burgos, M. Paulis, M. Mirari Antxustegi, M. Montes, Appl. Catal. B
Environ. 38 (2002) 251.
Secondly, the oxidation of toluene was investigated
under static conditions in order to expose human lung
cells. On the one hand, the results show that the amount of
benzene emitted remains relatively stable, between 4 and
5 ppm. On the other hand, the cells show a toxic response
partly due to benzene, even when a dilution is performed.
This one simulates a quite realistic exhaust dilution in the
atmosphere at the output of the treatment process.
This study confirmed that some by-products might be
emitted by the catalytic process under certain operating
conditions. It is necessary to take into account these by-
products emissions in the evaluation of the catalytic
performances. Thus, in the case of applications to real
industrial effluents, it is necessary to make compromises
between performances, operating costs, and potential
health impacts. Moreover, this increases the relevance of
performing a toxicological validation of these systems.
[9] S. Ordo´n˜ez, L. Bello, H. Sastre, R.F. Rosal, V. Diez, Appl. Catal. B Environ.
38 (2002) 139.
[10] S.C. Kim, W.G. Shim, Appl. Catal. B Environ. 92 (2009) 429.
[11] E.M. Cordi, JꢀL. Falconer, J. Catal. 162 (1996) 104.
[12] G. Centi, J. Mol. Catal. A Chem. 173 (2001) 287.
[13] J. Gonza´lez-Velasco, Appl. Catal. B Environ. 19 (1998) 189.
[14] P. Papaefthimiou, T. Ioannides, X. Verykios, Appl. Therm. Eng. 18 (1998)
1005.
[15] A. Rusu, E. Dumitriu, Environ. Eng. Manag. J. 2 (2003) 273.
[16] T. Machold, W.Y. Suprun, H. Papp, J. Mol. Catal. A Chem. 280 (2008) 122.
[17] G. Arzamendi, V. Delapenaoshea, M. Alvarezgalvan, J. Fierro, P. Arias, L.
Gandia, J. Catal. 261 (2009) 50.
[18] H.-J. Sedjame, G. Lafaye, J. Barbier, Appl. Catal. B Environ. 132–133
(2013) 132.
[19] S. Lars, T. Andersson, J. Catal. 98 (1986) 138.
[20] R. Beauchet, P. Magnoux, J. Mijoin, Catal. Today 124 (2007) 118.
[21] C. Gennequin, S. Kouassi, L. Tidahy, R.-F. Cousin, J. Lamonier, G. Garcon,
P. Shirali, F. Cazier, A. Abouka¨ıs, S. Siffert, C.R. Chimie 13 (2010) 494.
[22] S. Billet, G. Garcon, Z. Dagher, A. Verdin, F. Ledoux, F. Cazier, D. Courcot,
A. Aboukais, P. Shirali, Environ. Res. 105 (2007) 212.
[23] S. Billet, V. Paget, G. Garcon, N. Heutte, V. Andre, P. Shirali, F. Sichel,
Arch. Toxicol. 84 (2010) 99.
[24] S.T. Saarikoski, S.P. Rivera, O. Hankinson, K. Husgafvel-Pursiainen,
Toxicol. Appl. Pharmacol. 207 (2007) 62.
[25] M. N’Diaye, EꢀL. Ferrec, D. Lagadic-Gossman, S. Corre, D. Gilot, V.
Lecureur, P. Monteiro, C. Rauch, M.-D. Galibert, O. Fardel, J. Biol. Chem.
281 (2006) 19906.
[26] C. Sandoz, P. Lesca, A. Carpy, M. Laguionie, J.F. Narbonne, Int. J. Mol.
Med. 2 (1998) 615.
[27] J.F. Greene, J. Zheng, D.F. Grant, B.D. Hammock, Toxicol. Sci. 53 (2000)
352.
Acknowledgements
The authors want to acknowledge the French Environ-
ment and Energy Management Agency (ADEME) for the
financial support of the CORTEA PROBTEX project
(1281 C0095), as well as the Nord-Pas-de-Calais Region
Please cite this article in press as: Brunet J, et al. Identification of by-products issued from the catalytic oxidation of
toluene by chemical and biological methods. C. R. Chimie (2015), http://dx.doi.org/10.1016/j.crci.2015.09.001