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Journal of Environmental Chemical Engineering 2 (2014) 1104–1122

Contents lists available at ScienceDirect

Journal of Environmental Chemical Engineering

journal homepage: www.elsevier.com/locate/jece

Aerobic biodegradation of BTEX: Progresses and Prospects

Muftah H. El-Naas* , Janice A. Acio, Ayat E. El Telib
Chemical & Petroleum Engineering Department, United Arab Emirates University, P.O. Box 15551, Al Ain, United Arab Emirates

a r t i c l e i n f o a b s t r a c t

Article history: Monoaromatic pollutants such as benzene, toluene, ethyl benzene and xylenes are the most commonly cited
Received 22 January 2014 environmental contaminants in recent years and have attracted the attention of numerous researchers as well
Accepted 14 April 2014 as environmental agencies. Recently, considerable amount of research has been devoted to the development
of effective and reliable approaches for the containment of these toxic substances. Biotechnology has proven
Keywords:
to be a cost-effective and highly efficient method to remove petroleum and petroleum related pollutants
BTEX
Aerobic such as BTEX. This article offers a comprehensive review of old and recent literature dealing with the aerobic
Biodegradation biodegradation of BTEX. Special attention is given to the conditions that influence the overall degradation
Bioreactors efficiency and a discussion on recent research development such as innovative approaches, reactors and
Kinetics microorganisms. In addition, important aspects of BTEX biodegradation such as kinetics, mechanisms and
Modeling mathematical modeling are discussed in detail to develop a better understanding of BTEX as an environmental
challenge and compare the available options for tackling such a challenge.
c 2014 Elsevier Ltd. All rights reserved.

Introduction salt); the energy source for contaminant decomposition is provided

Volatile organic compounds (VOCs) particularly benzene, toluene,
ethyl benzene and xylenes (BTEX) have been considered as major
contributors to the deterioration of water and air quality. BTEX are
prevalent in the environment due to the fact that they are among the
anthropogenic emission of combustion processes as well as vehicle
exhausts. They are also used as industrial solvents for the synthesis
of several organic compounds (e.g. plastics, synthetic fibers, and pes-
ticides) and are the major aromatic components in many petroleum
products. Taking into account the very harmful effects of VOCs on the
environment and on living organisms, governments in many parts
of the world have been implementing very stringent environmental
standards. Consequently, there is an urgent need for the develop-
ment of efficient methodologies that are able to minimize or elimi-
nate the harmful effect of these compounds. Conventional treatment
techniques such as absorption, adsorption, combustion and conden-
sation suffer from several drawbacks, including high capital, operat-
ing and maintenance costs, high energy input, difficulty in handling
low-concentration pollutants, and production of toxic byproducts.
Recently, biological treatment processes that use the natural capa-
bility of microorganisms to degrade pollutants to less harmful prod-
ucts and utilize the carbon contained in these toxic compounds are
believed to be an attractive alternative. The numerous advantages of
biological methods include direct degradation, thus preventing the
increase in contamination of the environment; reduction of the pol-
lutants into less harmful reaction products (biomass, CO2 , H2 O and
* Corresponding author.
E-mail address: Muftah@uaeu.ac.ae (M.H. El-Naas).
by the contaminant themselves; and investment and operating costs
are low compared with other technologies. These can also be very
effective for treating contaminants with high flow rates and low pol-
lutant concentrations.
BTEX
The monoaromatic hydrocarbons, abbreviated BTEX, which stands
for benzene, toluene, ethyl benzene and the three xylene isomers,
are groundwater, soil and air pollutants, commonly associated with
petroleum and petrochemical production. BTEX are volatile, mono-
cyclic aromatic compounds that are usually present in coal tar,
petroleum products, and various organic chemical product formu-
lations (see Table 1 for physical and chemical properties of BTEX)
[1]. They are often found in air emission of several sources such as
refiners, petrochemical units, chemical plants, storage tanks, vehi-
cle exhaust [2], waste incinerators and composting facilities [3]. BTEX
contamination of soil and groundwater is usually related to petroleum
leakages and fuel oil from underground storage tanks, manufactur-
ing of solvent-based paints, lacquers and varnishes and the activities
of manufactured gas plants [4]. Significant quantities of these con-
taminants inevitably enter the environment during the production
process. BTEX compounds represent as high as 80% of the total VOC
in petrochemical plants [5] and account for up to 59% (w/w) of gaso-
line pollutants [6]. Aromatic compounds are widely distributed in the
environment due to natural and synthetic processes. Nonetheless,
substances produced through human activities are of greater concern
due to their toxicity and recalcitrance [4].
VOCs constitute a significant portion of hazardous waste being

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M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122 1105

Table 1
Physical and chemical properties of BTEX.

Compound Benzene Toluene Ethylbenzene o-Xylene m-Xylene p-Xylene

Molecular formula C6 H6 C7 H 8 C8 H10 C8 H10 C8 H10 C8 H10

Chemical structurea
MWb (g/mole) 78.11 92.13 106.16 106.16 106.16 106.16
BPc (◦ C) 80.10 110.60 136.20 144.40 139.30 137.00
MPd (◦ C) 5.50 −95.00 −94.97 −25.00 −47.40 13.00
VPe (mm Hg) 95.19 28.40 4.53 6.60 8.30 3.15
Densityf (g/mL) 0.87 0.87 0.87 0.88 0.87 0.86
Solubilityg (mg/L) 1791.00 535.00 161.00 175.00 146.00 156.00
Henry’s law 0.557 0.660 0.843 0.551 0.730 0.690
constanth (kPa
m3 /mol)
Overall reaction C6 H6 + 7.5O2 → C7 H8 + 9O2 →
6CO2 + 3H2 O 7CO2 + 4H2 O C8 H10 + 10.5O2 → 8CO2 + 5H2 O
b
Molecular weight [18].
c
Boiling point [19].
d
Melting point [19].
e
Vapor pressure [20].
f
Density [19].
g
Solubility at 25 ◦ C [18,20].
h
Henry’s law constant at 25 ◦ C [21].

treated globally today. They are harmful to ecosystem, human health

and atmosphere [7] and are among the major air pollutants due to
their malodorous and hazardous properties [8]. They readily volatize
to atmosphere and distribute over large regions because of their rel-
atively high vapor pressure. Their emission in the atmosphere causes
different environmental problems such as ground level ozone forma-
tion, stratospheric ozone depletion, photo chemical reactions, green-
house effect [9] and global warming [8]. VOCs emissions are heavily
regulated by federal, state, and regional air quality agencies [5]. This
group is included as regulated hazardous air pollutants in the US
Clean Air Act Amendments proposed in the 1990 [10]. Aromatic com-
pounds such as benzene are classified as hazardous air pollutants and
are limited to 25 tons/year total aromatics and 10 tons/year of any
individual aromatic [11].
Since these toxic substances easily move in air, they have direct
and indirect impacts on human health. Short term (acute) hazards
of BTEX include potential acute toxicity to aquatic life in the water
column (especially in relatively confined areas) as well as potential
inhalation hazards. Long term (chronic) potential hazards of these
compounds include changes in the liver and harmful effects on the Fig. 1. Generalized aerobic BTEX biodegradation pathway [29].
kidneys, heart, lungs, and nervous system [12]. Human exposure to
these compounds as a mixture can lead to neurological, respiratory,
genetic and excretory system damage and other health problems
Mechanism of BTEX biodegradation
ranging from irritation of the eyes, mucous membranes and skin,
to weakened nervous systems, reduced bone marrow function and
BTEX are highly receptive to microbial attack and the degradation
cancers. Usage of BTEX has persisted despite all these adverse effect
mostly occurs under aerobic conditions [34]. Toluene had been iden-
because of the extent of applications.
tified as the most easily biodegradable among the six compounds.
BTEX can pose a serious risk to soil and groundwater [13] because
This may possibly be due to the presence of the substituent group on
of their toxicity effect and high water solubility [14,15]. Due to their
the ring that offers an alternative route of attack on the side chain or
serious adverse impact on human health, the United States Environ-
oxidize the aromatic ring. The process requires dissolved oxygen (DO)
mental Protection Agency [16] has established maximum allowable
to utilize for both ring activation and cleavage of the aromatic nucleus
levels of these contaminants in water for public consumption. Since
and as the electron acceptor for its complete degradation by bacteria,
the frequency of groundwater contamination with hydrocarbons, in-
fungi or algae [22,23]. The overall reactions for BTEX biodegradation
cluding BTEX, has been increasing, there has been a demand for the
stoichiometries in aerobic conditions are given in Table 1. An aromatic
development of more efficient methods to remove or minimize the
compound can only be considered completely biodegraded if the ring
damages caused by these compounds [17].
undergoes cleavage (see Fig. 1) [24].
Metabolic pathways for the degradation of BTEX are provided
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1106 M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122
by two enzymatic systems: dioxygenases and monooxygenases. The
monooxygenase, also referred to as “tol” pathway, attacks methyl
or ethyl substituents of the aromatic ring [25,26] which are subse-
quently transformed by several oxidations to corresponding substi-
tuted pyrocatechols or phenyl glyoxal, respectively. The dioxygenase,
also referred to as the “tod” pathway, attacks aromatic ring with for-
mation of 2-hydroxy-substituted compounds [25–27]. The first step
of benzene oxidation is a hydroxylation catalyzed by a dioxygenase.
The presence of a substituent group onto the benzene ring allows
for two possible mechanisms: attacking side chains or oxidizing the
aromatic ring [28]. All these pathways converge in the formation of
(substituted) catechol intermediates [21,23].
For benzene, the main intermediate product is catechol [25,29,30]
while toluene and ethylbenzene are degraded on a separate pathways
leading to the production of their respective main intermediates, 3-
methylcatechol and 3-ethylcatechol [27,30]. Xylenes are all metabo-
lized to mono-methylated catechols; for example, m-xylene degrades
to 3-methylcatechol [31]. In contrast, some reports showed p-xylene
leads to 3,6 dimethylcatechol [32].
Subsequently, these catechol intermediates are mineralized by ei-
ther enzyme catechol 1,2-dioxygenase (also termed ortho- or intra-
diol cleavage or “upper” pathway) and then via the majority of the
pathways by β-ketoadipate or enzyme catechol 2,3-dioxygenase (also
termed meta- or extadiol-cleavage or “lower” pathway) [23,27,33].
The ring is opened and then degraded [34,35]. Finally, producing
low molecular weight compounds such as pyruvate and acetalde-
hyde, which can be further oxidized via the Krebs cycle [36]. Enzymes
catalyzing key steps in a catabolic pathway, such as catechol 1,2-
dioxygenase (C120) and catechol 2,3-dioxygenase (C230), could be
used for detecting BTEX [4,21,37].
Tsao et al. [25] reported that enriched soil culture use the tod
pathway only to metabolize benzene, while toluene and xylenes
may be oxidized by either the tod or tol pathway. However, p-
xylene can only be biodegraded via tod pathway, producing 3,6-
dimethylcatechol as an intermediate. Similar transformation was ob-
served in the metabolism of p-xylene and o-xylene by Pseudomonas
putida PPO1. Deeb et al. [38] conducted studies on BTEX mineraliza-
tion by the two Rhodococcus strains and showed that the pathway in-
volved is via a TOD-like involving dioxygenase attack on the aromatic
ring, leading to the formation of the corresponding catechols that are
then cleaved by either catechol-1,2- or 2,3-dioxygenase. Based on the
studied done by Mazzeo et al. [17] P. putida was able to break down all
the BTEX components and took the metabolic pathway based on the
direct oxidation of the aromatic ring by means of mono-oxygenases
or di-oxygenases to form a catechol, which is subsequently broken by
2,3-dioxygenase, and the metabolites generated in this second stage
are consumed by the Krebs cycle.
Zhang et al. [30] used a new strain Mycobacterium cosmeticum
byf-4 to simultaneously degrade (BTE(o-)X). This organism efficiently
degraded all the BTE(o-)X components either individually or as a com-
posite mixture and showed preference for toluene followed by ben-
zene, ethylbenzene and then o-xylene. In their study, isolation of
metabolites suggested that the BTE(o-)X compounds were initially
converted by a dioxygenase to their respective catechols. Thus, they
proposed that the initial attack on BTEX compounds by these bac-
teria appeared to be a dioxygenation reaction rather than sequen-
tial monooxygenations. Other strains that involve a dioxygenase en-
zyme system have also been reported by several authors. In another
study of BTEX biodegradation by P. putida ppF1, it was shown that
the strain utilizes a dioxygenase attack that oxidizes benzene ring to
form 3-methylcatechol which is then degraded via the meta-cleavage
pathway [39]. Whereas, Rhodococcus sp. strain DK17 has an o-xylene
dioxygenase that is involved in metabolism of o-xylene, toluene, and
ethylbenzene [40].
It is worth noting that bacteria and fungi degrade aromatic hydro-
carbons in different ways. Bacteria are able to utilize the compounds
as a sole source of carbon and energy, whereas fungi appear to come-
tabolize aromatic hydrocarbons to hydroxylated products [36]. Ligni-
nolytic fungi convert oxygen to hydrogen peroxide which is then used
for the formation of an aryl cation radical undergoing spontaneous re-
arrangements and degradation [41].
Factors affecting biodegradation of BTEX
Aerobic degradation can be affected by many physical, chemical
and biological conditions that influence the overall pollutant degrada-
tion efficiency. Several factors, such as pollutant concentration, tem-
perature, pH, availability of inorganic nutrients and microbial adap-
tation influence the rate and extent of biodegradation of BTEX [42].
Pollutant concentration and interaction
It is particularly important to study substrate interaction at dif-
ferent concentration since substrate toxicity is experienced by the
cells, especially at high concentration. Substrate inhibition due to
critical concentration is also said to be cell strain dependent. Li et al.
[43] showed inhibition by Planococcus sp. strain ZD22 for benzene
>80 mg/L. They confirmed that an inhibitory effect with increas-
ing benzene concentration can be obtained. Similarly, Abu Hamed
et al. [44] stated that specific growth rate of P. putida in batch sys-
tems has been found to be a decreasing function of concentration.
They reported that P. putida F1 could not degrade benzene completely
>380 mg/L and toluene >420 mg/L. BTEX compounds upon reaching
certain concentrations can inhibit the microbial activity due to com-
plex micro- and macro-level interactions [45]. Mathur and Majumder
[46] also reported that at high initial concentrations (>150 mg/L
benzene and 200 mg/L toluene), there was relatively less degrada-
tion rate of the substrates.
Substrate interactions can alter degradation rates of individual
contaminants either synergistically or antagonistically [47,48]. Syn-
ergistic interactions improve the degradation rates of individual con-
taminants by inducing the required catabolic enzyme. Another benefi-
cial substrate interaction which could enhance cometabolism would
be α BTX compound acting as a primary growth substrate. On the
other hand, antagonistic interactions inhibit the degradation rates
of another through exerting toxicity, diauxy, catabolite repression,
competitive inhibition for enzymes, or depletion of electron accep-
tors [49]. Antagonistic interactions such as preferential degradation
or diauxie, which is utilized in order of preferred substrates, can result
to lag phases before other substrates are consumed [50].
Abu hamed et al. [44] studied the biodegradation of benzene,
toluene and phenol as binary and tertiary mixtures. They discover
that the presence of benzene and phenol as co-substrate did not sig-
nificantly affect the biodegradation of toluene, but toluene and phe-
nol affected the biodegradation of benzene negatively. Jo et al. [45]
evaluated the antagonistic and synergistic effect of BTEX as mixed
substrate and demonstrated that increasing the concentrations of xy-
lene in the mixture showed good synergistic effect on the removal
of other compounds; however, the opposite occurred when benzene
concentration were increased. Toluene and ethylbenzene showed a
mixed response in the total BTEX removal pattern. Deeb et al. [38]
conducted studies with enriched consortium and R. rhodochrous and
evaluated the potential substrate interactions caused by the presence
of multiple BTEX compounds revealed a range of substrate interac-
tion. It was reported that benzene and toluene degradation rates were
slightly enhanced by the presence of o-xylene; whereas the presence
of toluene, benzene, or ethylbenzene inhibited the degradation of xy-
lene. Ethylbenzene was shown to be an inhibitor of BTEX degradation;
whereas, BTX was found to have negligible effect on the biodegrada-
tion of ethylbenzene by both cultures. At high levels of toluene, the
rest of the components in the mixture were inhibited. In a similar
study, Guo et al. [51] reported that in a binary mixtures of BT, BE and
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M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122
BX, T and X were completely removed; while B and E were not com-
pletely biodegraded. The presence of T and X increased the biodegra-
dation of B, but it was inhibited by E. The decreased degradation of B
in BE, compared with the degradation of B alone, was attributed to an
increase in the toxicity of B in the presence of E.
Aside from substrate interactions, degradation can also be inhib-
ited by the presence of structurally dissimilar compounds as well [50].
According to Corseuil et al. [52], the presence of ethanol was found
to inhibit the production of the enzymes that are needed for starting
BTEX degradation. Ethanol can be degraded using constitutive en-
zymes and long term exposure to it reduces the need of the bacteria
to produce the needed enzymes for BTEX degradation. Understanding
these interactions would lead to the understanding of why a partic-
ular BTEX compound still persist in a contaminated site, while other
BTEX compounds are degraded.
Temperature
Physicochemical condition, specifically temperature is important
when studying degradation rates [53]. Temperature is responsible for
controlling the nature and extent of microbial metabolism in hydro-
carbons as well as diffusion rates, bioavailability and solubility. BTEX
degradation rate under conditions of different temperatures could
affect the growth of bacteria, inactivation of enzymes, biotransfor-
mation and substrate concentrations and deprotonation of bacteria
[38,54]. The solubility of BTEX in an aqueous state decreases as the
temperature increases [55]. Studies on microbial growth and degrada-
tion activity of a microbial consortium from a gasoline-contaminated
aquifer was conducted and revealed cell growth increased with tem-
perature from 7 to 35 ◦ C and decreased sharply at 36–40 ◦ C [56].
Deeb et al. [38] optimized the temperature for bacterial growth and
toluene degradation activity to be 35 ◦ C. Alagappan et al. [57] deter-
mined the influence of temperature on the growth rate and benzene,
toluene degradation by P. putida over a range of 15–35 ◦ C. The opti-
mum temperature obtained is 33 ◦ C for both substrates. Therefore, P.
putida was identified to fall within the range typical for mesophilic
microorganism [58].
Mohammad et al. [9] showed that it was possible to treat high
loads of BTEX compounds under mesophilic (ambient conditions, ap-
proximately 20 ◦ C) and thermophilic (50 ◦ C) conditions. They studied
the effect on the elimination capacity (EC) of the biofilter and BTEX
removal efficiency and reported that thermophilic biofilter showed
effectiveness in treating BTEX gases at high loads and specifically su-
perior benzene removal. They also observed, within the mesophilic
range (15–30 ◦ C), that even a small increase in the temperature im-
proves pollutant removal in biofilters and biotrickling filters. They
indicated that the reason could be the lack of adaptation of the in-
oculated and dominant microorganisms to high temperatures, and
at high temperatures, the tolerance to substrate toxicity is reduced.
Only few studies are available on thermophilic biotreatment of BTEX
removal. Yoon and Park [59] showed that the removal efficiency of
VOCs in a peat packed biofilter was optimum at 32 ◦ C and decreased
at 45 ◦ C. These agreed with the study of Lu et al. [60] that the BTEX re-
moval of a trickle-bed biofilter increased in the range of 15–30 ◦ C and
decreased between 30◦ and 50 ◦ C. VOC degradation was inhibited at
temperatures above 40 ◦ C and reached optimum in the temperature
range of 25–35 ◦ C. Leson and Winer [61] also showed that microbial
activity was optimal at 20–40 ◦ C. In most laboratories, research is
carried out under mesophilic conditions (15–30 ◦ C); see Table 2.
pH
Microorganisms used in biodegradation usually survived in a cer-
tain pH range. For biodegradation of BTEX mixture by P. putida, You
et al. [53] study revealed that bacterial activity was completely inhib-
ited at pH 5, 9 and 10 and the optimum is at pH 6–8. In the case of
1107
trickle-bed air biofilter, Lu et al. [62] showed that B, T and X removal
efficiencies increased as the pH of the nutrient feed increased in the
range of 5–8 and decreased for pH between 8 and 8.5. The removal
efficiencies of each compound were greater than 80% in the pH range
of 7.5–8. Most bacteria are neutrophils, so the optimum pH at which
the highest degree of BTEX biodegradation is achieved is 7.5 [63].
These findings are consistent with Leson et al. [61] who reported that
bacteria and actinomycete activity was optimal in a range typically
between 7 and 8. Carbon dioxide is evolved during the metabolic
reaction of aerobic microorganisms which tends to lower the pH of
the system. Thus, if the waste gas or its intermediate byproducts do
not provide sufficient buffering capacity, the addition of chemicals
may be necessary for pH control. In general, the optimal pH range
for biological treatment systems is roughly 6.5–8.0 [64]. Thus, BTEX
treatment should be tested to optimize the pH value.
On the other hand, the biodegradation performance of fungi
showed no adverse effect associated with low pH and several au-
thors reported a good biofilter performance at low pH. Mohammad
et al. [9] showed that there was no adverse effect on the performance
of the reactor associated with the drop of pH to 4. Aizpuru et al. [65]
studied the biofiltration of a mixture of volatile compounds using a
peat biofilter. It reached a maximum removal and observed that even
under acidic conditions (pH 3.5–4.5), this did not seem to inhibit
or disturb the microorganism growth. Kennes et al. [66] evaluated
the biofiltration of VOCs and showed that good elimination efficien-
cies are reached with pH values lower than 4.5 and comparable to
those obtained with neutral pH values. Yadav and Reddy [67] studied
the degradation of BTEX by a lignin-degrading basidiomycete, Phane-
rochaete chrysosporium, and showed that pH variations between 4.5
and 7, had little effect on the extent of the BTEX degradation.
Availability of inorganic nutrient
Microorganisms consume organic contaminants like BTEX to fulfill
their carbon and energy requirements for ensuring biological activ-
ity. However, for their correct development, they also need nutrients
in solid form or as aqueous solutions which they cannot synthesize
for themselves. The usual solutions contain macronutrients (KH2 PO4 ,
KNO3 (NH4 )2 SO4 , NH4 Cl, NH4 HCO3 , CaCl2 , MgSO4 , MnSO4 , FeSO4 ,
NaMoO4 ) and micronutrients (vitamins and metals) [63]. In biofiltra-
tion, inorganic media such as rock, activated carbon, plastic or foam
do not contain appropriate nutrients, hence the microorganisms re-
quire the necessary nutrients to be provided [68]. However, a typical
biofilter uses a compost-based filter material that will provide suf-
ficient inorganic nutrients for microorganisms. Mudliar et al. [69]
reported that long term utilization of compost-based beds may lead
to exhaustion of nutritive resources and therefore becomes a limiting
factor for long-term biofilter performance. Other researchers are also
in agreement that depending on the target pollutant and the source of
the filter material, the availability of specific nutrients might become
process limiting [61,70]. Cho et al. [71] reported the most significant
decrease in the elimination capacity of biofilter and BTEX biodegrad-
ability due to nutrient limitation in the system. Thus, in order to have
a stable biofilter performance, it is encouraged that nutrient be fed
regularly and provided in the form and quantities that will support
most favorable microbial activity.
Microbial adaptation and processes
Microbial adaptation has been widely studied, because prior adap-
tation history significantly affects the degradation pattern. It was
shown that adaptation of microbial communities to specific aro-
matic carbon sources enhanced their degradative performance. Yeom
et al. [72] reported that when adapted to benzene, Alcaligenes xy-
losoxidans Y234 degraded benzene, toluene, and m-xylene better
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Table 2
Examples of BTEX biodegradation studies and process conditions.

Removal Degradation
VOC Microorganism Type of reactor Concentration pH Temp. (◦ C) efficiency (%) time Ref.

BTEX Free and Batch (shake 10% (v/v) 7.5 37 100 benzene, 48 h [42]
mixed flask) benzene, 10% 80 toluene,100
bacterial (v/v) toluene, ethylbenzene,
strain, Bb5 2–5% (v/v) 70 xylene
ethylbenzene,
0.5% (v/v)
xylene
BTEX Free Janibacter Batch (shake 240 mg/L pH 7 optimum 25–35 45.5 BTEX 60 h [74]
sp. SB2 flask) BTEX optimum
BTEX Free Batch (shake 15 and 7.0 28–30 97 benzene, 93 50 h [45]
Pseudomonas flask) 75 mg/L BTEX toluene, 90
sp.,Yarroia sp., ethylbenzene,
Acinetobacter 98 xylene
sp., Corynebac-
terium sp.,
Sphingomonas
sp.
BTEX Bacterial Batch (shake 50 mg/L BTE, 7.2 ± 0.5 36 ± 2 ◦ C 99.8 60 h [75]
consortium; flask) 15 mg/L
Strain FMB08; m-,p-xylene,
P. putida F1; 20 mg/L
and Escherichia o-xylene
coli strain
DH5a
BTEX Immobilized Batch (shake 24.68 mg/L NA 25 ◦ C 97.8 benzene, 24 h [76]
Mycobacterium flask) benzene, 94.2 toluene,
sp. CHXY119 23.67 mg/L 84.7
Pseudomonas toluene, ethylbenzene,
sp. YATO411 21.97 mg/L 87.4 p-xylene
ethylbenzene
BTEX Free Batch 100 mg/L 7.2–7.4 28 C 82–100 BTEX 36–42 h [30]
Mycobacterium (shake flask) BTEX
cosmeticum
byf-4
BTX Free and Batch 15, 30, 60, 7 30 ◦ C 100 BT, 6–14 h [77]
immobilized P. (shake flask) 90 mg/L 60–80
Putida F1 B, T, o-Xylene o-Xylene
BTEX Immobilized Continuous 120 mg/L 7–8 28–30 ◦ C 67 benzene, [78]
Mycobacterium reactor packed BTEX 81–90 TEX
sp. (CHXY119) with oxygen-
and releasing
Pseudomonas immobilized
sp. (YATO411) cell bead
(ORICB)
BTEX Free Bacillus Continuous, 0.0970, 0.0978, 30 ± 2 ◦ C > 99.85% [79]
sphaericus Bench Scale 0.0971 and
corn cob-based 0.0968 mL/L
biofilter BTEX
column
B Free Continuous 24.8 g/L 6.8 ± 0.2 19–22 ◦ C 81% [80]
gram-positive laboratory-
Gram-negative scale
streptomyces biofiltration
column
BTEX Free Continuous 2.3–4.3 mg/L 6.4–7.2 22–25 ◦ C > 97% [81]
filamentous shallow,
bacteria (A-1, sparged
ATCC No. bioreactor
55581)
BTEX Free Fungus Batch (shake 30–60 mg/L 7.5 30 ◦ C 100% B, E 240–288 h [82]
Paecilomyces flask) 45% B,
variotii 45%m-,p-
CBS115145 xylene
30% o-xylene
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M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122
than non-adapted and toluene-adapted cells. The reason was dis-
cussed in terms of catechol 1,2-dioxygenase induction; when adapted
to benzene, the cells induced more catechol 1,2-dioxygenase than
those adapted to toluene, which led to rapid degradation of benzene,
toluene and m-xylene. Babaarslan et al. [73] utilized a mixed cul-
ture of microorganisms obtained from petroleum production wells.
They showed that toluene as a single component and ethyl benzene
in a multi-component were biodegraded the fastest by non-adapted
microorganisms. They also reported that toluene-adapted cells have
higher rate of removal for single component than the non-adapted
and benzene-adapted microorganisms, resulting in a faster overall
biodegradation rate.
Advances in biodegradation of BTEX
Immobilization
Cell immobilization has been demonstrated to offer obvious ad-
vantages over conventional biological systems using freely suspended
cells. Immobilization has several advantages such as increasing the
biodegradation rate through a higher cell loading [83], protecting mi-
croorganisms from harsh environmental conditions, allowing higher
biomass density, providing a greater opportunity for reuse and re-
covery, and reducing reactor volume [84,85]. The immobilization of
cells can be achieved by different methods: covalent coupling, cross-
linking, physical entrapment, and the natural process of adhesion.
Recently, one alternate option that has turned to be important is the
production of natural fiber-polymer materials, the main application
of which is the production of construction materials [77]. Tsai et al.
[86] isolated Pseudomonas sp. YATO411 from a bioreactor through en-
richment using methyl tert-butyl ether (MTBE) and BTEX mixture as
sole carbon sources. The microorganisms were then immobilized in
phosphorylated PVA alginate gel beads using the method described
by Chen and Lin [87] and An and Lo [88] with small modifications. El-
Naas et al. [89] studied the characteristics and viability of polyvinyl
alcohol (PVA) as a support material for biomass immobilization. PVA
gel pellets were prepared by iterative freezing-thawing method us-
ing different PVA compositions. The study revealed that the porous
structure and mechanical properties of the PVA depended heavily on
the cross-linking process and the PVA composition. Robledo-Ortı́z et
al. [77] investigated the BTX degradation by immobilized P. putida F1
of postconsumer agave-fiber/polymer foamed-composites (AFPFC)
and suspended cultures in a controlled conditions. Analyses showed
that P. putida F1 adhered onto the composite surface and developed
a biofilm.
The microbial processes for BTEX degradation employ free
[30,42,45,74,81] or immobilized cells. Microorganisms have been im-
mobilized on carrier materials like porous polypropylene pellets, Ca/
Na alginate, polyvinyl alcohol, agave-fiber/polymer foamed compos-
ites, PVA-alginate beads to enhance the viability of microorganisms
[76,77,86,90]. Tsai et al. [86] investigated the characteristics and ki-
netics of benzene and toluene biodegradation using Pseudomonas sp.
YATO411 immobilized with PVA-alginate beads and compared these
experiments with freely suspended cells. The results indicated the
benefits of using a cell-immobilized system to treat high concentra-
tion of toxic xenobiotics, i.e. benzene, as it can protect cells from
shocks due to high concentrations. A cell-suspended system can be
used to treat a relatively non-toxic xenobiotic, i.e. toluene, using
Pseudomonas sp. YATO411. As toluene is relatively non-toxic com-
pared to benzene, mass transfer resistance of toluene that occurred
in cell-immobilized beads becomes an important rate-limiting step,
especially as the toluene concentration increases. These experimental
results established that whole cell immobilization offer obvious ad-
vantages over conventional biological systems using freely suspended
cells, especially for recalcitrant compounds like BTEX.
1109
High BTEX concentration
High concentrations of toxic pollutants typically inhibit biodegra-
dation and often affect the structure of a microbial community in a
contaminated aquifer [91]. Also, it is difficult to evaluate the changes
in microbial communities by toxic pollutants using conventional mi-
crobial identification methods, such as plate counting, because of co-
existence of several species of indigenous microorganisms in aquifers
[92]. Xin et al. [76] carried out a study for applying bioaugmenta-
tion technology for situ remediation of the high concentration BTEX-
contaminated groundwater with approximately 100 mg/L in total
concentration. Bioaugmentation with Mycobacterium sp. CHXY119
and Pseudomonas sp. YATO411 immobilized in bead was used to re-
mediate BTEX-contaminated groundwater. The batch experiments re-
sults showed that the CHXY119 and YATO411completely biodegraded
each BTEX compound, and degradation rates achieved by the bioaug-
mented permeable reactive barrier (Bio-PRB) were 97.8% for benzene,
94.2% for toluene, 84.7% for ethylbenzene and 87.4% for p-xylene. Ac-
cording to a study by Lin et al. [78], BTEX concentration of 120 mg/
L obviously distorted the structure of the indigenous microbial com-
munity. However, at BTEX concentration of 120 mg/L, 67% of benzene
and 81–90% of TEX were removed using novel oxygen-releasing bead
(ORB) and oxygen-releasing immobilized cell bead (ORICB). ORICBs-
column rapidly degraded BTEX after a 2–5 day acclimation period.
Singh and Fulekar [93] developed a two-phase partitioning bioreac-
tor system to overcome the problem of adding substrate at too high
concentration, which inhibits or even kills the microorganism, by
adding substrate at too low rate causing the microbial cell to starve
and resulting in a sub optimal process performance.
Bacteria from contaminated sites
Bacterial biomasses are usually isolated from sludge and soil sam-
ples from local area such as oil refineries and wastewater treatment
plants. Biomasses that are indigenous to the contaminated sites are
more capable of dealing with local environmental conditions than
those that may be acquired from abroad. Pseudomonas group is one of
the biomasses that are known for their organics-degrading abilities.
Martino et al. [94] isolated two Pseudomonas strains from an oil refin-
ery wastewater using several hydrocarbons as sole carbon source and
to accumulate polyhydroxyalkanoates. Both strains were capable to
synthesize rhamnolipids as surfactant compounds. One of these iso-
lates, Pseudomonas sp. KA, was able to degrade benzene, toluene, and
xylene, and to tolerate them at high concentrations. In the study of
Morlett-Chávez [75], the efficiency of BTEX biodegradation by a con-
sortium acclimatized to unleaded gasoline and the bacterial strains
that isolated from it was evaluated. The isolates were recognized by
PCR-amplification of 16S rRNA genes, where the BTEX biodegradation
was confirmed by the identification of dioxygenase-related proteins.
The results indicated that the consortium degraded 95% of the total
BTEX, and Strain FMB08 was able to remove 90% of the total BTEX; its
16S rDNA analysis was similar to that of Pseudomonas. Jin et al. [95]
generated an enrichment culture to isolate a BTEX-degrading bac-
terium from contaminated sea-tidal flat using seawater containing
BTEX compounds. In the enriched microbial communities, a Janibac-
ter species was dominant during the enrichment. Data showed that
for the initial concentration of 240 mg/L BTEX in a slurry system con-
taining 3.0 × 107 cells/L, 45.5% BTEX removal was observed under
the optimum condition of NH4 Cl and NaH2 PO4 ; whereas 32.2% BTEX
removal was observed under the uncontrolled condition of NH4 Cl and
NaH2 PO4 .
Utilization of mixed cultures
Broad spectrum of microorganisms using mixed cultures maybe
necessary for the complete mineralization of BTEX. Previous studies
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1110 M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122
have reported complex interaction patterns during BTEX biodegra-
dation using mixed/pure cultures, despite similarities in chemical
and structural properties [32,45]. Oh and Bartha [96] described the
complementary metabolic properties of a two-member consortium,
P. putida PPO1 and P. putida ATCC 33015. PPO1 strain followed the
TOD pathway consuming toluene and benzene, while the ATCC33015
the TOL pathway consuming toluene and p-xylene only. However,
when the two strains were used together in equal quantities, all of the
three components were completely removed. Another report showed
similar results, wherein a co-culture of two P. putida isolates was un-
able to utilize o-xylene; however, when the two cultures were mixed
together, all the BTEX compounds were removed collectively with
an enhanced cometabolic removal of o-xylene [97]. Liu et al. [98]
evaluated a pure and mixed cultures on the degradation of benzene.
Bacterial strains of pure culture (L4, N3, and N6) were isolated from
oil sludge and identified as Bacillus spp. Investigations showed that
(L4 + N3) mixed cultures exhibited greater efficacy in degrading ben-
zene than any other mixture or single culture examined and achieved
total degradation. It is evident that in this case, a mixture of two
bacterial strains has improved the efficacy of benzene degradation.
In some cases, the benzene degradation rates actually declined over
single strain which is perhaps due to inter- and/or intra-species in-
teraction of the bacteria. Deeb and Alvarez-Cohen [99] explained the
mineralization potentials of mixed and pure cultures. Two isolates in
their study exhibited a biodegradation pattern that was a subset of
that exhibited by the mixed culture. This was attributed to either the
presence of different microbial species with a number of metabolic
pathways or to interspecies interactions. Therefore, mixed cultures
may be more effective than pure cultures in biotreatment systems for
the complete biodegradation of multicomponent hydrocarbon mix-
tures.
Microorganism in BTEX biodegradation
Biological processes have been used as effective, eco-friendly and
potentially cost savings approaches for treating VOCs. Microorgan-
isms are important in biological process. They have the physiologi-
cal and metabolic capabilities to be highly effective in the removal
of these pollutants. A large number of microorganisms including
bacteria, fungi and algae are capable of degrading BTEX. Microbial
degradation of BTEX by aerobic [97] and anaerobic bacteria [100–
103] has been extensively studied for two decades. However, this
review focuses on the aerobic biodegradation of these compounds
(Table 3). The degradation of BTEX was discovered when the bac-
teria Bacillus hexabovorum grew aerobically in a medium containing
toluene and xylene. Gray and Thronton in 1928 demonstrated the
ability of microorganism, found naturally in soil, to degrade BTEX.
According to Gibson and Subramanian [104] and Corseuil and Alvarez
[105], researchers found more than 200 species of bacteria present in
non-contaminated soil samples were able to degrade hydrocarbons.
Among the bacteria of the genus Pseudomonas, P. putida [57,106–109]
is the most common bacterial genus employed in degrading aromatic
hydrocarbon. P. putida is gram negative bacterium able to metabo-
lize BTEX and other aromatics as the only carbon and energy source
[17,110,111]. The strain demonstrates a diverse metabolism, and it is
non-pathogenic compared to other species.
BTEX degraders that have been isolated in different environ-
ments include Rhodococcus [112], Marinobacter [113] and Acine-
tobacter [114]. Other BTEX degraders detected in soil include Al-
caligenes, Arthrobacter, Acidovorax, Agrobacterium, Aquaspirillum, Bre-
vibacterium, Bradyrhizobium, Variovorax and Stenotrophomonas [115].
Common BTEX degraders from sewage and fresh water are Ralstonia
[116], Microbacterium, Mycobacterium, Azoarcus [117], Thauera [118]
Burkholderia [119] and Sphingomonas [120]. However, there are some
limited reports in degrading potential of Janibacter species though it
exhibited obvious BTEX-degradation ability. This bacterial strain, Jani-
bacter sp. SB2, was isolated from a contaminated sea-tidal flat through
an enrichment process. Strain SB2 was able to degrade all BTEX com-
pounds effectively but xylene compounds (o-, m- and p-xylene) were
degraded more slowly than other compounds and occurred almost
simultaneously [74].
Aside from bacteria, fungi have also shown the capability to
degrade hydrocarbons [121]. The ability to utilize hydrocarbons
has been observed in numerous types of fungi. Prenafeta-Boldu et
al. [122] isolated five fungal strains from enrichment culture. The
isolates were identified as deuteromycetes belonging to the gen-
era Cladophialophora, Exophiala and Leptodontium, the ascomycete
Pseudeurotium zonatum and the toluene-degrading fungus Cladospo-
rium sphaerospermum. The study showed the advantages possessed
by fungi over some bacteria is that the former can adapt more readily
to adverse environment (e.g. conditions of low moisture and low pH)
and when near zero net growth is preferred. Prenafeta-Boldu et al.
[123] have investigated the removal of BTEX compounds as mixtures
using fungi. The deuteromycete Cladophialophora sp. strain T1 was
able to degrade toluene and ethylbenzene but not benzene, while dif-
ferent amounts of the xylene isomers were co-metabolized. The lack
of benzene degradation appears to be the main drawback for applica-
tion of this fungus; however this strain possessed a metabolic capacity
for the degradation of BTEX similar in many aspects to that of bacteria.
A more recent study by Nikolova and Nenov [124] showed the poten-
tial of Cladophialophora sp. and Cladosporium sp. for BTEX degradation.
They reported that ethyl benzene was easily degraded in all cases, but
neither strain was able to degrade benzene. Cladophialophora sp. fully
degraded o- and m-xylene both as single substrates and in mixtures
with toluene, while Cladosporium sp. was able to degrade them fully
only in the presence of toluene. p-Xylene was only partially assimi-
lated in all tests.
Fungal ability to metabolize the individual BTEX compounds was
evaluated by Paecilomyces variotii CBS115145 in batch and in biofil-
tration experiments. Toluene was completely degraded, followed by
ethyl benzene; benzene was partially metabolized along with xylene
isomers. Binary mixtures were also used to determine the effects of
interactions in the degradation pattern. Degradation followed the or-
der of toluene, ethyl benzene, and benzene in binary toluene–benzene
and ethyl benzene–benzene mixtures. The overall reduction in rates
in both cases suggests competitive inhibition. When toluene was used
as initial substrate, m-xylene was completely assimilated; however,
benzene and o- and p-xylene were still partially degraded [82]. One
of the main advantages of preferring the growth of fungi rather than
bacteria for the removal of VOC is their ability to degrade these pol-
lutants under a wide range of process conditions [125]. With this
development, further studies have been performed to improve the
efficiency of these microorganisms for BTEX degradation [17] and has
led to the advancement of numerous biologically based technologies
for their control and treatment from wastewaters and, in particular,
waste gases.
Recent advances in kinetics and modeling
Kinetics and modeling
Significant amount of studies has been reported on the kinetics and
simulation of degradation of BTEX as individual component or mix-
ture. Accurate investigations of biodegradation kinetics are required
for improvement of biodegradation process which depends largely
upon consideration of these kinetic characteristics. Also, a good de-
termination of biodegradation kinetics is important to design and
optimize a cost effective biological reactors in order to treat contam-
inated groundwater, contaminated soil, or industrial wastewaters. A
variety of kinetic models have been used to describe the dynamics of
microbial growth on BTEX as shown in Table 4.
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M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122 1111

Table 3
Example of microorganisms and substrate.

Microorganism Substrate References

Bacteria
Rhodococcus rhodochrous BTEX [38]
Alcaligenes xylosoxidans Y234 Benzene, Toluene and Phenol [126]
Rhodococcus sp. DK17 BTE, phenol, isopropylbenzene, and other [112]
alkylbenzene isomers
Pseudomonas putida BTEX [97]
Pseudomonas putida BTE(o-)X [106]
Pseudomonas fluorescens BTE(o-)X [106]
Pseudomonas aeruginosa Benzene [127]
Pseudomonas putida F1 ATCC 700007 BT, phenol [108]
Pseudomonas putida F1 BT [57]
Burkholderia (Ralstonia) pickettii PKO1 BT [57]
Rhodococcus pyridinovorans PYJ-1 Benzene, toluene, and m-xylene [128]
Pseudomonas putida CCMI 852 BTX [109]
Achromobacter xylosoxidans BTEX [129]
Pseudomonas spp. BTX [130]
Pseudoxanthomonas spadix BD-a59 Benzene, toluene, ethylbenzene, o-, m-, p-xylene [131]
Alcaligenes (Achromobacter) xylosoxidas BTX [132]
BTEX [78]

Pseudomonas sp. (YATO411)

Mycobacterium sp. (CHXY119)

Pseudomonas putida YNS1 BTEX [53]

Bacillus sphaericus (MTCC 8103) BTEX [133]
Mycobacterium cosmeticum byf-4 BTE(o-)X [30]
Fungi
Phanerochaete chrysosporium BTEX [67]
Cladophialophora sp. T1 TEX [123]
TE (o-, m-, p) X [124]

Cladosporium sp.
Cladophialophora sp.

Paecilomyces variotii CBS115145 BTE (o-, m-, p) X [82]

Cladophialophora psammophila TEX [134]

Table 4
Biodegradation kinetics models.

Model name Equation References

μmax S
Monod μ= K s +S
[A] [135–137]

μmax S
Andrews μ= K s +S+S2 /K i
[B] [108,137,138]

μmax S
Andrews and Noack μ= ( K s + S)(1+ S [C] [139]
Ki )
n
μmax [1− KSi ]
Han-Levenspiel μ= K s + S−[1− S
]
m [D] [139]
Ki

μmax S S n
Luong μ= K s +S
[1 − Sm
] [E] [43,140,141]
μmax S
Yano and Koga μ= K s +S+S3 /K i 2
[F] [141–143]
μmax S (− KSi )
Aiba et al. μ= K s +S
e [G] [43,141,144]
μmax S
Wayman and Tsenga μ= K s +S
− i ( S − Sθ ) [H] [43,141,145]

μmax S
Alagappan and Cowan μ= K s +S+S2 /K i
− i( S − Sθ ) [I] [141,146,147]
μmax S
Michaelis–Menten: two substrate μ= S+ K s (1+ KI )
[J] [128,135,148,149]
i
reaction, competitive inhibition
μmax S
Two substrate, non-competitive μ= ( S+ K s )(1+ I
)
[K] [149]
Ki
inhibition
μmax Si
Mixture of substrate, competitive μ= K si + Si +

Sj ( K si
)
[L] [137,149–151]
j=i K sj
inhibition
μ max Si
Mixture of substrate, non-competitive μ=  K si Si S j [M] [137,149–151]
K si + Si + j=i [ S j ( K sj )+ K sj ]
inhibition
μmax Si
Mixture of substrate, uncompetitive μ=  Si S j [N] [149–151]
K si + Si + j=i K sj
inhibition
μmax
Si
SKIP, unspecific interaction μ= K si + Si +
[O] [107,149,151,152]
j=i Si I ij

a
Sθ = threshold m-xylene concentration below which there is no inhibition [mM].
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1112 M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122
Based on material balance, the growth is presumed to be pro-
portional to the size of the cell population and can be described as
follows:
dX
= μnet X = μ X − kd X
dt
d ln X
or = μnet X
dt
dS μX
= −
dt Y 20 mg/L, cell concentrations of about 106 –107 mL−1 , and for Ks half
where t, X, μ and μnet denote the time, concentration of biomass (mg/
L), specific growth rate (h−1 ) and the net specific growth rate (h−1 ),
respectively. S is the substrate concentration (mg/L) and Y is the
biomass yield coefficient must also be determined experimentally.
The Monod equation is the most widely used kinetic expression
to model liquids or gases biodegradation. Monod equation expresses
the microbial growth rate as a function of the nutrient that limits
growth [135]. The equation has a similar form to that of the Michaelis–
Menton equation for enzyme kinetics, except that Michaelis–Menton
equation was derived empirically [135]. The Monod model is given in
Table 4 as equation A, where μ is the specific growth rate (h−1 ), μmax
is the maximum specific growth rate (h−1 ), Si is the substrate con-
centration (mg/L), and Ks is the half saturation constant (mg/L). The
model is based on the assumption that only one limiting substrate can
be considered, and a variation of its concentration causes alterations
of the culture behavior. The yield (Y) is constant over the concentra-
tion range when the initial substrate concentration is much higher
than the critical substrate concentration (So ≫ (Ks ,KI 2 )1/3 ) [141]. The
Monod model has limiting condition that the substrate concentra-
tion should be sufficiently low that Ks ≫ S [137]. Whereas in the
cases of high pollutant concentrations Ks ≪ S, an inhibiting effect on
biodegradation will appear, and the Haldane Model (some time re-
ferred to Andrews) is recommended (Table 4, equation B), where Ki
is the inhibition constant (mg/L); high Ki value means the culture
is less sensitive to substrate inhibition and vice versa. The biodegra-
dation rate passes the maximum with increasing concentration. The
μ value is determined based on the exponential phase (first-order)
[135]. When the substrate concentration increases, this equation can
predict Monod behavior until the substrate concentration reaches a
maximum. The rate then decreases because of the S2 term in the
denominator.
Mathur et al. [139] used Han-Levenspiel (Table 4, Equation D)
which is used to observe that the growth ceases altogether at suffi-
ciently high concentrations of inhibitory substrates. The culture fol-
lowed substrate inhibition kinetics could be fitted to Haldane, An-
drews and Noack and Han-Levenspiel models. Among the three mod-
els, the Han-Levenspiel model is best suited system of the biodegra-
dation of Benzene, toluene and phenol, because as substrate concen-
tration increases the term (μmax /Ks + S − [1 − S/Ki ]m ) becomes
close to 1. Moreover, it has been reported that at high concentration,
the biodegradation rate decreases due to: (1) deficit in availability of
oxygen, and hence the culture cannot biodegrade benzene, toluene
and phenol under hypoxic conditions. (2) The fall in pH of the so-
lution inhibits cell growth at initial substrate concentration above
√
So ≫ KS Ki [153,154]. Another substrate-inhibition kinetic model is
Luong (Table 4, equation E) where Sm is the maximum substrate con-
centration above which growth ceases. Luong’s equation is the same
as Han-Levenspiel, except that m = 0. There are number of empirical
inhibition models, such as Aiba (Table 4, equation G), Wayman and
Tseng and Alagappan and Cowan (Table 4, equations H and I), that de-
scribe the substrate inhibition of microbial growth [147]. Alagappan
and Cowan [147] indicated that the Luong model best represented
substrate inhibition of specific growth rate for benzene, comparing
with the other four inhibition models mention above. He Li et al.
[43] selected Luong model among different known kinetic models to
describe substrate inhibition behavior for improving the prediction
of Pseudomonas putida with the use of a gene expression regulation
model of the TOL plasmid. Koutinas et al. [141] chose Yano and Koga
model from the models propagated. There were small differences be-
(1) tween the tested models: Andrews, Yano and Koga, and Wayman and
Tseng.
It has been reported in the Literature that for different ranges of
BTEX concentration, the Monod and Haldane (Andrews) models could
simulate BTEX degradation profiles (Table 5). The Monod model gives
good results for initial substrate concentration So in the range of 1–
(2)
saturation coefficient less than 1 mg/L. Ellis et al. [155] showed that
when So /Ks ratio is greater than 1 for experimental data fitted by the
Monod model, one unique maximum of specific growth rate μmax
and Ks values will be found. Jihyeon et al. [156] illustrated that the
half-saturation coefficient for toluene was low compared to values
in the Literature, because the half-saturation coefficient is strongly
dependent on the type of microorganisms and on the physiological
stages. Andrews model shows better performance than the Monod
model in batch operation, at a given value of Ki and when the initial
concentration of toxic substrates increases with Si2 / K i > 0 [149,157].
Trigueros et al. [137] confirmed that for concentrations above 40 mg/
L, Andrews model gives favorable fitting. In addition, in most of the
studies listed in Table 4, it was assumed that BTEX concentration is
the only limiting for growth rate; a sufficient amount of headspace
was provided, and the liquid phase was well mixed so that the oxygen
supply from the gas phase to the bulk liquid was not a growth limiting
[43,77,137,141,151].
Biodegradation kinetics for mixtures
Although microbial growth on substrate mixtures is commonly en-
countered in the biological treatment processes, the number of stud-
ies on the mathematical modeling of the biodegradation of mixed
substrates is still limited compared to single substrate biodegrada-
tion [141]. Deeb RA et al. [158], Chi-Wen et al. [149], Trigueros et
al. [137] and Yoon et al. [152] proposed that the performance of the
unstructured kinetic models for the mixture of BTEX as homologous
substrates can be represented by competitive inhibition and SKIP
models. Chi-Wen et al. [149] and Segel [159] used noncompetitive
and uncompetitive inhibition models to describe dual substrate in-
teraction. Moreover, Abuhamed et al. [160] and Reardon et al. [107]
investigated the kinetics of P. putida F1 growing on benzene, toluene,
phenol and their mixture and compared the different mathematical
models to describe the results.
The competitive inhibition model (Table 4, equation J) applies
when two or more compounds serve as substrates, and the com-
pounds can be degraded simultaneously with competition. While the
noncompetitive inhibition model (Table 4, equation K) describes the
processes where two or more substrates are simultaneously bound
to enzyme forming a nonreactive complex. The model of uncompeti-
tive inhibition, presented in (Table 4, equation N), can be used when
only the inhibiting substrate binds to the enzyme–substrate complex
not the free enzyme. When the interaction between the substrates
does not specify the type of interaction, then an alternative model
can be applied. The model is known as SKIP (sum kinetics with inter-
action parameters) (Table 4, equation O), and it formulated by Yoon
et al. [152]. This model includes an interaction parameter Ij,k in each
Monod term indicating the degree to which substrate j affects the
biodegradation of substrate k. The SKIP model accurately shows the
biomass dry cell weight concentration [141]. In a study by Daniela et
al. [137], SKIP model perfectly described biodegradation kinetics of
BTEX mixtures. However, the weakness of SKIP is modeling the sys-
tem on the population level without taking into account the specific
metabolic controlling key steps. Full understanding of the interac-
tions between substrates can be achieved by studying the metabolic
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M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122 1113

Table 5
Summary of growth kinetics from literature.

Concentrations Interaction
Microorganism range (mg/L) Model type Compounds μmax (h−1 ) Ks (mg/L) Ki (mg/L) parameters Temp. (◦ C) Ref.
P. putida 10–400 Monod B 0.1631 71.18 – – 30 [139]
T 0.1722 62.56 – –
P. putida F1 15–30–60–90 Monod B 0.5 10.11 – – 30 [77]
T 0.58 10.8 – –
P. putida F1 2–250 Andrews T 0.78 5 753 – 32 [138]
P. putida 10–400 Han- B 0.3626 – 64.12 – 30 [139]
Levenspiel
T 0.3003 – 57.40
Rhodococcus 25–30–50 competitive T-B 0.13 6.2 12.4 – 32 [128]
pyridinovo-
rans
T-m-X 0.12 6.2 17.2 –
B-T 0.080 4.8 6.7 –
m-X-T 0.035 4.7 9.6 –
P. putida F1 90 Andrews B 0.62 1.65 180 – 30 [108]
T 0.61 6.47 88
Pesudomonas 46–274 Andrews B 0.0194 8.349 191.89 – 30 [149]
aeruginosa
T 0.0064 9.851 48.48 –
E 0.0340 211.791 310.56 –
X 0.0060 1.427 153.55 –
Pseudomonas 0.9–13.6–1.04 Yano and m-X 0.979 0.096 0.465 – 30 [141]
putida mt-2 mM Koga
Planococcus 0–11 mM Luong B 0.34 0.041 – n = 1.21, Sm 20 [43]
sp. strain zd22 = 10.2 mM
Pesudomonas 46–274 Competitive M + B + T 0.0292 1087.795 – - 30 [149]
aeruginosa + E + X
Non- 0.0302 1088.291 – –
competivite
Rhodococcus 0–80 SKIP B 0.41 1.11 – Itb = 1, Ieb = 35 [137,158]
rhodochrous 10, Ixb =
0.007
T 0.42 1.24 – Ibt = 0.0023,
Iet = 4.5, Ixt =
0.0005
E 0.45 1.75 – Ibe = 0.175,
Ite = 0.025,
Ixe = 0.10
X 0.05 20 – Ibx = 1.10, Itx
= 1.7136, Iex
= 7.075
Consortium 80 SKIP, B 0.44 27.57 – IT,B = 2, IB,T = 30 [151]
cometabolism -0.4
T 0.60 34.12 –
E 0.13 0.36 – IE,B = 4, IX,B =
-0.7
o-X 0.85 0.85 –
P. putida DSM 39.5 ± 5.9 SKIP, B 0.88 0.3 – IT,B = 2.2, IX,B 30 [156]
921T unspecific = 0.3
interaction
T 1.91 0.5 – IB,T = 0.8, IX,T
= 0.9
p-X 0.1 – – IX,B = 0.7, IX,T
= 0.7
(cometabolism)
B: benzene; T: toluene; E: ethylbenzene; X: xylene; M = MTBE (methyl t-butyl ether).

pathways of microorganism [137]. Michalis et al. [141] divided the
limitation of models into three categories: “Category 1: the lag period
is not modeled and models are used to predict only the post-phase
data; Category 2: there is no comparison of model’s prediction against
an independent experiment; Category 3: the model predictions do not
fit accurately the experimental results and it is not possible to predict
a variety of multisubstrate experiments using a single set of parame-
ters”. However, Littlejohns et al. [151] and Reardon et al. [107] were
categorized in Category 1.
Temperature significance in kinetics
Temperature is the most important environmental parameter af-
fecting microbial growth and activity [161]. Relatively few studies
have been published in the Literature totally assessing the effect of
temperature on the kinetic and stoichiometric coefficients of aero-
bic microorganisms, such as maximum specific growth rate, specific
decay rate, growth yield and half-saturation coefficient [57]. Some
studies point out that optimal growth temperature reflecting the
temperature range which favors bacterial growth, lies between 20
and 40 ◦ C [162]. The well-known Arrhenius equation [57], as given in
Eq. (3), is commonly used to present the effect of temperature on the
maximum specific growth rate:
μˆ = Ae E a /RT (3)
where μ is the temperature-dependent maximum specific growth
rate (h−1 ), A is an exponential factor, Ea is the activation energy for
cellular multiplication (J/mol), R is the universal gas constant (J/mol
K), and T is the absolute temperature (K). This growth model implies
an exponential increase in the growth rate of the organisms with
rising temperature. However, the Arrhenius function is unsuccessful
when the temperature approaches the value of optimum activity,
because it cannot characterize the fall in rates at higher temperatures.
However, alternate model has been proposed by Topiwala and Sinclair
[57], as given in Eq. (4), to overcome the limitations of Arrhenius and
predict the drop in biodegradation rate following the optimum.
μˆ = Ae−E a /RT − Be−E b /RT (4)
where Eb is the activation energy for thermal denaturation processes,
which is usually higher than the activation energy for multiplication.
It is assumed that when the temperature rises above the optimum, the
cellular decomposition reactions are favored, resulting in irreversible
damage to plasma membranes, loss of metabolites, and decrease in
metabolic function.
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1114 M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122
Alagappan and Cowan [57] conducted batch experiments to an-
alyze the effect of temperature and dissolved oxygen concentration
on the rates of growth and benzene and toluene degradation by the
toluene-degrading strain, P. putida F1. For the temperature range of
15–35 ◦ C, the maximum specific growth rate followed the Topiwala–
Sinclair relationship when either benzene or toluene was considered
as the sole carbon source. Raikos et al. [163] evaluated the growth pro-
file of Staphylococcus epidermidis, R. picketii and Agrobacterium tume-
faciens based on cell counts and reported that the temperature had a
profound impact on the growth profile of all three bacterial species;
the highest increase in bacterial cells was observed at 30 ◦ C.
Response surface methodology (RSM)
The aim of RSM is to characterize the relationship between a re-
sponse and group of quantitative factors of interest to the researcher.
This is achieved by building a model that describes the response over
the applicable ranges of the factors of interest. The response surface
referred to as the fitted model in many industrial applications, be-
cause the response can then be graphed as a curve in one dimension
(one factor of interest) or a surface in two dimensions (two factors of
interest) [45,164]. By definition, response surface methodology (RSM)
is a graphical statistical approach to identify factor settings (operating
conditions) that produce the best response and satisfy operating or
process specifications. It can also be used to identify new conditions
that improve product quality over that achieved by current condi-
tions, while simultaneously modeling the relationship between the
quantitative factors and the response [165].
After running a full central composite design (CCD) experiments,
a second-degree quadratic polynomial can be used to represent the
function in the range of interest:
   
Y = β0 + βi X i + βii X i2 + βij X i X j
i=1 i=1 j=i+1
where Y is the predicted response, β 0 is the offset term, β i is the
coefficient of the linear effect, β ii is the coefficient of squared effect,
Xi is the coded value of variable i, Xj is the coded value of variable j,
and β ij is the coefficient of interaction effect.
Since the degradation of BTEX by microorganisms is strongly af-
fected by many parameters, it is important to search for the key in-
fluencing factors. Conventional techniques such as “one-factor-at-a-
time” do not guarantee the determination of optimum conditions and
are unable to detect synergistic interactions between two or more
factors. Thus, by using the response surface method (RSM), one can
optimize all the affecting parameters and eliminate the limitations of
single factor optimization [166]. RSM has been used in various fields
such as machining techniques [166], chemical reactions [167,168]
enzyme and catalytic reactions [169–171] optimization of water and
water treatment process [172–174], biochemistry [175] toxicology
studies [176–178] and biotechnology for studying the biodegrada-
tion of phenol [179]. However, only a few studies were reported on
optimizing the influencing factors and their interactions to improve
BTEX’s removal efficiency.
Recent advances in reactor types
Biological treatment is an increasingly popular technique com-
pared with physical and chemical methods, since it offers a cost ef-
fective option, and it is positively perceived by the general public as an
environmentally friendly alternative to conventional removal meth-
ods [180]. It is an alternative to conventional air pollution control
technologies such as thermal or catalytic oxidation, wet scrubbing,
and adsorption onto activated carbon. Biological treatment methods
do not only involve bacterial microorganisms, but other biomasses
such fungi and plants are being used [181]. Biological treatment is
achieved at ambient temperatures and does not generate secondary
pollutants; it converts volatile organic compounds to carbon diox-
ide, reduce sulfur compounds to sulfate, and chlorinated compounds
to CO2 and chloride. Several bioreactors have been developed for
treating volatile organic compounds and odorous compounds in va-
por phase. The different types of air phase biological reactors include
biofilters, biotrickling filters and bioscrubbers. Among the newly de-
veloped reactors are the membrane reactors, novel rotating rope
bioreactor, bioactive foam emulsion reactor, an flat plate vapor phase
bioreactor using oxygen micro sensors, two liquid phase bioreactor.
Some other examples are the external loop airlift bioreactor, fluidized
bed bioreactors, spouted bed bioreactor (SBBR) and monolith biore-
actor.
A summary of the main advantages and disadvantages of different
types of bioreactors is given in Table 6. Moreover, brief descriptions
of major bioreactors are given in following sections.
Biotrickling filters
In a trickle bed reactor, the liquid phase and nutrients are fed
at the top. The aqueous phase trickles from the top of the reactor,
over the biological substrate in order to maintain the maximum level
of nutrients and degree of wetness. The influent gas is allowed to
flow into the reactor, either with the current of the water (liquid
phase) or against it. The absorbed contaminant in the liquid are in
constant contact with the packing material and microorganisms on
it; therefore, they are rapidly biodegraded [63]. Raquel et al. [182]
showed that mass transfer characterization was a powerful tool to
optimize both biotrickling filters design and operation. The KLa values
were obtained by fitting the model to experimental data of toluene
absorption obtained at empty bed residence times (EBRT) from 7
to 50 s. The model resulting KLa values ranged from 35 to 113 h−1 .
Chungsying Lu et al. [62] studied the effects of pH, moisture and flow
pattern on the performance of a trickle-bed air biofilter. In the pH
range of 7.5–8, removal efficiencies of each compound were greater
than 80% with a loading of 143 m3 /h.
Biofilters
Biofilters are the most widely used bioreactors for air pollution
control, where a humid polluted air stream is passed through a porous
packed bed on which a mixed culture of pollutant-degrading organ-
isms is naturally immobilized. Garcı´a-Pen˜a et al. [181] evaluated the
BTEX degradation as individual substrates and in mixtures, in liquid
culture, using packed biofilters with the filamentous fungus P. variotii
CBS115145. The results illustrated that BTEX are differentially utilized
by P. variotii; toluene was completely degraded, followed by ethyl
benzene; benzene and m-p-xylenes were partially assimilated (45%),
whereas o-xylene was 30% metabolized in liquid culture. Moreover,
carbon recoveries as CO2 were 48, 40, and 53% for toluene, benzene,
and ethyl benzene, respectively.
Rahul et al. [133] evaluated the degradation of BTEX for a period
of 86 days in a laboratory scale corn-cob based biofilter containing
Bacillus sphaericus (MTCC 8103). Using a 3-D mesh technique, the
overall performance of a biofilter was estimated in terms of its elimi-
nation capacity. Maximum removal efficiency was found to be more
than 96% for all BTEX compounds. The optimum elimination capacity
of 60.89 g/m3 /h of the biofilter was obtained at inlet BTEX load of
63.14 g/m3 /h. In another study, Gallastegui et al. [212] examined the
interactions between toluene and p-xylene in air treatment biofil-
ters, packed with an inert filter media. Three lab-scale biofilters were
used to evaluate the effect of the inlet load of toluene, p-xylene and
mixtures of both compounds on the biodegradation rate. A maximum
elimination capacity of 26.5 and 40.3 g C/m3 /h for an inlet load of 65.6
and 57.8 g C/m3 /h was obtained for p-xylene and toluene biofilters,
respectively. The presence of toluene seemed to inhibit the biodegra-
dation of p-xylene when the mixture was treated; while the presence
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M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122 1115

Table 6
Main features of various bioreactors.

Reactor type Advantages Disadvantages Ref.

Biotrickling - Effective treatment of acid-producing pollutants - Accumulation of excess biomass in the filter bed [69,180,182]
filters - Lower pressure drop during long term operation - Complexity in construction and operation
- A high removal performance for hydrophilic VOCs - Production of secondary waste streams
Bio-filters - Lack of mobile aqueous phase - Clogging of the medium [69,183,184]
- Suitable for low water solubility gases - Medium deterioration
- High efficiency in BOD removal - Less treatment efficiency at high concentrations of pollutants
- Large area for mass transfer between the phases - Limited flexibility and control
- Low operating and capital costs
Bioscrubbers - Excellent stability of process parameters (pH, temperature, - Elevated production of wastes, [69,185–188]
nutrients) - Contaminants in the liquid state
- Relatively small pressure drop - Low efficiency in the case of substances poorly soluble in
- Relatively small size of equipment water
- Necessity to control the growth of biomass so as to restrict
the amounts of solid waste being produced.
Hollow - Compact with a high interfacial area between air and biofilm - High construction costs [69,189,190]
membrane phase - Long-term operational stability
reactors - High cell densities - High cost of membrane fabrication
- Independent control of air and nutrient flow rates with no - Membrane fouling
flooding

- Control of biomass concentration is easy

- Operated without clogging
Novel rotating - High volatility along with high water-solubility. - Technology is not well established [191,192]
rope bioreactor - Higher interfacial area
- High oxygen mass transfer rate
- Greater microbial culture stability
- Higher substrate loadings and removal rates
Bioactive foam - No packing in the reactor - Stability problem at high air velocity [193]
emulsion reactor - Not subjected to clogging - Oxygen limitation
- Surpasses the performance of existing gas phase bioreactors
- Reuse of emulsion cells
- Rapid mass transfer

Flat plate vapor - Low cost - Accumulation of dead cells on the top of the biofilm [194]
phase bioreactor - Good performance. - The lack of activity in the surface film

Two-phase - Robust and reliable - Scale up of mechanically agitated may not be feasible [195–197]
partitioning - Enhancing the productivity in fermentation technology - Requires large quantities of organic solvent
bioreactors - A higher overall concentration gradient which increases the - Excessive energy inputs
(TPPBs) driving force for VOC transfers to the aqueous phase
Airlift loop - Less energy - Hydrodynamics, mass transfer and bioreaction are complex [188,197–
reactor (ALR) - Ease design scale up and they strongly couple together 199]
- Poor mixing
External loop - Versatility - Oxygen mass transfer rate is smaller than that in well-mixed [200,201]
airlift bioreactor - Simple construction bioreactors
(ELAB) - Ease of operation - Limit the growth rate of cells
- Fewer chances of media contamination
- Lower energy consumption
- Absence of regions of high sheer exist near the impeller.
Internal loop - Preferentially at large scale - Sparging can damage mammalian cells and insect cells [202,203]
airlift bioreactor - High and readily controllable liquid circulation velocity - Agitation may have detrimental effect on animal cell
(ILAB) - High efficiency of homogenization bioreactors
- Intense mixing - Damage to cells on macrocarriers is found to result from the
- Better mass transfer performance power dissipation in the form of turbulent eddies.
Fluidized bed - Immobilization of microorganisms on small, porous - Relatively high energy consumption [204–207]
bioreactors fluidized media as biofilms results in higher biomass
concentration
- Reducing hydraulic retention time (HRT) with high
treatment efficiency
- No bed clogging, high pressure drop, poor mixing and
oxygen transfer
- Provide larger surface area for nutrient transfer
Spouted bed - Systematic intense mixing - May be difficult to maintain the bed fluid dynamics in large [208,209]
bioreactor (SBBR) - Better contact between substrate and cells beds
- Faster oxygen transfer rate
Packed bed - Efficiency and stability - Have large dead zones [210]
reactor - Easy scale-up - Channeling
- High pressure drop across the column
Monolith - Low pressure drop - Clogging of the channels for long-term stable operation [211]
bioreactor - Large pore size
- Large specific surface area and thin walls
- Better liquid distribution at low liquid flow rates
- High mechanical strength
- Scaling up relatively easy
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1116 M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122
of p-xylene enhanced the toluene removal efficiency.
Bioscrubbers
A bioscrubber consists of two units which are the absorption unit
and the bioreactor unit. In the absorption unit, the gas and liquid
flow countercurrently and influent gases are transferred to the liq-
uid phase. The depolluted gas is released at the top of the column,
while the contaminated liquid phase is pumped to an agitated, aer-
ated bioreactor. This reactor unit contains suitable microbial strains
suspended in nutrient solution [69]. Littlejohns et al. [188] devel-
oped a dynamic mathematical model to predict the performance of a
stirred tank, solid-liquid two-phase partitioning bioreactor (SL-TPPB)
for the treatment the BTEX contaminated gases. The SL-TPPB system
contains a bacterial consortium and a solid phase of silicone rubber
beads (10%, v/v) with a high affinity for BTEX compounds. The model
was developed from mass balances on BTEX components in the gas,
aqueous and polymer phases and biomass in the aqueous phase. The
model was capable of estimating the off-gas BTEX concentrations with
reasonable accuracy.
Hollow fiber membrane reactors
Microporous hydrophobic hollow fiber membranes are usually
made of material such as polyethylene that serves as a support for
the microbial population and provides a large surface area for VOCs
and oxygen mass transfer [189]. Dong-Jin and Kim [189] examined the
biodegradation of toluene using Pseudomonas putida type A1, which
was isolated and enriched to treat toluene vapor in a hollow fiber
membrane bioreactor. Batch experiments were conducted to study
the effect of oxygen levels on the microbial activity. The results show
that a low oxygen level does limit the degradation rate of toluene.
However, a hydrophobic polyethylene microfiltration hollow fiber
membrane bioreactor showed that toluene removal efficiencies were
constantly kept in the range of 86–97% at the loads of 0.85–4.3 kg Tol.
/m3 day for 150 days.
Kumar et al. [213] studied the performance of laboratory-scale
biofilm membrane bioreactor inoculated with Burkholderia viet-
namiensis G4 to treat toluene vapors in a waste gas stream over a
period of 165 days. The gas feed and nutrient solution were separated
by a composite membrane consisting of a porous polyacrylonitrile
(PAN) support layer coated with a very thin (0.3 µ m) dense poly-
dimethylsiloxane (PDMS) top layer. The biofilm membrane bioreactor
was operated at different residence time (28–2 s) and loading rates
(1.2–26.7 kg−3 /day), with inlet toluene concentrations ranging from
0.21 to 4.1 g−3 . The removal efficiencies ranged from 78% to 99%.
Novel rotating rope bioreactor
Mudlair et al. [191] developed a novel immobilized bioreactor for
the treatment of pollutants that have high volatility along with high
water solubility and low microbial yields. The rotating rope bioreac-
tor is characterized by higher interfacial area (per unit reactor liquid
volume) along with higher oxygen mass transfer rate; larger micro-
bial culture stability; and as a result, higher substrate loadings and
removal rates in comparison to other conventional rectors for the
treatment VOCs. The researchers reported that the RRB system was
able to degrade pyridine with removal efficiency of more than 85%
at high pyridine concentration (up to 1000 mg/L) and loading (up to
400 mg/m2 /h (66.86 g/m3 /h)), with a short hydraulic retention time
(9–18 h).
Bioactive foam emulsion reactor
Foam emulsion bioreactor (FEBR) consists of an emulsion of highly
active pollutant degrading microorganisms and a water-immiscible
organic phase. The FEBR is like the two phase partitioning bioreactor,
except that the amount of organic phase is low and it uses a bio-
compatible surfactant for foam production [132]. Shahna et al. [132]
proposed a novel bioactive foam emulsion bioreactor for treating con-
taminated gases as benzene, toluene and xylene (BTX). The gas–liquid
interfacial area was improved in FEBR by biocompatible foam and
driving force for mass transfer by a water immiscible organic phase.
Shahna et al. studied the effect of several parameters such as gas
residence time, oxygen content, and organic phase concentration on
bioreactor performance. They reported an average elimination ca-
pacity of 220 g/m3 /h with removal efficiency of 89.59% for BTX inlet
concentration of 1 g/m3 at 15 s gas residence time in the bioreactor.
Moreover, the optimum elimination capacity of the reactor for BTX
reached to 423.45 g/m3 /h.
In another study, Kan and Deshusses [193] developed a mathemat-
ical model and a proof of concept using the foam emulsion bioreactor
for treating toluene. The data showed a toluene elimination capac-
ity as high as 285 g/m3 /h with a removal efficiency of 95% at a gas
residence time of 15 s and a toluene inlet concentration of 1–1.3 g/
m3 . Oxygen limited the reactor performance at toluene concentra-
tion above about 0.7–1.0 g/m3 . The elimination capacity increased
from 204 to 408 g/m3 /h with >77% toluene removal at toluene inlet
concentrations of 2–2.2 g/m3 .
Flat plate vapor phase bioreactor
In the flat plate biofilm reactor, the gas stream contains humidi-
fied air with toluene-saturated which is supplied to the system as a
sole source of carbon and energy. Mineral salts medium flowed to the
reactor at a constant flow rate and the reactor is operated in coun-
tercurrent mode [194,214]. Villaverde et al. [194] studied the toluene
degradation process in a flat plate vapor phase bioreactor (VPBR) us-
ing a P. putida 54G biofilm. The results show a linear shape of the
dissolved oxygen concentration profile in the outer 87% of the biofilm
thickness. However, the oxygen consumption in the remaining basal
13% (0.3 mm) followed zero order kinetics with a rate constant of
102.2 g/m3 /h, for toluene gas concentration of 1.5 g M-3 .
Two-phase partitioning bioreactors (TPPBs)
The concept of two phase partitioning bioreactor system is based
on the use of a water immiscible and biocompatible organic solvent
that is allowed to float on the surface of a cell containing aqueous
phase. The solvent is used to dissolve large concentrations of haz-
ardous wastes, which then reduced into the aqueous phase with low
levels [93]. Singh et al. [93] used the two phase partitioning biore-
actor (TPPB) to biodegrade benzene at high concentrations. In TPPB,
5000 mg/L of benzene was biodegraded up to 50.17% over a period
of 168 h. Pseudomonas putida MHF 7109 was isolated from cow dung
microflora as potential benzene degrader and its ability to degrade
benzene at different concentrations was analyzed. The experimental
results indicated 100%, 81% and 65% degradation at the concentra-
tions of 50, 100, 250 mg/L within the time period of 24 h, 96 h and
168 h, respectively.
Airlift loop reactor (ALR)
The airlift loop reactor (ALR) is a bubble column with a draft tube
allowing the fluid to circulate in the reactor. The draft tube can be
placed inside or outside the bubble column, which is defined as the in-
ternal ALR (IALR) and the external ALR (EALR), respectively [199]. Pour
et al. [215] used an immobilized cell airlift bioreactor for the aerobic
bioremediation of simulated diesel fuel contaminated groundwater
and tested with p-xylene and naphthalene in batch and continuous
regimes. The study consisted of two stages; in the first stage, immobi-
lized soil bioreactor (ISBR) was used to develop an efficient microbial
 Author's personal copy
M.H. El-Naas et al. / Journal of Environmental Chemical Engineering 2 (2014) 1104–1122
consortium from the indigenous microorganisms, which existed in
diesel fuel contaminated soil. The immobilized cell air lift bioreac-
tor was then used in the second stage, with the cultivated microbial
consortia of the first stage. Volumetric biodegradation rates of p-
xylene and naphthalene, at biomass density of 720 mg/L were 15 and
16 mg/L h, respectively. Wang et al. [199] studied the biotreatment
of toluene in a gas–liquid–solid three-phase airlift loop reactor (ALR)
using the computational fluid dynamics (CFD) method. A transient 3D
CFD model was used for the simulation including the k–ǫ turbulence
model, interphase forces, momentum and mass transfer, bubble co-
alescence and break up, and the bioreaction. Optimized simulation
had been done based on the rate-limiting step and had found that the
decreased solid bead diameter could effectively develop the removal
efficiency.
Spouted bed bioreactor (SBBR)
The spouted bed bio-reactor (SBBR) is characterized by a system-
atic intense mixing due the cyclic motion of particles within the bed,
which is generated by a single air jet injected through an orifice in the
bottom of the reactor. It has many advantages over the conventional
bubble column and other flow bioreactors, including better mixing
and contact between substrate and cells, and faster oxygen trans-
fer rate, which lead to higher biodegradation rates. The reactor was
proved to be effective for the biodegradation of phenol [208].
Packed bed reactor
Packed bed bioreactor is a common natural filter bed, where pol-
luted air is moved in either a downflow or an upflow mode through
the bioreactor; the pollutants are then biodegraded by the biocat-
alyst present in the packed bed. Packed bed reactor is particularly
suitable for the treatment of hydrophobic and weakly water soluble
compounds with a Henry’s constant up to about 1 [216].
Monolith bioreactor
The monolith is used as catalyst support for VOCs treatment. The
Modified monolith bioreactor could provide relatively inexpensive,
light weight, inert, bioreactor packing which gives a high specific
surface area to improve the mass transfer rate. “The flow in monolithic
channels is bubble-train or Taylor. The gas and liquid flow through the
channels as separate slugs approaching plug flow behavior. Between
the gas bubble and the biocatalyst wall, a thin film is set up, through
which gas is readily transferred to the bio-catalytically active wall.
Inside the liquid slug itself, a recirculation pattern is observed. This
recirculation enhance transfer of gas from the caps of the bubble to
the biocatalyst” [211].
Summary
This review showed that the biological treatment can play a ma-
jor role in the elimination of harmful VOCs from industrial processes.
BTEX removal efficiencies up to 99% and above can be achieved by aer-
obic processes. Biological treatment is environmental friendly, very
efficient, cost effective and has gained increasing attention in pol-
lution prevention. These applications techniques in the laboratory,
large pilot plants and industrial scale opens the doors for future re-
search and explorations and offers new frontiers for developing new
and efficient technologies. Although many approaches are available
for BTEX removal, the search continues for the development and im-
provement in the future works. Further study should focus on the
microorganisms, considering more research for fungi and algae. Ar-
eas that could benefit from further research mostly revolve around
developing new bioreactors.
1117
Conflict of interest
None.
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