Journal of Sciences, Islamic Republic of Iran 28(3): 205 - 219 (2017) http://jsciences.ut.ac.

ir
University of Tehran, ISSN 1016-1104

Review Article
The Study of Biological Technologies for the Removal of
Sulfur Compounds

F. Nazari1, ME. Kefayati2, and J. Raheb1*

1
Department of Industrial and Environmental Biotechnology, National Institute of Genetic Engineering
and Biotechnology (NIGEB), Tehran, Islamic Republic of Iran
2
Department of Chemical Engineering, Faculty of Engineering, University of Tehran, Tehran, Islamic
Republic of Iran

Received: 19 November 2016 / Revised: 3 July 2017 / Accepted: 29 July 2017

Abstract
Combustion of sulfur components of fossil fuels such as oil causes the emission of SO2
in the atmosphere and lead to the formation of acidic rain in the environment. The
conventional approach for desulfurization of fossil fuels is the chemical procedure of
hydrodesulfurization (HDS). However, this method has low efficiency for desulfurization
of ring components of sulfur such as dibenzothiophene (DBT) that include a significant
percentage of the total sulfur content of fossil fuel. biodesulfurization (BDS), is a biological
method proposed for desulfurization of ring components of sulfur which is a non-
destructive pathway to remove sulfur from hydrocarbons of petroleum in the mild
conditions which potentially used as complementary with HDS. For industrial application
of BDS, the approach needs the new challenge to enhance desulfurization activity by
genetic engineering methods and bioreactor development to achieve from a fantasy
technique to an industrial and reality method for reduction of sulfur from fossil fuels. In this
review, we studied and evaluated the BDS and advances in the two last decades.

Keywords: Biodesulfurization; Hydrodesulfurization; Dibenzothiophene; Fossil fuels.

During the combustion of sulfur compounds Sulfur-

Introduction oxide gases [mostly SO2] are released and this results in
Particulate emissions from the combustion of fossil both serious air pollution and poisonous metal catalysts
fuels into the atmosphere have created serious problems [2]. Benzothiophene [BT], dibenzothiophene [DBT],
for our planet [1]. Particles of carbon dioxide and and their alkylated derivatives make up more than 50%
nitrogen and sulfur oxides play a role in the heating of of the total sulfur content of commercial diesel [3,4].
the earth and the creation of acid rain, destruction of These aromatic thiophenes as recalcitrant organic sulfur
forests, poisoning of lakes and destruction of buildings. compounds have higher resistance to hydrodesulfuri-
Also, due to the high percentage of sulfur in fossil fuels, zation [HDS] treatment than other sulfur compounds
and a high percentage of sulfur oxides in gases, they can such as mercaptans and sulfides [5].
put human health in danger strongly (For example, they Among sulfur oxides, SO2 is widely abundant and
can infiltrateinto the respiratory system and disrupt it). produced in the lower atmosphere. Moreover, this sulfur

*
Corresponding author: Tel: +982144787387; Fax: +982144787399; Email: jamshid@nigeb.ac.ir

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oxide can cause the formation of sulfate aerosol. Aerosol conventional method for removal of sulfur compounds
particles have an average diameter of 2.5 micrometers such as kerosene and diesel oil is the
and can be transferred to the lungs and cause breathing Hydrodesulphurization chemical methods [Hydrogen].
problems (World Bank, 1999 Atlas 2001). The hydrogenic method is a catalytic process done
SO2 can react with atmospheric moisture and cause under high pressure [150-3000 lb/in2] and high
acid rain or fog with a low pH. The acid formed in this temperature [290-455˚C] and also uses hydrogen gas in
way can cause corrosion of monuments. It can also be the presence of a metal catalyst, where finally the sulfur
transferred to the soil, cause damage to trees or affect the of oil compounds is converted to hydrogen sulfide. But
acidity of lakes with low buffering capacity and threaten as mentioned above, a significant proportion of sulfur in
marine life (EPA, 2006). some oil products [especially gasoline and diesel] is in
If SO2 is transported by air currents, it can be the form of heterocyclic organic compounds [such as
produced in one region and leave devastating effects in Benzothiophenes] that are resistant to this approach
the thousands of kilometers [in a remote location]. So it [12,13,14,15, 72, 73] and in order to remove them via
is necessary to develop international cooperation to hydrogenic method, higher temperature, pressure, and
control SO2 emissions. rates of catalyst are required, which considerably
Also, sulfur oxides emitted from burning of sulfur- increase the desulfurization costs.
containing compounds in fossil fuels can cause unknown
effects on the environment and the economy. Therefore, Biological Removal of Sulfur from Oil
in order to reduce emissions of sulfur oxides governments The biological method can be considered as a non-
around the world imposed new laws based on which oil destructive method in which microbes specifically
products with low sulfur content must be used [6,7]. remove sulfur from biodegradable hydrocarbons [16-
In1979, Canada, the United States and Europe in 21]. This method could be used in mild conditions and
particular, signed a number of agreements to reduce and has presented a proper potential for industrial
control emissions of SO2. Most of the agreements desulfurization industry.
focused on the means of transportation fuels because In the hydrogenic method, in addition, to taking
these means are one of the world's most important higher cost and energy, sulfur is not separated
sources of SO2 emission factors. completely from heterocyclic poly-aromatic
The highest sulfur content in diesel was set to be 10 compounds. Hence researchers are focused on the use of
ppm in 2010 in America [8]. It is also important to microbial desulfurization [Biodesulfurization] where
remove sulfur for engines equipped with nitrogen oxide sulfur removal reaction [under the very mild condition
[NOx] storage catalysts because the sulfate produced by and with lower costs], is performed by bacteria and
the sulfur in the fuel has toxic effects on these catalysts. enzymes, produced by them. So that they can fix the
Sulfate is unusually stable in thermal conditions and flaws in the hydrogenic method through microbial
saturates the reduction sites on the catalyst. So accessible desulfurization and in fact replace [full or partial] the
space for NOx reduction is reduced and their hydrogenic method, or complete it as a supplement.
effectiveness is lowered. To overcome this problem, Microorganisms require sulfur [in the structure of
desulfurization processes should be designed or some cofactors, amino acids, and proteins] for the growth
promoted to achieve sulfur content less than 10 ppm [9, of and their biological activities [22,23]. In addition,
10]. There are generally two different methods to reduce microorganisms are dependent on enzymes and metabolic
the impact of emissions of sulfur oxides: Pre-combustion pathways and have the ability to provide their sulfur
and post-combustion. requirements by various sources using enzymes.
The pre-combustion method has more advantages Some of these microorganisms are able to consume
than the post-combustion method. For example, in flue sulfur compounds in thiophene components by enzymes
gas desulfurization [post-combustion method] applied in the metabolic pathways and reduce sulfur content in
on hot corrosive exhaust gases is an expensive method. fuel. In general, desulfurization by microorganisms has
Quality control of treated flue gas in any part of the advantages, firstly because it is done in low temperature
treatment is virtually impossible. In addition, this and gentle pressure and this can be considered as an
method is limited and cannot be extended for everyone. energy-saving process. Second, biocatalysts are
In other words, pre-combustion of treatment method involved in biological activities that can be greatly
reduces the sulfur dioxide emissions from fossil fuel specific to their substrate.
without being dependant on combustion process type. In addition to structural applications of sulfur in
Also because the desulfurization solutions are not very biological macro molecules, some microorganisms take
frequent, their quality control it is much easier [11]. The sulfur as an energy source, so the microorganisms

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should obtain their sulfur from sulfur compounds such thiophene compounds were used as a source of carbon
as oil sulfur compounds. The problem is that the and sulfur, it was believed that attempts to
bacteria may take hydrocarbon compounds of oil as biodesulfurization were failed [31.32]. In this way,
carbon source that reduces the value of the fuel. Yamada and colleagues conducted a systematic study
Accordingly, the scientist searched for bacteria that do on microbial desulfurization. They reported that
not consume the oil hydrocarbon compounds and only Pseudomonas genus microorganisms, break
consume isolate and use sulfur from the compounds. In Dibenzothiophene down into a water-soluble product.
Iran also some strains of of Gordonia RIPI [24-26] were Their Pseudomonas strains were Pseudomonas
separated and characterized by the Department of abikonasis and Pseudomonas jianii. On the other hand,
Biotechnology technology of Research Institute of Kodama et al. reported that amino acids and other
Petroleum Industry [Rhodococcous FMF] [27, 25] and carbon compounds are essential as co-substrate for
the National Institute for Genetic Engineering and oxidation of Dibenzothiophene and the growth of
Research Organization for Science and Technology to microorganisms. Microorganisms with desulfurization
remove isolated sulfur from benzothiophene compounds activity have a metabolism that breaks c-c links in
model and efforts are made for higher molecular heterocyclic sulfur compounds such as
identification for the to modify the microorganisms Dibenzothiophene, thereby decompose benzene ring and
genetically. finally, sulfate is released in a series of cascade
Recent discoveries about microbial desulfurization oxidative reactions. The reaction mechanism that
mechanisms can lead to the commercial application of attacks carbon skeleton is called “Kodama" pathway
desulfurization process using novel combination that includes hydroxylation of an aromatic ring, splitting
engineered components for high expression of the ring and oxidation. Finally, a water-soluble product
desulfurization genes. Production inhibitors are finally is obtained. Reactions related to the pathway are briefly
separated or improve biocatalytic processes via connection
of related structures in environmental conditions.
Few studies have been conducted in all fields of
biodesulfurization, for example, some studies are
conducted on biodesulfurization of spent engine oil [26].
The sulfur level in fossil fuel is 1000-3000 ppm. The
concentration of sulfur in diesel fuel is often higher than
5000 ppm. In the past decade, the level of sulfur in these
fuels is reduced from ppm 2000 - 5000 to less than ppm
500 ppm. Recently, using new mechanisms it becomes
much less than 350 ppm. During the period [2005-2007]
it was expected that refineries produce low sulfur fuel
[sulfur content less than 10-15]. Thus during the last 10
years, there was a significant interest in the
development of technologies for removal of sulfur from
fossil fuels and their derivative products.
There are many refinery equipments traditionally
used for the removal of sulfur. In addition, several new
technologies have been developed for the further
reduction of the sulfur content of fossil fuels. One
solution is to use the ability of enzymes specific for the
production of diesel fuel and gasoline with low sulfur
content. The pathway known as biocatalytic
desulfurization [BDS] has a good potential for the
removal of sulfur from petroleum compounds [29,30].

Molecular destructive biodesulfurization

As mentioned, in the first step, by separating the
bacteria that have the ability to use DBT, since the
isolated microbial species could not specifically Figure 1. Desulfurization pathway by destroying the
separate the sulfur from DBT and in addition some molecular structure (34)

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described in Figure 1. plasmid with 9 open reading frames [open reading

In this reaction, other aromatic molecules in the oil frame], and the open reading frames are known as sox
are also attacked. And thus, a substantial amount of and also they found that only genetic pathways control
hydrocarbons enter the aqueous phase [33]. This type of the DBT, naphthalene and phenanthrene metabolize.
microorganisms generates a component of water-soluble The other pathway of destruction that resulted in
thiophene by oxidative decomposition of the mineralization of DBT was introduced by Van Afferden
Dibenzothiophene, as the production of oxidation that is et al. in 1993 along the pathway of DBT mineralization,
difficult to be separated from water [Kodama et al.]. there are three types of metabolic materials including
The microorganisms do not target the enzymatic DBT sulfoxide [DBTO], DBT sulfone [DBTO2] and
reactions of sulfur and sulfur so it is not practical to benzoate. This method of desulfurization is possible by
separate sulfur organic compounds with high molecular destroying the molecular structure and may be valuable
weight in crude oil. The reasons for these limitations for Biodesulfurization of DBT in the environment [34].
can be summarized as follows:
1) The attack on the carbon ring of Dibenzothiophene Sulfur-specific desulfurization pathway
often is done in positions 2 and 3 of Dibenzothiophene In this desulfurization pathway, Dibenzothiophene
replaced by alkyl or allyl groups as these situations do sulfoxide, Dibenzothiophene sulfonate, 2,2-
not apply to a substrate on Kodama pathway. 2) the dihydroxybiphenyls were characterized as intermediates
destruction pathway of the carbon skeleton reduces the of Dibenzothiophene in desulfurization. Apart from this,
amount of fuel energy. 3) The main product of the strains of Rhodoccucos rhodochrous ATCC53968 is
decomposition pathway of the carbon skeleton is 3- isolated that has a pathway to convert Dibenzothiophene
Hydroxy-2-Formyl benzothiophene and plenty of to hydroxyphenyl and sulfate.
Dibenzothiophene is decomposed to form sulfate and It has been reported that 70% of the organic sulfur in
thus sufficient desulfurization does not occur. crude oil and coal is reduced by these microorganisms.
Monticello and Denote concluded that the Kodama Also, in the case of Corynebacteria SP, A separate
pathway genes are for decomposition of DBT on a pathway to break Dibenzothiophene is described that

Figure 2. Four step pathway of desulphurization process (37)

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through the oxidation of Dibenzothiophene converts it middle host for transfer of genetic material. For this
to Dibenzothiophene sulfoxide, then Dibenzothiophene reason, expression plasmids in Pseudomonas have a
sulfone and finally the 2-Hydroxybiphenyls sulfate [35]. wide range of host and are more stable in enteric and
Many researchers have reported that special pathway non-enteric bacteria. Today, the expression vectors used
of desulfurization can be done by aerobic and anaerobic in Pseudomonas are mostly plasmids of incompatible
bacteria. However, there are reports that using the tests groups such as RP4 or IncQ groups that can be
under controlled anaerobic sulfate reducing conditions, transferred to conjugation method [51].
no significant reduction has been observed in the
amount of sulfur of vacuum gas oil. Since there is little Recombinant bacteria for Commercial
evidence in terms of the significant potential of Biodesulfurization
anaerobic desulfurization commercially, aerobic Advanced technologies help the development of bio-
desulfurization is widely considered. biodesulfurization for the treatment of oil components,
The 4s pathway is a special desulfurization pathway where biocatalysts are activated at 40 to 60 °C leading
where DBT is desulfurized and converted to 2HBP [2- to the removal of nitrogen and heavy metals, but the
Hydroxybiphenyl] [36, 74,75, 76, 77, 78, 79]. The 4S most important problem of biocatalysts is their
pathway condition along with factors and involved instability because they will die after 2-3 days [31]. So
enzymes, the carbon chain remains unchanged and the many works have been dedicated to making
fuel does not lose its value in the case of energy and desulfurization strains commercially advantageous.
burning value. Also, the sulfate ions formed in the 4S Unlike, the processes are taken for modification of
pathway are released in the culture medium or may be expression of effective enzymes in this pathway, this
adsorbed by other microorganisms. Figure 2. process is so slow to be used as an industrial
R.erythropolis IGTS8 [38,39,40] and Rhodococcus sp. application, both in the case of rate and efficiency of
X 309 [13] are the first strains of bacteria that desulfurization. Desulfurization rate higher than 20
researchers obtained lots of information at the molecular micromoles per gram of substrate in a minute per gram
level about the 4S desulfurization pathway. In this of catalyst is required. Successful separation of sulfur
regard, a gene cluster that could complete the mutant from petroleum compounds needs enzymes with high
bactericidal activity [IGTS8] were cloned and specificity and low Km in dsz system for various
sequenced, [14] also, it was found that this gene cluster bacteria [52.53]. Successful separation of sulfur from
contained three open reading frames on a plasmid petroleum compounds needs enzymes with high
identified as ABC dsz. specificity and low Km in dsz system. Recently, some
notes about gene shuffling of dsz genes have been
Desulfurization process in the pathway of development introduced in different bacteria to create novel genes
An investigation carried out after cloning revealed and more substrate specificity. Results showed that the
that dsz C product, converts DBT directly to DBTO2 developed techniques can be used create new hybrid
and also converts the gene products of dsz A and dsz B enzymes with required activity in refinery applications
DBTO2 to 2HBP [2-hydroxybiphenyl] [41]. The gene and remove the needs of bacteria in removal of sulfur
fragment is related to desulfurization phenotype on a components from petroleum compounds [54]. Use of
large plasmid that contains three genes of B, C dsz A the developed biocatalysts is essential for commercial
and is about 4kb long. In Rhodococcus bacteria, dsz application of BDS, however, in most chemical
genes of plasmid are close to entering sequences desulfurization processes, oil components are under HP
[42,34]. and HT and reaction cooling for microbial processes is
The important point about this genetic not practically limited and they can be stable for a long
desulfurization is that the set of genes related to time. There are a number of very thermophilic enzymes
desulfurization are in the form of one operon with three- that are active at temperatures higher than 100 °C and
genes [dsz A and dsz B and dsz C] organized by a single can be stable for a long time and on the other hand their
promoter [43,39]. stability may be increased by pressure. The main cause
Due to the weaknesses of Rhodococcus IGTS8 and in of this behavior is not clear and no creation of thermal
order to commercialization and optimization of stability in desulfurization enzymes has been done so
desulfurization, 4S pathway genes of Pseudomonas [44- far, however Clinical isolates of Bacillus have been
50, 41] have been cloned. known recently which desulfurized DBT at 55 °C. In
Generally, due to the transfer of genetic material into order to optimize and commercialize recombined
non-enteric gram-negative bacteria with very low and bacteria, researches aim to use thermophilic enzymes
poor efficiency, the best way is to use E.coli bacteria as and biosurfactant genes, more than before [51].

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Considerable efforts have been taken to make hydrocyclone or in combination with other low-cost
appropriate strains for commercial use. In 1999, the first centrifuge or precipitator systems.
patent on the accession of desulfurization genes [dsz] The entire procedure included reactor design,
was conducted in Pseudomonas in the United States. production, recycling and separation of water –oil
The other work on the accession of flavin reductase in a phase. New hypotheses are significant from several
synthetic operon containing all the genes required for aspects.
BDS pathway with a single transcript was released. First, use of multi-stage reactors to cope with weak
Since both the accession of genes in Pseudomonas and kinetic of reactions in low sulfur concentrations, second,
use of flavin reductase have been recently released, it keeping the fossil fuel value and ultimately use of the
seems these patents contain remarkable points for continued growth systems and biocatalysts regeneration
creation of useful business processes. Unlike, the in the reaction system and separation of them in external
processes for modifying the expression of key enzymes tanks, Figure 4.
in this pathway, the process is a very slow for
commercial use; this is a problem in the rate and the Reactor design and implementation of a process for
amount of desulfurization. Desulfurization rate more biological conditions
than 20 mM substrate, per minute, per gram of catalyst In addition to the genetic engineering, several other
is also needed [54]. conditions should be considered for industrial use of a
biocatalyst. First, the already grown cells should be
Development of biodesulfurization processes considered as none- grown biocatalysts. This reduces
Recently, a significant development has happened in the probability of producing agents that are active in
microbial desulfurization process and its economic their surface area, in response to cell growth in oil.
discussion. A hypothetical form of industrial These agents make it difficult to isolate production
Biodesulfurization is shown in Figure 3. 3 sets of Pathways. Biocatalyst cells should not need the
reactors are required to obtain a petroleum product with presence of expensive induction agents, such as
very low sulfur content, due to the high rate of Km of naphthalene or salicylate to achieve maximum activity.
dsz system. In this scenario, the cells are grown first and - In order to have a low-cost biocatalyst production,
then find resurgence in the process. This was a cells must grow on a low-cost carbon source and
necessary step for the long-term commercial use of produce a large amount of active biomass for each
biocatalyst. Isolation stages [boxes] can be used just by substrate.

Figure 3. A hypothetical form for microbial desulphurization of oil (55)

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Figure 4. A hypothetical diagram for microbial desulfurization of oil (54)

- Biocatalysts' substrates should be non-toxic. used in the presence of 40 to 50% of normal

Fortunately, inappropriate ring sulfur toxicity is Tetradecane or kerosene and 50% of diesel or 96%
decreased by dissolving in these compounds in oil, as a hexadecane. It has been found that emulsions of the oil
result, these components have lower toxicity during the phase and the upper layer contain significant amounts
desulfurization of oil, compared to when used in pure of bacterial particles that form droplets with a diameter
form. of 1 to 10 micrometers, which accumulate in high
 The lack of precision in the use of proper hosts concentrations of hexadecane during desulfurization of
[bacteria], could lead to the creation of substrate DBT. This requires the use of bio-surfactants for
resistant biocatalysts. separation of parts effective in a commercial process.
 Biocatalysts should not be under Feedback effect Result of microbial desulfurization of petroleum
(Effect [positive or negative] that the product can put compounds was about removal of 30 to 70% of oil
on the expression of genes) of the product. It was sulfur content for middle oil, 40 to 90% of diesel oil, 65
observed that using ion exchange resins and removal of to 70% for diesel oil treated with HDS, 20 to 60% for
reaction products, oxidation of DBT has been increased light gas-oil, 75 to 90% for refined oil and 25 to 60%
10 times. In addition, by selection of suitable strains, crude oil. Although this is a significant amount of
genetic engineering or mutagenesis of biocatalyst cells, separation, according to the sulfur content the amount of
it is possible to prevent the feedback effect of the desulfurization of fuels is not effective. For example, in
product. the case of crude oil, a chemical mechanism used for
Although reports about the isolation of various DBT separation of heteroatoms from asphalt and polar
decomposing microorganisms and enzymes with fractions lead to a a reduction of 24 to 40% of sulfur,
different substrate specificity, continues, still several nitrogen, oxygen and metallic elements together with
important issues remain to be resolved. The most hydrocarbon fractions [56].
important thing is how these highly hydrophobic
molecules find their way onto the primary enzyme [55]. Economic benefits of BDS
In new studies Desulfurization of a model compound BDS is more economical compared to the existing
[e.g, DBT], often was carried out in aqueous systems, HDS method and other desulfurization technologies.
resulting in a little resemblance to conditions of Advanced technologies help to develop a BDS method
biocatalysts is used in commercial affairs. Recently, for the treatment of oil components, where bio-catalysts
two solution phases are used. In aqueous systems in are active at 40 to 65 °C and lead to the removal of
order to increase the rate of desulfurization, DBT was nitrogen and metals from the oil. Another consideration

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for economic acceptability of BDS, includes safety Biological Removal of Sulfur Compounds From Gas
issues, transportation, storage and use of living bacteria Natural gas extracted from wells is different from
on the large scale for the refinery to use. Of course, what is known as regular gas. In underground
BDS process is not accepted in this view, as the cells reservoirs, gas is available in the free form [isolated]
remain alive for 1 to 2 days maximally. A common way and dissolved in an oil or in contact with oil and water.
to solve this problem is to produce and regenerate So, the gas extracted from the ground is in the form of
biocatalysts within the BDS process. In this way, the associated gas or dry gas and wet gas, respectively.
lifetime of biocatalysts reaches the 200 to 400 hours [8 Natural gas with a high percentage of methane, from
to 17 days]. any source, contains hydrocarbons such as ethane,
Studies to design a reactor led to the creation of propane, butane, and pentane after oil extraction and
modified condition, which reduces the mass transfer separation. In addition, the gas usually contains other
limitations and increases the volumetric reaction rate. substances such as water vapor, hydrogen sulfide [H2S],
The design of reactors required for the BDS process carbon dioxide, helium, nitrogen and other elements.
included designing tanks, appropriate aeration and Natural gas before the transfer and use should be
optimal environment with the low water-to-oil ratio. separated from these compounds so that some isolated
This process reduces the size of the reactor, but at the compounds are very precious and have other expenses.
same time, other processes are required to break the Natural gas is transported to the refineries via pipeline
resulting emulsion. Increased concentration of network. The main natural gas processing involves the
biocatalysts [up to 50 g dry cell weight per liter] leads to separation of dense gas, water, gas liquids, carbon
stable emulsion formation and causes problems in dioxide and hydrogen sulfide, Figure 5, [58].
separation. The information about the products obtained The production and use of biogas have recently
by BDS is low and scattered and there is generally little increased as this fuel represents a valuable renewable
information in this regard. Pacheco et al. produced a energy source. Biogas utilization produces an indirect
type of diesel oil by BDS that had more fluidity than oil reduction of greenhouse gas emissions through the
in the HDS method [31]. replacement of fossil fuel [59]. However, the use of
biogas is limited by the presence of hydrogen sulfide
[H2S] at high concentrations [0.1-2%]. H2S is a

Figure 5. Simplified scheme of the Claus process for chemical desulfurization under pressure of sour gas (57)

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corrosive and toxic compound that has an adverse different conditions of environmental stress such as lack
environmental effect due to the sulfur oxides generated of oxygen, acidic conditions and other factors, are
during combustion. increasingly being used to convert H2S and sulfur
The main approaches employed for gas compounds in a biological process. Separation of
desulfurization are physicochemical methods. However, hydrogen sulfide using a system of biological gas
physicochemical methods are characterized by high treatment was conducted in two stages: first, adsorption
consumption of energy and/or chemicals, and these of hydrogen sulfide followed by the oxidation of
methods can lead to other pollution problems such as hydrogen sulfide by the absorbent solution.
the generation of large amounts of carbon dioxide Biological filters are used for separation of H2S in
[CO2], nitrogen oxides or exhausted adsorbents that wastewater treatment. The process is basically conducted
require disposal [60]. One of the most widely used by aeration and oxidation product is often sulfate rather
biological methods for the purification or treatment of than elemental sulfur. The disadvantage of this
gas streams is biofiltration. Biofiltration is a safer and technology is to reduce the pH of environment that has a
cleaner technology. The development of biofiltration negative impact on the process. The H2S anaerobic bio-
has been rapid in recent years because it is less filters, as well as aerobic biofilters are effective, with the
expensive than other technologies, has good advantage of using cheaper raw materials. The removal
performance at the pilot scale and in industrial means the removal of safety risks often associated with
applications, and is feasible for the treatment of a wide aerobic processes which are another advantage of
variety of gaseous effluents [61,62]. A biotrickling filter anaerobic methods [65].
[BTF] is a packed bed bioreactor with biomass
immobilized. The gas flows through a fixed bed usually Improving biological removal of sulfur from natural
counter-currently to a mobile liquid phase. Synthetic gas
carriers are usually used such plastic, ceramic, lava A technology was designed and developed in
rocks, polyurethane foam, etc. First of all, the pollutant Wageningen University's environmental technology
compound must be transfer from the gas to the liquid sector, in collaboration with Delft University
phase and finally, the degradation is carried out in the Department of Biotechnology. The process is designed
biofilm. Fresh medium is fed to provide nutrients and in alkaline pH and high concentration of sodium which
remove the oxidation products [60].The biological has high performance for desulphurization of natural
removal of H2S from biogas has been mainly studied gas. First, the biological oxidation of sulfide was used to
under aerobic conditions [63,64, 69-71]. study sulfide separation in anaerobic form, then
Biodesulfurization of gas varies depending on Wageningen University used the process for the
different sources. In order to separate H2S from biogas, separation of H2S from biogases produced in anaerobic
anaerobic methods via phototroph bacteria are used. In form by wastewater. Then, by collaboration with Shell
this way, no environmental problems are caused. In this Co the process was developed for desulfurization of
method, the bacteria [cholorobium limicola] is used natural gas, refinery gas and other types of gases [Figure
which is very convenient to use. Due to the use of the 6] [57].
non-organic environment for bacterial growth, it has
high efficiency in converting the sulfide to elemental The mechanism of the process
sulfur [57]. In this process, the H2S carrier gas is passed through
The method because of being used to separate the an alkaline solution and this solution absorbs H2S which
biomass and being used in high-volume, fixed reactors form disulfide later. Absorbed carrier solution is then
are economically suitable to convert sulfide to elemental pumped to a bioreactor such that sulfide is oxidized to
sulfur. If the light is fully under control, the product of elemental sulfur in the bioreactor. This is done by
oxidation will be non-toxic to a large amount of sulfur. bacteria oxidizing sulfur components [Chemical
The light source plays an important and valuable role in autotrophics]. Insoluble sulfur components were
the process. It is important to use an economic light separated from the water flow and immediately used as
source in this method so that it can be a good alternative sulfur fertilizers [57]. Biological oxidation of sulfide to
to chemical methods [58]. In the case of chemotrophic disulfide is conducted in by hypoxic conditions while
bacteria, there is no need to more control over the use of the formation of sulfate is done in extra oxygen
oxygen to produce elemental sulfur rather than sulfate. condition.
Even in the best circumstances, conversion of sulfide to
sulfur, still relatively unknown sulfate contents are 1) H2O+OH- HS-+H2O
produced. Thiobacillus species that can grow in 2) HS-+½O2 S+OH-

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Figure 6. Gas desulphurization biological process to remove H2S: the right side after the cross lines is the sign of the decline of
thiosulfate (57)

3) HS-+2O2 SO42-+H problems will appear. This problem is due to

2-
Thiosulfate [S2O3 ] is generated in hypoxic acidification of the alkaline solution. Therefore, because
conditions as low-sulfur oxidative compounds. the solubility of potassium salt is more than three times
The main disadvantage of sulfate and thiosulfate are: of sodium salt, a mixture of NaOH or KOH solution is
1) Less consumable elemental Sulfur is produced. used to prevent acidification. The flow used to drain
2) Production of protons [H +] causes acidification of sulfate and thiosulfate can be removed by a direct route
alkaline solution. of biological sulfate reduction.
3) In order to prevent the accumulation of sulfate and In the second anaerobic bioreactor, sulfate reductant
thiosulfate, these ions must be removed by a fluid flow. bacteria cause conversion of sulfate and thiosulfate to
By the flow, bicarbonate ions are also eliminated and sulfide which can again be directed straight back to the
this loss should be compensated [57]. The existing manufacturer aerobic sulfate bioreactor. As a result, the
process is activated in an alkaline medium of pH [8.2- water cycle of the process can be eliminated by reducing
8.5] and 5.0 M of salt. However, in high pH and high water consumption. In order to form sulfide in the step
salt concentration, following benefits are obtained: of reduction of thiosulfate, a donor electron such as
1) Removing sulfur from gases with high CO2 content ethanol or hydrogen is required. In terms of using high
such as natural gas is performed easier. pH and high concentrations of sodium and potassium,
2) Cumulative capacity of sulfide alkaline solution just natran-alkaliphilic bacteria are able to grow. In
significantly increases at high pH and high concentration of recent decades, different types of bacteria have been
carbonates. This reduces energy used in absorbing columns. isolated from alkaline lakes that have high performance.
3) In this case there is no need for energy
consumption to compensate for lost bicarbonate and in Biological Removal of Sulfur Compounds From Coal
terms of energy consumption, it is more economic Coal is an inhomogeneous fossil fuel that includes
because reactor solution contains a higher concentration varying amounts of oxygen, sulfur, nitrogen and
of sulfate and thiosulfate ions [57]. minerals in addition to carbon and hydrogen. Sulfur
If only sodium ions are used to be combined with content of coal deposits depends on the formation place
bicarbonate, sulfate, and thiosulfate, due to sediment and associated sediments, which varie between 0.5 and
accumulation of sodium bicarbonate higher scales 10% and sometimes more. Sulfur in coal rocks is in two

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 The Study of Biological Technologies for the Removal of Sulfur …

Organic and inorganic forms. not able to remove organic sulfur. The self-sufficient [in
terms of food] and aerobic bacteria require relatively
An economic analysis of desulfurization of coal rocks thick oxygen, and if the amount of oxygen in the
Investigating the cost of removing sulfur and ambient air is less than 5% their activity is reduced and
comparison with the rising price of coal due to they are destroyed. Suitable pH range for their activity
reductions in the sulfur content is economically is 1.5-3.5 and outside the range diminishes their
interesting. Generally, the value of coal depends on activity. The bacteria are naturally activated in
these factors some of which can be improved by temperatures of 25 to 40 degrees Celsius, abundant in
desulfurization. the coal mines and output of many mine waters. The
1) Thermal power of coal [by increased thermal other category of desulfurization bacteria are
power, its value will be enhanced] Sulfolobus. The bacteria are found in the warm waters
2) The ash content [coal value has a reverse and adapted to temperatures higher than 50 °C and are
relationship with the ash content. By increasing the ash, active. The bacteria are not self-sufficient in terms of
its value decreases] food and nutrition is needed. These bacteria are capable
3) The sulfur content [sulfur and coal costs are also of removing organic sulfur and from this point of view
correlated inversely, by reducing sulfur content, its Thiobacillus has an advantage over Thiobacillus
value will enhance] ferrooxidans [31].
4) Coal dimensions [overall value of coal has a direct Laptev bacteria are also ideally suited for
relationship with a particle size and by the larger desulfurization. These bacteria are free from iron
dimensions, its value also rises] oxidation effect, but in the presence of sulfur oxidative
5) Moisture content [moisture content decreases its bacteria such as Thiobacillus they quickly oxidize pyrite
value] and remove its sulfur content. The thermophilic bacteria
6) Coke dealer capability [the value of coal with remove sulfur from coal more quickly which may be
Coking capability is several times greater than thermal due to the activity of this type of bacteria at a higher
coal value] [31]. temperature.
Microbial desulfurization method is new and by the In general, most bacteria quickly remove mineral
application of bacteria, almost all inorganic sulfur [at sulfur. But in the case of organic sulfur they are less
least 90 percent] can be removed in a relatively short active and don not have the potential for its removal or
time [a few days]. Organic sulfur removal is more perform it slowly and incomplete. In practice, the active
difficult because its elimination needs special type of bacteria on organic sulfur require feeding. So the main
bacteria mostly self-sufficient in terms of food factors in choosing the type of bacteria include replication
[autotrophic bacteria which obtain their required rate and self-nutrition should be considered [31].
oxygen, nitrogen and carbon from carbon dioxide and Application of different methods to remove sulfur
nitrogen in the air] and time to remove is very long from coal depends on the type of fuel, form of the
[several days] and is also very costly. Furthermore, sulfur, sulfur percentage, environmental standards and
nowadays the microorganisms used for this purpose are so on. Therefore, evaluating the advantages and
not able to remove organic sulfur completely and at the disadvantages of microbial method, performance of
best condition, they can remove sulfur at 50 to 60 bacteria and bacterial factors should be examined in
percent. Today, using genetic engineering, developed designing applications. [31]
researches have been conducted to cultivate bacteria,
which are self-sufficient for providing food, apply all Advantages and disadvantages of using bacteria to
organic sulfur components and have the maximum rate remove sulfur from coal
for removal of sulfur. But still no applicable result has The use of microorganisms to remove sulfur from
been presented in the industry. Since the sulfur removal coal, like any other method, has advantages and
bacteria produce sulfuric acid which affects coal ash disadvantages that must be considered in the course of
more or less and solves part of it, they decrease the ash their use. The advantages of using bacteria to remove
content to some extent, too [31]. sulfur from coal can be summarized as follows:
1. Working at low temperature
Types of bacteria used to remove coal sulfur 2. Low power consumption
Thiobacillus ferrooxidans [TF] bacteria are the most 3. High efficiency of coal recovery
common microorganisms used for the removal of sulfur 4. The relatively simple technology
from coal. These bacteria are easily capable of oxidizing 5. Lower investment
pyrite and sulfuric acid production, but they are actually 6. Reducing transport cost

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2. Some of the microorganisms used for this purpose

are not self-sufficient in terms of food and nutrition,
which increases the price of desulfurization by bacteria.
3. Microorganisms are sensitive to the presence of
heavy metals in coal and if the amount of these metals
exceeds the limit, they are a not able to function and are
destroyed. Therefore, the bacteria cannot be used on all
types of coal.
The first semi-commercial continuous bio-
depyritization operation was designed With a capacity
of 50 kg raw coal per hour and consisted of three main
sections: a comminution bay, a bioreactor bay, and a
reject water purification [66, 67] and disposal system
[Figures 7 and 8].

Results and Discussion

Natural energy resources include fossil fuels [oil,
coal and gas], uranium [nuclear energy], water [source
of electrical power], and other cases such as wind, solar
and etc. Due to limited energy resources on the one
hand and heavy losses [such as environmental pollution
or corrosion] of energy consuming on the other hand,
Figure 7. Porto Torres biodepyritization pilot plant: Coal researches have been performed in different areas on the
comminution bay. 1 head bin, 2 belt feeder, 3 automatic efficient use and reduction of the damage. According to
scale, 4 magnetic separator, 5 hammer mill, 6 belt conveyor,
current and emerging issues especially in the field of
7 wet ball mill, 8 preparation tank (Modified and redrawn
from Rossi (66), and Loi et al. (67) environmental concerns, it is predicted that more
researches would be conducted in this context in future.
7. Lower side-products A significant part of energy is generated through
8. Conversion of coals with high sulfur content to fossil fuels. About 80 million barrels of crude oil are
sale coals extracted from the ground daily, including oil
Advantages and disadvantages of using bacteria: derivatives such as natural gas, crude oil, coal, tar and
1. Sulfur removal microorganisms are aerobic and etc. Almost 90% of the oil extracted from the ground is
need oxygen. Supply of oxygen in the desulfurization used as an energy source. Because much of the oil
environment is only feasible for chemical reactors and contains significant amounts of sulfur, its derived fuels'
can be done by injecting air, but in other cases the combustion causes a release of high amounts of sulfur
supply of oxygen for the bacteria is associate oxides into the atmosphere.
withproblems and increase the cost of desulphurization Now most sulfur in fossil fuels can be removed
because of mass transfer limitations and effectively low easily. However, there is a section known as
solubility of oxygen in the bioreactor. biodegradable organic sulfur that it is very difficult to

Figure 8. Porto Torres biodepyritization pilot plant: bioreactor and coal dewatering bay. 1 propagator, 2 pump, 3–9 stirred tank
bioreactors, 10 pump, 11 rake thickener, 12 diaphragm pump,13 settling pond, 14 to stock-pile (Modified and redrawn from Rossi
(66), and Loi et al. (67)

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 The Study of Biological Technologies for the Removal of Sulfur …

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