Research on Coal Desulfurization of Pseudomonas Stutzeri
Tingting Hu
1, a
, Yu Yang
1, 2, b
, Mengjun Zhang
1
, Qian Cheng
1
and Yu Gao
1
1
School of Minerals Processing and Bioengineering, Central South University
2
Key Laboratory of Biometallurgy, Ministry of Education, 932 South Lushan Road, Changsha, Hunan, China, 410083
Email:
a b
Keywords: High-sulfur coal, Biodesulfurization, Pseudomonas stutzeri, Ultraviolet mutagenesis.
Abstract: The burning of high-sulfur coal releases sulfur dioxide and causes environmental pollution. An efficient
desulfurization strain is the key to the development of coal biological desulfurization. In this study,
Pseudomonas stutzeri LH-42 was employed as the experimental bacterial, and the mutant strain
Pseudomonas stutzeri ZW-15 with the highest desulfurization efficiency was selected by UV mutagenesis.
The mutant strain ZW-15 was applied to the bioleaching experiment for coal from Liupanshui mine, Guizhou
Province, China, and the result showed that 41% of the total sulfur and 93.25% of the organic sulfur in coal
was removed in a 15-days experiment, which indicated that the mutant strain ZW-15 do have an application
potential for the biodesulfurization of high-sulfur coal.
1 INTRODUCTION
China is one of the countries with the largest
production and consumption of coal in the world.
Chinese coal consumption accounted for half of the
world's total in 2015 (Yang et al., 2012, Liu et al.,
2016). However, the combustion of high sulfur coal
will cause serious environmental problems, such as
acid rain. (Burns et al., 2016). With the depletion of
high-quality coal resources, the proportion of high-
sulfur coal consumption is getting higher. Therefore,
how to reduce sulfur content of high-sulfur coal has
become a hotspot in environmental science
research(Zhang et al., 2013).
The best method to limit the amount of sulfur
oxides emitted into the atmosphere is to reduce the
content of sulfur in coal before combustion(He et al.,
2012). Coal sulfur has inorganic and organic sulfur
in two forms. Inorganic sulfur mainly exists in the
form of pyrite in coal, organic sulfur is mainly in the
form of Dibenzothiophene (DBT) (Mishra et al.,
2017). In the past years, we ofen took chemical and
physical methods to remove the sulfur from the coal,
however, these ways are high-cost, energy-intensive
and inefficient for removing organic
sulfur(Gonsalvesh et al., 2012). Thus, more and
more attention has been focused on the
biodesulfurization of high sulfur coal since it offers
a clean alternative method to remove sulfur from
coal (He et al., 2012, Khanna et al., 2011, Kodama
et al., 2000).
At present the biodesulfurization technology is
still in the laboratory research stage, because the
stable and efficient desulfurization strain is not easy
to obtain, there are some challenges in industrial
applications of coal biodesulfurization. However,
coal biodesulfurization technology still has great
potential for development and application prospects
with the exploitation of desulfurization strains and
improvement of biotechnology desulfurization
process.
In this article, Pseudomonas stutzeri LH-42 was
employed as the experimental bacterial. And we got
mutant strain ZW-15 with the highest
desulfurization efficiency after UV mutagenesis, and
described its characteristics of coal leaching
desulfurization experiment, and demonstrated the
possibility of its application for the
biodesulfurization of high sulfur coal.
2 EXPERIMENTAL SECTION
2.1 Coal Sample
The coal sample used in the experiment was
collected from Liupanshui, Guizhou Province,
China. X-ray diffraction (XRD) was used to analyze
the component of coal sample. These results showed
Hu, T., Yang, Y., Zhang, M., Cheng, Q. and Gao, Y.
Research on Coal Desulfurization of Pseudomonas Stutzeri.
In 3rd International Conference on Electromechanical Control Technology and Transportation (ICECTT 2018), pages 41-45
ISBN: 978-989-758-312-4
Copyright © 2018 by SCITEPRESS – Science and Technology Publications, Lda. All rights reserved
41

in Table 1. The total sulfur of coal samples is 4.973
%. The sample was pulverized in a ball mill and size
to less than 0.5 mm for desulfurization test (Cara et
al., 2003).
Table 1: Phase analysis results of sulfur in raw coal.
Sulfur forms
Content
(%)
Proportion
(%)
Total sulfu
r
4.973
Organic
sulfu
r
1.979 39.795
Sulfate 0.144 2.896
2.2 Media
The BSM medium was used for cultivation of
Pseudomonas stutzeri LH-42 contained: 2.44 g of
KH
2
PO
4
, 12.03 g of Na
2
HPO
4
•12H
2
O, 0.36 g of
MgCl
2
•6H
2
O, 2.00 g of NH
4
Cl, 0.004 g of
MnCl
2
•4H
2
O, 0.001 g of FeCl
3
•6H
2
O, 0.001 g of
CaCl
2
, 1.62 g of glycerin, and one liter of deionized
water (Zhang et al., 2017). Final pH value was 7.2.
Sulfur sources, DBT, were added to the medium at
the concentration of 0.3 mM. The BSM medium
with 0.3 mM of DBT was labeled as BSM(D) in the
following experiments.
2.3 Microorganisms and Cultivation
The strain Pseudomonas stutzeri LH-42 originally
isolated from the petroleum-contaminated soil was
routinely maintained in our laboratory(Yang et
al., 2013). The strain was inoculated into 100 ml
medium and cultured at 30℃ with constant agitation
(170 rpm) for 48h.
2.4 UV Mutagenesis
A mutant was obtained by UV mutagenesis to
increase the efficiency of the organic sulfur
degradation of Pseudomonas stutzeri LH-42. A cell
suspension (1 loopful/ml) was spread on the surface
of the agar plate. Then, the cells were irradiated by
UV light (30 W) for 0s, 10s, 15s, 20s, 25s, 30s, 45s,
60s (All experiments were run in triplicate) at a
distance of 30 cm, and cultivated at 30°C.
2.5 Coal Desulfurization Comparison
Test
The row coal used in the experiment were sterilized
and the biodesulfurization process of different
mutants was carried out in flasks with a volume
capacity of 100 mL in 250 mL of BSM medium,
15% w/v pulp density, the initial cell concentration
of 1.0×10
6
cells/mL and processing time of 20 d.
The wild strain was used as control. The sulfur
content of coal in this biodesulfurization system
were detected in the first five days, and after that,
the sulfur content was detected in every five days.
The best mutant strain was chosen to the following
experiment by comparing the desulfurization
efficiency.
2.6 Coal Bioleaching Experiment
To remove the sulfur of coal, the best mutant strain
Mutant ZW-15 cells were inoculated into column
containing 2L sterilized culture medium
supplemented with 1000g coal sample (the initial
cell concentration was 1.0×10
6
cells.mL−1), and
were incubated at room temperature, processing time
of 15 days. At the bottom of the column, layered
1000g of coal which was 10 Tyler mesh, and 1000g
of coal which was 20 Tyler mesh. The leaching
system sprayed for 1 minute per hour, and 2.5 L
liquid per minute. Triplicate leaching experiments
were performed under identical conditions. Parallel
experiments (without cells; but the same culture
medium) were prepared as sterile control. The pH
value and the redox potential of bioleaching system
were determined every day. After the bioleaching,
the sulfur content was detected again to calculate the
efficiency of biodesulfurization.
2.7 X-ray Diffraction Analysis of Coal
In order to further study the influence of sulfur
removal in coal of the induced strain ZW-15. Coal
sample of the initial coal and biological
desulfurization sample were tested by X-Ray
diffraction analysis (XRD), respectively.
3 RESULTS AND DISCUSSION
3.1 Purification and culture bacteria.
In order to obtain individual bacterial colony,
Pseudomonas stutzeri LH-42 was cultured in
BSM(D) solid medium. And several single colonies
were identified by analyzing their 16S RNA
sequences. The result of BLAST indicated that this
strain is Pseudomonas stutzeri (Yang et al., 2013).
The growth curves of the strain LH-42 which
cultured in BSM liquid medium containing 0.3
mmol/L of DBT was shown in Figure 1.
ICECTT 2018 - 3rd International Conference on Electromechanical Control Technology and Transportation
42

0 20406080100
0
20
40
60
80
100
120
Cell Concentrate (×10^6/ml)
Time (h)
Growth curve of LH-42
Abiotic control
Figure 1: Growth curves of the strain LH-42.
3.2 The results of UV mutagenesis
Pseudomonas stutzeri LH-42 was cultured in
BSM(D) solid medium and treated by Ultraviolet
ray. Lethality rate of spores of Pseudomonas stutzeri
LH-42 by Ultraviolet ray was shown in Figure 2.
The lethality rate of LH-42 could achieve 89.3%
when the strain radiated for 15 seconds, and it could
be further improved to 100% by increased the
induced time to 30s.
0 102030405060
0
20
40
60
80
100
Lethal Rate (%)
Time (s)
Lethal Rate
Figure 2: Lethality rate of spores of Pseudomonas
stutzeri LH-42 by Ultraviolet ray
.
3.3 The results of coal desulfurization
comparison test
The UV-mutated colony whose diameter was greater
than 2mm was selected to be inoculated into
BSM(D) medium supplemented with sterile coal
sample and cultured for 20 days.
Desulfurization rates
of wild and mutant stains were shown in figure 3. By
comparing the efficiency of desulfurization, we
found that a mutant strain named Mutant ZW-15
was the most efficient one. A comparison test
between wild strain and Mutant ZW-15 was proceed
to verify whether the mutant strain actually more
efficient. The desulfurization rate of Mutant ZW-15
was 49.00% compared with wild strain’s rate
21.36% (shown in Figure 3). Especially in the first
five days, the desulfurization rate of Mutant ZW-15
increased massively and there was not significant
change in the total sulfur desulfurization after 10
days’ leaching, which suggested that the period of
biodesulfurization could be shortened greatly.
0 5 10 15 20 25
0
5
10
15
20
25
30
35
40
LH-42(Mutant ZW-15)
LH-42(Wild type)
Control
LH-42(Mutant ZW-10)
LH-42(Mutant ZW-8)
LH-42(Mutant ZW-5)
Total Sulfur Desulfurization Rate (%)
Time (d)
Figure 3: Comparison of desulfurization rates between
wild and mutant.
3.4 The change of pH and redox
potential of coal bioleaching system
The change of pH value and the redox potential of
leaching desulfurization system were shown in
Figure 4. Compared with the control group, the pH
value of the leaching system showed significant
downward trend. The coal sample itself contains a
certain amount of humic acid which caused the
sharp decrease of pH in the first one hour after the
sample’s dissolved in water. The pH value of test
group dropped to 4.3 finally, which might because
the bacterial released H
+
while decomposed the
organic sulfur of coal. The slow oxidation of the
pyrite in coal also contributed to the reduction of
pH.
Research on Coal Desulfurization of Pseudomonas Stutzeri
43

0 2 4 6 8 10121416
4.5
5.0
5.5
6.0
pH of leaching system
pH of control group
pH
Time
(
d
)
-20
0
20
40
60
80
100
120
140
160
redox potential of leaching system
redox potential of control group
Eh (mV)
Figure 4:
The pH and redox potential of leaching
desulfurization test.
Compared with the control group, the redox
potential of the leaching system changed
significantly. The initial redox potential of test group
was 69 mv, but after 2 days the potential jumped to
152 mV. Due to the initial stage of leaching system
was unstable, there had been a small range of
fluctuation of redox potential. Along with the
process of bioeaching, the bacteria released metal
ion like ferric ion and ferrous ion while decomposed
the organic sulfur of coal, and it would increased the
redox potential(Hu et al., 2006).
3.5 Desulfurization rate of coal
bioleaching system
The mutant strain Mutant ZW-15 showed the better
performance compared with the wild strain in
desulfurization within 15 days, it degraded 41% total
sulfur and 93.25% organic sulfur from the coal
sample.
3.6 The results of mineralogical X-ray
diffraction analysis of coal
The XRD graphs collected from the bioleached
particles are shown in Figure 5. The Figure shows
that the pyrite content was high before leaching but
reduced after 15 days leaching process. The results
further validated that biodesulfurization by induced
strain ZW-1 can effectively remove inorganic sulfur
from coal.
0 102030405060708090
0
5
10
15
20
25
30
0 102030405060708090
0
5
10
15
20
25
30
Peak intensity(n/s)
A
Pyrite
initial coal sample
Peak intensity(n/s)
B
biodesulfurization sample
Figure 5: The XRD results of initial coal sample (A) and
biodesulfurization sample (B).
4 CONCLUSIONS
The application of biodesulfurization is still
premature so far, however, it has shown some
enormous potential in the development of energy
industry and environment protection. More active
microbial cultures with higher desulfurization
efficiency which can degrade a wide variety of
sulfur compounds are needed for process
development.
The mutant strain Pseudomonas stutzeri LH-
42(Mutant ZW-15) was used for the
biodesulfurization of coal. After 15 days’
processing, it degraded 93.25% of organic sulfur and
41% of total sulfur. Thus, we can conclude that
Mutant ZW-15 can be used as the efficient strain in
the coal biodesulfurization.
ACKNOWLEDGEMENTS
This work was supported by the Fundamental
Research Funds for the Central Universities of
Central South University (2017zzts383).
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