Application of Guar Gum Degrading Bacteria in Microbial

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Application of Guar Gum degrading bacteria in Microbial Remediation of Guar-based Fracturing Fluid Damage Xin Ma, Zhihui Wang, Qi'an Da, Mingming Cheng, Chuanjin Yao, and Guanglun Lei Energy Fuels, Just Accepted Manuscript • DOI: 10.1021/acs.energyfuels.7b00999 • Publication Date (Web): 27 Jun 2017 Downloaded from http://pubs.acs.org on July 2, 2017

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Application of Guar Gum degrading bacteria in Microbial Remediation of Guar-based Fracturing Fluid Damage Xin Ma,† Zhihui Wang,† Qi’an Da,† Mingming Cheng,‡ Chuanjin Yao,*,† and Guanglun Lei*,† †

School of Petroleum Engineering, China University of Petroleum, Qingdao 266580, China

‡

Department of Resources and Environment, Binzhou University, Binzhou 256600, China

ABSTRACT: Microbial remediation of fracturing fluid damage is first time to be presented in this paper. One guar gum degrading bacterial strain, Bacillus sonorensis, numbered by XSJ, was isolated from oilfield produced water and identified through 16S rDNA. The characterizations show that strain XSJ is mesophilic, alkalophilic, facultative and halophilic, which is suitable for the reservoir environment. In addition, the guar gum degrading performance of this strain was investigated through apparent viscosity of the guar gum solution, the average molecular weight of guar gum and gas chromatography. Besides, the degrading performance of insoluble residue of guar gum was analyzed through the measurement of particle size distribution using laser particle size analyzer and total weight of insoluble residue using weighing method. Finally, a kind of sand-pack column was designed to determine the recovery of permeability. The results indicate that: 1. Bacillus sonorensis can efficiently degrade guar-based fracturing fluids to less than 5.0 mPas of apparent viscosity and 50,000 of average molecular weight. 2. The bacterial

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strain can decrease the average particle size of insoluble residue from 105.56 μm to 72.40 μm and total weight of insoluble residue with the microbial degradation rate of 25.30%. 3. This strain can improve effectively the permeability of sand-pack column (max 90.32%) in laboratory simulation experiments. In conclusion, it is believed that the microbial remediation seems to be a new approach for remediation of guar-based fracturing fluid damage and has great potential in oilfield application.

1. INTRODUCTION Oil supplies the vast majority of the world’s energy as an essential nonrenewable resource.1 As the worldwide demand is continually rising, there is a dire need to develop unconventional oil and gas reservoirs, such as low or ultra-low permeability deposits, shale oil and gas, et al.. For example, in China, 72.8% of crude oil is trapped in low permeability reservoirs of all the proven reserves. However, the low permeability reservoirs account for 60.8% in the unused but proven reservoirs.2 To achieve commercial production, hydraulic fracturing has become a critical component in the successful development of these reservoirs.3 However, amounts of fracturing fluid residue are remained underground. Typically, only 10% ~ 70% of fracturing fluids is subsequently recovered as flow-back. 4 The fracturing fluid residue (e.g. filter cake, insoluble residue) remained in fractures or rock matrix have seriously limited the production rate of oil and gas. The fracturing fluid damage directly influences the development of unconventional oil and gas reservoirs and therefore the overall development tendency of hydraulic fracturing is to reduce the fracturing fluid damage to the reservoirs.


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Guar gum (galactomannan) is extracted from the seeds of Cyamopsis tetragonoloba, as a natural, water-soluble and readily biodegradable polysaccharide. 5 It consists of a linear backbone of α-1,4 linked mannose units and randomly β-1,6 linked galactose units attached as side chains.6 Guar gum is widely used as a thickener in the fracturing fluid. However, filter cakes forming on the two faces of fractures and insoluble residue after gel breaking will be the main damage mechanisms for fracturing fluid damage.7 To avoid the fracturing fluid damage, some reservoir simulators are also used to investigate the effects of unbroken fracture fluid on well performances to control fracturing fluid damage or recommend the method of fracture cleanup.8,9 In addition, fracturing fluid researches are shifting toward viscoelastic surfactant fracturing liquids due to non-damaging effects on oil reservoir or optimization of the fracturing-fluid system.10,11,12 However, guar gum and its derivatives are still used commonly in water-based fracturing fluids duo to the low cost and high technological maturity.13,14 To remediate the fracturing fluid damage, the early methods include chemical treatments such as strong acids or oxidizing materials (e.g. ammonium persulfate) to degrade the fracturing fluid residue through the redox reaction.15 However, the application of chemical treatments are limited duo to the non-specific chemical reactivity, such as incompatibility for acid sensitive formation, corrosion of tubular goods, etc., with finite effectiveness and requirement of activators.16 Recently, enzyme is also used to be gel breaker to remove the guar gum residue.17,18 However, enzyme activity is easily influenced by pH, temperature, heavy metal iron and enzyme can not reproduce and grow as microbes.19,20 As the development of biotechnology, microbial remediation of fracturing fluid damage provides an environmentally sustainable, highly efficient and cost-effective alternative to conventional treatment methods. In this novel method, microbes are injected into the formation


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after hydraulic fracturing. Filter cakes and insoluble residue will be the carbon source of microbial growth, which means that the biodegradation will clean the fracturing fluid residue in the pore-fracture media. The fracture conductivity and the rock matrix permeability will be regained, while the production rate of oil and gas will be recovered. To the authors’ knowledge, nobody else has proposed the hypothesis of microbial remediation of fracturing fluid damage and no study has examined the effect of microbial treatment on guar gum residue. The goal of this study was to demonstrate the application potential of guar gum degrading bacteria in the microbial remediation of guar-based fracturing fluid damage. (a) Guar gum degrading bacterial strains was isolated from oilfield produced water. (b) The effect of biodegradation on physicochemical property of guar gum and insoluble residue was measured. (c) The effectiveness of biodegradation on the permeability recovery was tested through sand pack column laboratory experiments. 2. MATERIALS AND METHODS 2.1. Materials The reagents of inorganic salt used in this study were analytical grade and produced by Chinese Medicine Group Chemical Reagent Co., Ltd.. Guar gum was supplied by Wanbo Chemical Products Co., Ltd., Henan, China. All reagents were used without any further purification. Deionized (DI) water was used for the preparation of all aqueous solutions and the injection water in simulation experiments. The formation water sample collected from an oil well in Gudong reservoir of Shengli Oilfield in Eastern China was used as inoculum for the enrichment of guar gum degrading bacteria. The temperature of the oil reservoir is approximately 53 °C and formation depth reaches 1000-1100 m. The artificial brine water (shown in Table 1) with the total salinity being 91173 mgL-1 used in the experiment was prepared according to the 4 ACS Paragon Plus Environment

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salinity of the brine in Ansai Oilfield where is one of the important ultra–low permeability reservoirs in China.21 Table. 1 Salinity Analysis of Artificial Brine Water pH 6.8

water type

salt ion concentration (mgL-1) CO32-

HCO3-

Cl-

SO42-

Ba2+

Ca2+

Mg2+

K++Na+

Salinity

0

80

56380

0

650

21000

80

12980

91173

CaCl2

2.2. Bacterial isolation, identification and characterization 2.2.1 Enrichment. The formation water sample was used as inoculum for the enrichment of guar gum degrading bacteria. 10 mL of formation water sample was added into a 250 mL glass bottle containing 100 mL of the enrichment medium [Guar gum 4.0 g/L; Salt solution (KNO3 2.0 g/L, (NH4)2SO4 2.0 g/L, K2HPO4 2.0 g/L, MgCl2 0.15 g/L, CaCl2 0.15 g/L); pH 7.0]. Enrichment was incubated at 50 °C in hot air oven for 7 days. 2.2.2 Isolation. The bacterial isolation was set up by dilution plate method.22 50 µL diluted enrichment liquid was spread on the isolation medium [Guar gum 4.0 g/L; Ager 10.0 g/L; Salt solution (KNO3 2.0 g/L, (NH4)2SO4 2.0 g/L, K2HPO4 2.0 g/L, MgCl2 0.15 g/L, CaCl2 0.15 g/L); pH 7.0] and incubated at 50 °C in hot air oven for 3 days. The growing fastest and biggest morphologically distinct colonies were picked and purified by streak inoculation in the beef extract-peptone ager medium (Beef extract 3.0 g/L, Peptone 10.0 g/L, NaCl 5.0 g/L, Ager 10.0 g/L; pH 7.0). The purified cultures were stored at 4 °C on beef extract-peptone ager slope medium in test tubes. 2.2.3 Identification. Bacterial strain numbered XSJ was identified by PCR-sequencing analysis of 16S rDNA gene fragments of XSJ consortium as the method of Sachdey et.al.23 with some modifications. Firstly, DNA extraction of XSJ consortium was done by CTAB method.24 Then PCR amplification of 16s rDNA was carried out using 27F(5’-AGA GTT TGA TCC TGG CTC


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AG-3’) and 1492R(5’-TAC GGC TAC CTT GTT ACG ACT T-3’) in a gradient thermal cycler (Eppendorf, Germany). The following time–temperature profile was used. Initial denaturation at 95℃ for 5 min, 35 cycles of denaturation at 95℃ for 30s, annealing at 55℃ for 30 s and elongation at 72℃ for 1min. A final extension step of 10 min at 72℃ was included. Finally, agarose gel electrophoresis was used to detected the production of PCR amplification. The electrophoresis included 3 µL samples, 1% agarose gel with DNA mark (100, 250, 500, 750, 1000, 2000, 3000, 5000bp). The PCR production was sequenced by the Beijing Genomics Institute. Combined with observation experiments of physiological and biochemical examination. A neighbor-joining tree was constructed in MEGA 4.0 using 16S rDNA sequences of this bacterial culture and their close relatives in the Gen-Bank database. Bootstrap values were shown at nodes for frequencies at or above a 50% threshold (1000 bootstrap re-sampling).25 2.2.4 Characterization All the characterization experiments referred to the method of Cheng’s26. The growth curve was measured using cell count technique or the turbidimetry using spectrophotometer (721, produced by Shanghai Precision Scientific Instrument Co., Ltd.) with wavelength of 440 nm. Since bacteria grow exponentially, it is often useful to plot the logarithm of the relative population size [In(N/N0)] against time [t]. Based on the modeling of bacterial growth curve, the three phases of the growth curve include lag phase, logarithmic phase and stationary phase. They can be described by three parameters: the maximum specific growth rate, µm, is defined as the tangent in the inflection point; the lag time, λ, is defined as the x-axis intercept of this tangent; and the asymptote, A[A=In (N∞/N0)], is the maximal value reached.27 The mathematical model of Logistic equation28 for fitting the growth curve is following in Eq.(1).


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 N  A In  =  N 0  1 + exp  4 µ m ( λ − t ) + 2     A    

(1)

Where N is the real-time cell density ; N0 and N∞ are the cell density of the initial inoculum and in the stationary phase, respectively; t is the culture time; A is the maximal value reached; µm is the maximum specific growth rate; λ is the lag time. 2.3. Guar gum degradation 2.3.1 Biodegradation experiment. Viscosity method29 was used to measure the guar gum degradation of bacteria. Firstly, one piece of lawn was inoculated in 100 mL seed liquid medium (Beef extract 3.0 g/L, Peptone 5.0 g/L, NaCl 5.0 g/L; pH 7.0) and cultured at 50℃for 12 hours. The seed liquid medium after culturing was the bacterial seed liquid. Then, 0.5 mL bacterial seed liquid was inoculated in a 250mL triangular flask with 100 mL guar gum degradation medium [Beef extract 1.5 g/L Guar gum 4.0 g/L; Salt solution (KNO3 2.0 g/L, (NH4)2SO4 2.0 g/L, K2HPO4 2.0 g/L, MgCl2 0.15 g/L, CaCl2 0.15 g/L); pH 7.0]. They were incubated in water bath shaker at 50℃ and 100 rpm. The control treatment was prepared in the same way without inoculation. Finally, the sample was collected as following: (1) The liquid medium after degrading for a period of time was used to measure the apparent viscosity. (2) The guar gum degradation medium after apparent viscosity measurement was collected and centrifuged at 3000 rpm for 10 min and the supernatant was filtered through a 5.0 µm micro-porous membrane to remove impurities by vacuum filter. The filtrate was collected to determine the average molecular weight. (3) Gas production in triangular flask after the degradation for 3 days was collected using aluminum foil gas collecting bag for gas chromatography. 2.3.2 Apparent viscosity measurement. The apparent viscosity of samples was determined with a digital rotating viscometer (NDJ-1B-1, produced by Shanghai Changji Co., Ltd.) equipped with


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a temperature controller. Rotation rate was set at 60 rpm. The sample (50~80 mL) was added in a circular cylindrical container. Appropriate concentric cylinder geometry rotator was used to measure the viscosity. For example, 0# or 1# rotator was used to measure the solution samples with 1.0~10.0 mPas or 10~100 mPas. 2.3.3 Average molecular weight measurement. Intrinsic viscosity is a measure of the inherent ability of a polymer to increase solution viscosity.30 It was determined by the experimental results of dilute solution viscosities and extrapolating to infinite dilution, according to Huggins’s Equation (Eq. (2)) and Kraemer’s equations (Eq. (3)), respectively. Then the molecular of polymer, M, can be calculated through Mark-Houwink Equation and Beer’s experimental results (Eq. (4)).31,32 Schott Gerate Ubbelohde viscometer (produced by Zhejiang Jiaojiang Co., Ltd., inner diameter of capillary is 0.6 mm, viscometer constant is 0.8~0.9 mm2/s2) was used to determine the intrinsic viscosity, [η], of guar gum. Absolute viscosity of sample, η, was measured contrasting the solvent viscosity, ηs, by the time of flowing through capillary. The filtrate sample collected from 2.3.1 Biodegradation experiment was equilibrated to 25℃(±0.1 °C) to analysis. 25 mL of fluid was used and tests were performed in triplicate.33 η sp

=[ η ]+ K ' [ η ]2 c

(2)

Inη r =[η ]-β [η ]2 c c

(3)

[η ]=5.13 × 10−4 M 0.72

(4)

c

Where ηr is the relative viscosity, defined as the ratio of solution viscosity and solvent viscosity, η/ηs; ηsp is the specific viscosity, defined as ηr-1; c is the concentration of guar gum, gdL-1; [η] is the intrinsic viscosity, dLg-1; K′ and β are Huggins and Kraemer constants, respectively. M is average molecular weight.


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2.3.4 Gas composition measurement. Lead acetate test paper (Chinese Medicine Group Chemical Reagent Co., Ltd., China) and gas chromatography (GC) were applied to test the gaseous products.34 The wet lead acetate test paper was placed in the gaseous products for 5 min. If the color of test paper changes into black or brown, this phenomenon indicates the existence of H2S. Otherwise, there is no H2S. Gaseous components were identified by GC analysis on a GC790A gas chromatograph (Agilent, USA) equipped with an on-column injector, an FID detector, an ECD detector, and a Porapak Q column (80-100 Mesh). Column temperature was set to 55 °C and nitrogen was used as gas carrier. A sample volume of 1mL was used. N2O was detected by ECD detector at 330 °C at the constant flow rate of 30 mL/min. CO2 and CH4 were analyzed by FID detector at 200 °C after nickel catalyst at 375 °C. The combustion gas was H2 at the flow of 45 mL/min, while the oxidant gas was air at the flow of 300 mL/min. 2.4 Insoluble residue degradation 2.4.1 Insoluble residue collection A glass beaker containing 1L 4.0 g/L guar gum solution was water-bathed at 80 °C, thereafter, 1.0 g ammonium peroxydisulfate (APS) was added and mixed round to dissolve. After keeping still for 1 hour, the viscosity of guar gum solution decreased and the white flocculent precipitate is the insoluble residue of guar gum. After centrifugation at 3000 r/min for 20 min, DI water was used to clean the precipitate. 10 mL supernatant was collected in a test tube and 0.2 mol/L BaCl2 was added dropwise into the test tube to test the SO42-. If there is no white precipitate in the test tube, the insoluble residue are cleaned absolutely and the precipitate in the centrifuge tube was the purified insoluble residue. 2.4.2 Biodegradation experiment 8.0 g insoluble residue was dissolved with 40 mL salt solution [(KNO3 2.0 g/L, (NH4)2SO4 2.0 g/L, K2HPO4 2.0 g/L, MgCl2 0.15 g/L, CaCl2 0.15 g/L); pH 7.0]. Then the suspension of insoluble residue was divided into two equal parts in two 50 mL


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plastic tubes (long 108mm, inner diameter 28 mm). 0.5 mL bacterial seed liquid of 2.3.1 Biodegradation experiment was added in one of the two tubes as the degradation treatment while the other tube was the control treatment without any inoculation. Finally, they were all incubated at 50 °C and 120 rpm in a water bath shaker. 2.4.3 Particle size measurement The shape of residue particle can be observed by microscope (XSP-8CA, Jiangnan photoelectric Co., Ltd., Shanghai, China) with the digital camera. Laser particle size analyzer was employed to measure the particle size of insoluble residue. Measurement range of size is 0.1~450 µm and laser wavelength is 600 nm. According to the instruction manual of the analyzer, 0.1mL suspension of insoluble residue was added into the quartz cuvette with 20~30 mL DI water. Background noise was cleared by the DI water with no sample. Data report of particle size distribution was outputted by related computer software of the analyzer. Average particle size (dz) shown as Eq. (5).35,36

dz =

d16 + d50 + d84 3

(5)

Where d16, d50 and d84 are particle sizes when cumulative percentages are 16%, 50% and 84% in the cumulative distribution curve. 2.4.4 Total weight measurement Tubes were centrifuged at 3000 r/min for 20 min and dried at 80℃ to constant weight and then the microbial degradation rate (Rmd, %) was calculated through the total weight of insoluble residue using the Eq. (6). Rmd =

w0 − w1 × 100% w0

(6)

Where, w0 and w1 are the weight of insoluble residue of control treatment and degradation treatment, respectively.


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2.5 Sand-pack column assays : Valve Pump 2

(B)

Core holder

(A)

Thermostat Pressure sensor

Culture liquid Computer Fracturing fluid

Pump 1

Measuring cylinder

Figure 1. Experimental flow for the microbial remediation of fracturing fluid damage 2.5.1 Equipments The main equipments and the experimental flow for microbial remediation were shown in Figure 1. Pump 1 was the 2PB00C constant-flux pump supplied by Beijing Satellite Factory, China. Other equipments included a hand sling pressure pump (Pump 2), two piston middle vessels, a Hassler core holder, two pressure sensors and a thermostat. They were all purchased from Petroleum Research Instrument Co., Ltd., Jiangsu, China. Proppant

Simulated the face between Simulated matrix fractures and matrix

Matrix Filter cake

Cemented sand zone

Filled sand zone

Acrylic tube Cemented sand zone

Figure 2. Schematic diagrams showing the structure of sand-pack column. 2.5.2 Design of sand-pack column The effectiveness of microbial remediation of fracturing fluid damage was tested in sand-pack columns which simulated the porous media of matrix with the surface between matrix and fracture after hydraulic fracturing. The sand-pack column used in this study was designed as shown in Figure 2. The sand-pack column was fabricated by acrylic


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(Polymethylmethacrylate, PMMA) tube. The out diameter (ϕout), inner diameter (ϕin) and length (l) of the sand pack, being similar to the size of Berea core, were 2.5 cm, 2.1 cm and 6.0 cm, respectively. The sand-pack columns were all packed with acid-washed quartz sands (large size quartz sands of 50/70 mesh; small size quartz sands of 100/110 mesh). Epoxy-resin glue (Shenzhen Junqi Chemical Co., Ltd., China) was used to bond the large size quartz sands at two ends of the column. The cemented sand zone can prevent the small size quartz sands in the filled sand zone from moving. The filled sand zone can simulate the realistic surface of porous media of the formation. Research results of Devine et al.37 suggested that the greatest post-treatment damage generally occurs in the intermediate permeability core, so three columns with initial permeability of approximately 50 mD were selected to test the remediation efficiency. Table 2 gives some key parameters of sand-pack columns. Table 2. Key Parameters of Sand-pack Column sand-pack

length

out diameter

inner diameter

pore volume

porosity

initial permeability

column

/cm

[ϕout]/cm

[ϕin] /cm

[PV] /cm3

[φ] /%

[k0] /×10-3 µm2

1

6.20

2.50

2.10

4.19

19.52

49.80

2

7.20

2.50

2.10

6.80

27.28

52.52

3

6.30

2.50

2.10

5.41

24.81

39.49

2.5.3 Experimental flow In this experiment, the flow of fluids in the core obeys Darcy law shown as Eq. (7). All the injection rates of Pump 1 were 0.2 mL/min and all permeability values were calculated in the steady state of pressure with DI water injection. The simulation experiments were performed as following: (1) Determination of the physical parameters of core: The sand core was saturated with DI water after vacuum pumping. The weight difference before and after saturation can be employed to calculated the porosity of core. In addition, the initial permeability, k0, can be calculated


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during DI water injection from (B) emulating formation matrix to (A) emulating the inlet well (shown as Figure 1). (2) Simulation of formation damage: Guar-based fracturing fluid, with 1.0 g/L ammonium persulphate as gel-breaker, was injected from (A) to (B) using Pump 1 with the injection rate of 0.5 mL/min. When the pressure was higher than 2 MPa, the injection was paused and when the pressure was lower than 2MPa, Pump 1 was started-up. If there was no fluids produced at the end of (B), the fracturing fluid injection step was finished. Then temperature of thermostat was turned up to 65 °C for 4 hours to realize adequately the function of breaker. Additionally, 1.0 pore volume (PV) of DI water was injected using Pump1 from (B) to (A) to simulate the flow-back and the damaged permeability, k1, was tested from (B) to (A). (3) Simulation of microbial remediation: 1.0 PV culture liquid of microbes was injected from (A) to (B) and microbial cultivation was conducted at 50 °C for 2 days. (4) Measurement of the effectiveness of remediation: 1.0 PV DI water was injected from (B) to (A) to simulate the second flow-back and the regained permeability, k2, was tested from (B) to (A). Then the damage rate (Rd, %) and regained rate (Rr, %) were described by Eq. (8) and Eq. (9).

k=

µ ×Q× A ∆P × L

×103

(7)

Rd =

k0 − k1 × 100% k0

(8)

Rr =

k2 ×100% k0

(9)

Where k, A, and L were the permeability (mD), sectional area (cm2) and length (cm) of sand pack column; µ and Q indicated the viscosity (mPa·s) and flow rate (cm3/s) of DI water; ∆P (1×105 Pa)


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was the differential pressure between two ends of sand pack column. 3. RESULTS AND DISCUSSION 3.1 Identification

XSJ

XSJ

M

Figure 3. Photos showing the PCR results and the phylogenetic relationship of strain XSJ by neighbour-joining tree based on 16S rDNA gene sequence. Guar gum degrading microbial consortia were enriched from the a formation water sample collected from oil wells in Shengli Oilfield in eastern China. Growth of the guar gum degrading bacteria was monitored in terms of the observation of bacterial lawn and cell density one such consortium designated as XSJ was selected for further studies as it showed maximum growth on the isolation medium. Analysis of 16S rDNA gene fragments performed to determine the composition of XSJ. One prominent band was identified as the phylogenetic affiliates of Bacillus sp. (>98%, 16s rDNA sequences homology with reference sequences in Gen-bank) as shown in Figure 3. The sequence was deposited in the Gen-bank database under the accession number of KY077256. From the phylogenetic tree in Figure 3 constructed based on 16S rDNA, strain XSJ was most similar to Bacillus sonorensis sp. nov. isolated from soil in the Sonoran desert of Arizona in America reported by Palmisano.38 Bacillus sonorensis39 was also used as plant


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growth promoting rhizo-bacterium and α-amylase or cellulose production bacterial stain40-42. However, this is the first report of Bacillus sonorensis in the guar gum degrading.

3.2 Characterization (a)

(b)

(a) beef extract-peptone ager

(b) starch salt ager medium

(c)

(d)

(c) guar gum salt ager, aerobic (e)

(d) guar gum salt ager, micro-aerobic (f)

20mm (e) CMC-Na salt ager, 50 °C

(f) CMC-Na salt ager, 55 °C

Figure 4. Colonial characteristics of strain XSJ on different ager media. CMC-Na was carboxymethylcellulose sodium. 0.05 mol/L iodine solution and 1.0 g/L Congo red solution were used as chromogenic agent in (b) and (e) (f), respectively.


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3.2.1 Carbon sources The growth of strain XSJ using the various carbon sources was investigated shown in Figure 4. Figure 4 (a) showed the distinctive colonial morphologies in beef-extract peptone ager medium. Colonies in Figure 4 (c) indicated that strain XSJ can grow in the aerobic environments utilizing the guar gum as the sole carbon source. Figure 4 (d) showed that strain XSJ can grow at the button of guar gum salt ager medium with the micro-aerobic environment. Degradation ring in Figure 4 (b) showed the capability of strain XSJ in starch degradation. Cellulose degradation rings in the Figure 4 (e) and Figure 4 (f) showed that XSJ also can grow using cellulose as the sole carbon source at 50℃ and 55℃. The insoluble residue of guar gum often contain impurity e.g. cellulose, starch, lignin. They account for approximately 3%43 depending on the isolation and modification technology of guar gum. Therefore, strain XSJ have the potential in degrading the impurity containing guar gum, cellulose or starch. 3.2.2 Growth curve. The growth and fitting curve of strain XSJ was showed in Figure 5. After fitting of the logistic curve, the microbiologically relevant parameters were showed in Table 3. The maximum specific growth rate [µm] in the guar gum salt medium is far less than in the beef-extract peptone medium. In addition, the lag time of guar gum salt medium is 8.71h far higher than the beef-extract peptone medium of 2.52h. The lag phase is related to the synthesis of induced enzyme to degrade the substrate. It is means that the substrate utilization of guar gum was conducted by induced enzyme. However, the maximal value of bacterial number was equal approximately, which means that guar gum is good carbon source for the growth of strain XSJ.


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8

In (N/N0)

7

8

(b)

(a)

7

6

6

5

5

4

4

3

3

2

Beef-extract peptone

Guar gum

1

In (N/N0)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Energy & Fuels

2 1

0

0 0

2

4

6 Time /h

8

10

12

0

10

20

30 Time /h

40

50

60

Figure 5. Growth and fitting curves of strain XSJ in the beef-extract peptone medium (a) and guar gum salt medium (b). Table 3. Parameters of Growth Curves of Strain XSJ. medium maximum specific growth rate (µm)

lag time (λ)

maximal value (A)

beef-extract peptone

13.15

2.52

7.30

guar gum salt

1.57

8.71

7.43

3.2.3 Salinity Strain XSJ can also grow under a wide range of salinity (

Application of Guar Gum Degrading Bacteria in Microbial

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