Manuscript 107501-0606 IJCEE-Biosurfactant Produced by Azotobacter Vinelandii

8/9/2019 Manuscript 107501-0606 IJCEE-Biosurfactant Produced by Azotobacter Vinelandii

1/8

International Journal of Civil & Environmental Engineering IJCEE Vol: 10 No: 1

Application of Biosurfactant Produced byAzotobacter vinelandii AV01 for

Enhanced Oil Recovery and Biodegradation of Oil Sludge

Qomarudin Helmy, Edwan Kardena, Zeily Nurachman and Wisjnuprapto

Abstract - The problem of petroleum waste managementis giving a due consideration of the national level. Largequantity of dehydrated oil sludge, generated in thedisposal process of oil-containing sewage in Indonesiathat needs to be rendered harmless to human and to theenvironment. Microbial degradation has been accepted asan important method for the treatment of oil sludge byemploying indigenous or extraneous microbial flora. Thepurpose of this study was to investigate the performanceof biosurfactant in its attempt to recover oil from oilsludge and in enhanced biodegradation of oil sludge process. Measurement of biosurfactant productionindicated that the maximum production occurred at theend of exponential growth phase (48h). The

emulsification capacity of the biosurfactant also stablesunder thermal and ionic strength treatment that meet anyrequirement for application in the oil recovery anddegradation. In the oil recovery preliminary test,biosurfactant have the capability to recover oil up to 15%from oil sludge. For oil sludge biodegradation assay, itwas found that addition of petrofilic consortia increasedthe removal efficiency up to 55%, while addition ofbiosurfactant in this reactor increased the total efficiencyof 70% after 70 days of incubation. These results suggestthat both petrofilic consortia and biosurfactant additionstimulate the biodegradation and overcome the limitationof petroleum hydrocarbon degradation process.

Keywords: A. vinelandii, biosurfactants, biodegradation,oil sludge, oil recovery, crude oil

1. INTRODUCTIONOil, being an essential energy source, is both the

lifeblood and a liability of many industrializednations. The use of crude oil as an energy source hasallowed many nations to develop a high standard ofliving. Continued economic growth will increase thedemand for oil, which must be met by current production technologies or by new discoveries.Traditional oil recovery technologies under theumbrella of chemically enhanced oil recovery

(CEOR) can recover a maximum of 4045% of theoil initially in place, in two stages, namely, primaryand secondary recovery[1, 2].

Manuscript received Januari 9, 2010. This work was supported inpart by ITB Research Grant and Directorate general of higher education(DIKTI), Government of Indonesia for the project grant

No:324/SP2H/DP2M/III/ 2008 in environmental biotechnology.Q. Helmy, E. Kardena and Wisjnuprapto is member of the Water

and Wastewater Engineering Research Group, Faculty of Civil andEnvironmental Engineering, Institut Teknologi Bandung (ITB)Indonesia. Ganesha 10 Bandung-West Java. Tel. No. +62 817 023 5878,[email protected]; [email protected]; [email protected].

Z. Nurachman is member of the Biochemistry Research Group,Faculty of Mathematics and Natural Sciences in the same Institute,[email protected]

One of the most encountered pollutants in oilproduction companies is the formation of oil sludgethat is entrapped with the effluents during treatmentand conditioning of the wells produced crude oilthrough treatment process facilities. Most of the oilsludge is piled up outdoor without any treatment,and poses a serious environmental problem. Thehydrocarbons in the sludge penetrate from the topsoil into the subsoil slowly, presenting a direct riskof contamination to subsoil and groundwater [3-5].On the other hand, the light hydrocarbons in the oilsludge vaporize, leaving behind a layer of oil-containing dust of soil which blows upwards to

pollute the air. Therefore, the oil sludge should betreated to prevent harm to environment. Although burning of the sludge may be simple and easilyadaptable, this technique has undesirable hazard inair pollution. Cleaner technologies are needed due tothe environmental friendly such as microbialdegradation concept [6-9].

The oil sludge is attributed to two major factorscontrolling in its formation. First is the inorganicresidue consisting of sediments, sands, scales, dustand the second is the precipitation of paraffinic wax.Since wax precipitates are sparingly soluble in crudeoil, temperature changes are the reason behind wax precipitation. In addition to the above reasons, theoxidation of heavy organic material in crude oil dueto climate changes or from oxidizingmicroorganisms and also the interruption in materialbalance due to losses of volatile components and thetendency of asphaltene, resin and polymericcompounds to precipitate all are the reasons of oilsludge formation.

As the oil price rised during year 2008, attemptto recover oil from oil sludge became advantageous both from economic and environmentally point ofview. Production of oil sludge as oil refinery by-

product in Indonesia exceeding 2,000 ton.day

-1

. Thishuge oil sludge production refer to environmentalproblem because of its characteristics and handling.Many technologies are employed to clean upcontaminated sites including various chemical and physical methods such as excavation, thermalevaporation and soil vapor extraction.Bioremediation has been accepted as an importantmethod for the treatment of oil sludge by employingindigenous or extraneous microbial flora. Undercertain conditions, living microorganisms primarily bacteria can metabolize various classes ofhydrocarbons compound [10]. Since hydrocarbons


2/8


contain high organic matter, it is not surprisingly if itcan be used as a carbon source by microorganism.The microorganisms in this process are effective inoxidizing the dissolve organic compounds, includingsome emulsified oil [11]. One of the importantfactors in oil biodegradation rate is its solubility.This related to the bioavailability of contaminant to

the microbial attack. One promising approach toincreasing the biodegradation rates of organiccompounds with limited water solubility is theaddition of biosurfactant [12, 13].

In this paper, we report the possible applicationof biosurfactant produced from A. vinelandii in theoil industry, investigated the performance of biosurfactant in its attempt to recover oil from oilsludge and in enhanced biodegradation of oil sludgeprocess.

2. MATERIAL AND METHODS

2.1. Reagents.All chemicals were of reagent grade, purchased

from Merck, J.T. Baker and Sigma Chem Co.Growth media were purchased from Oxoid ltd.

2.2. Crude Oil and Oil Sludge.Crude oil and oil sludge samples was obtained

from Duri Oil Field Pekanbaru and Balongan OilField Indramayu Indonesia, respectively.

2.3. Bacterial strain and Culture Conditions. Azotobacter vinelandii AV01 was used in

producing biosurfactant, while the petrofilicconsortia containing Bacillus cereus BL01,

Pseudomonas stuzeri BL02, Acinetobacter sp.BL03 and Bacillus sp BL04 were used in thebiodegradation assay of oil sludge. All bacteria wereobtained from the Culture Collection ofEnvironmental Biotechnology Laboratory-Environmental Engineering Department, InstituteTechnology of Bandung, Indonesia. A. vinelandiiwas maintained at 4C on mannitol enrichment agarslants containing (l-1): 20 g mannitol, 20 g yeastextract, 20 g tryptone, and 15 g of agar. While eachpetrofilic bacteria was maintained at 4C on NutrientAgar covering with 1 drop of crude oil. Sub-cultureswere made to fresh agar slants every 1 month to

maintain viability.

2.4. Biosurfactant Production.Cultures of A. vinelandii were grown on a

minimal basal medium (MB) which composed thefollowing components (l-1) of distilled water: 1.5 gof K2HPO4; 0.5 g of KH2PO4; 0.2 g of MgSO4; 0.25g of (NH4)2 SO4; and 20 g glucose as substrate. 10ml Trace Element solution was added per liter ofMB medium. The compositions of this trace element(l-1) are 12 g of Na2EDTA2.H2O; 1 g of CaCl2; 0.4 g

of ZnSO4.7H2O; 10 g of NaSO4; 0.4 g ofMnSO4.4H2O; 0.1 g of CuSO4.5H2O; 0.5 g ofNa2MoO4.2H2O [14]. The medium was sterilized byautoclaving at 121C for 15 min. The inoculums of A. vinelandii was prepared by transferring cellsgrown on a slant to 250 ml Erlenmeyer flaskscontaining 50 ml of MB broth. Culture was

incubated in an orbital shaker at room temperature,110 rpm for 2 days. The MB containing 106 cells/mlwas used to initiate growth using 2% (v/v)inoculums. Biosurfactant production was carried outin a 10 liter capacity of fermentor at 37oC withagitation speed of 100 rpm and aeration rate of 0.176ft3.min-1 for 2 days.

2.5. Crude Biosurfactant IsolationThe fermentation broth was centrifuged at

13.000 rpm for 30 minute to obtain a cell free broth.After centrifugation, the supernatant was thendissolved in a 4 N hydrochloric solution and allowed

to stand overnight at 4C, followed by the biosurfactant extraction step with a chloroformsolvent at room temperature [15]. The organic layerwas transferred to a round-bottom flask and theaqueous layer was re-extracted two times forcomplete recovery of biosurfactant. The organic phases were combined yielding a viscous brown-colored crude biosurfactant product and thenevaporated to remove the solvent; the residue wascollected and weighted. Vermani [16] method wasused to determine the exopolysaccharide fraction of biosurfactant. A mixture of 1:2 (v/v) biosurfactantand chilled acetone were agitated and stand

overnight to precipitate. Formed precipitate werefiltered and gravimetrically analyzed.

2.6. Emulsification Index (E24).To determine the emulsification index, Batista

[17] method was applied. Centrifugation at 13,000rpm to separate biosurfactant from microorganismcells yielding a cell free broth. A mixture of 1:1 between biosurfactant and crude oil is agitated forabout 2 minute then stabilized for 24 hour.Emulsification index (%) determined by measuringthe column height of emulsified oil against its totalheight multiplied by 100 times.

2.7. Emulsification Stability TestStability studies as described by [18], were done

using the cell free broth obtained by centrifugationprocess as mentioned above.Effect of extreme temperature. Five milliliter of cellfree broth was exposure with various temperatureconditions: 4oC, 27oC and 120oC for 30 min andallowed to cool to room temperature, then theemulsification index were measured using describedprotocol.


3/8


Effect of extreme ionic strength. To study the pHstability of biosurfactant, five milliliter of cell freebroth were exposure with various pH environments:pH value between 2-12.

2.8. Oil Recovery from Oil SludgeAdapting the soil washing method from [19]

with some modification, 1:1 ratio of oil sludge andbiosurfactant poured into 10 ml centrifuge test tubes.The test tube was then shaken with vortex for 5minute and allows settling. After that the test tubewas centrifuged at 6,000 rpm for 15 minute yielding3 phases of oil emulsion, remaining oil sludgematrix and liquid solution. Oil emulsion recoveredwas then measured in volume unit.

2.9. Total Petroleum Hydrocarbon (TPH)Measurements.

Measurement of TPH was conducted withgravimetric method as described by [20]. Samplewas extracted with n-hexane, the organic layer werepooled and dried by evaporation of solvents. Afterevaporation, the amount of residual TPH recoveredwas weighted.

2.10. Biodegradation AssayTo determine the performance of petrofilic bacteriain degrading oil sludge, a preliminary biodegradation assay developed and set up asfollows:

Control-l: without addition of both petrofilicinoculums/P and biosurfactant/B.

Control-2: without P; add 2% (v/v) B. Reactor-1: add 2% (v/v) P; without B. Reactor-2: add 2 % (v/v) each of P and B. Oil sludge initial concentration was 100.000

ppm of TPH and incubation time forbiodegradation assay was 70 days.TPH concentration and growth of petrofilic

bacteria were observed on certain time.

3. RESULT AND DISCUSSION

3.1 Biosurfactant ProductionThe reason we chooseA. vinelandii are first,A.

vinelandii is a rhizosphere bacterial strain that ableto produce biosurfactant constitutively from watersoluble substrate without any addition of inducersubstrate; second, A. vinelandii known have the

ability to utilized free nitrogen from the air; andthird, the ability to produce biosurfactant from watersoluble substrates is preferred because single-phasefermentation is simpler than biphasic fermentationthat usually occurred when hydrocarbon basedsubstrates were applied. Growth and biosurfactant production from A. vinelandii with 2% glucose assole carbon source was described in the Figure 1.

Fig 1. Growth (solid circle symbol), biosurfactant production (open circle symbol) and emulsificationactivity (solid square symbol) profiles ofA. vinelandii grown in fermentor with 2% (v/v) glucose as a carbonsource at 37oC.

The biosurfactant production started to increaseduring the exponential phase, reaching its maximumafter about 48 h (9.81 g/l). These results indicate thatthe maximum of biosurfactant biosynthesis fromglucose occurred predominantly at the end of theexponential growth phase. The emulsification

activity of the cell free broth increased up to 90% inthe first 24 hour of incubation, whereas surfactantaccumulation increased during this period and startto decrease after reaching its maximum synthesis.This might be due to biosurfactant were used ascarbon source by A. vinelandii. Similar result

0

10

20

30

40

50

60

70

80

90

100

EmulsificationIndex

(%

)

0

2

4

6

8

10

12

14

16

0

0.2

0.4

0.6

0.8

1

1.2

1.4

1.6

0 20 40 60 80 100

Biosurfactantproduction

(g/L)

Biomass(g/L)

Incubation Time (hour)


4/8


reported by [18], that grown C.lipolytica with 10%canola oil and 10% glucose. Biosurfactantconcentration reaching its maximum production after48h at the end of exponential phase and start todecrease in a longer incubation time. While [21],used the molasses and corn steep liquor as the primary carbon and nitrogen source to produce

rhamnolipid biosurfactant from Pseudomonasaeruginosa GS3. Maximum surfactant productionoccurred after 96h of incubation, when cells reachedthe stationary phase of growth.

3.2. Emulsification Stability Test: effect oftemperature and pH

Interest in biosurfactant is growing due to theirapplications in the protection of the environment andin the oil industry. Their environmental uses are principally to the bioremediation of petroleumhydrocarbon contaminated sites [22]. In the oilindustry, they are used in microbial enhanced oilrecovery (MEOR) and or biosurfactant-mediatedenhanced oil recovery, facilitate transportation ofheavy crude oil through pipelines and in the cleaningof contaminated vessels [1]. Application ofbiosurfactant in enhanced oil recovery process mustmeet any requirement involving the extremecondition of oil reservoir [23]. These processesfrequently involve exposure to extremes oftemperatures, pressure, salinity, pH and organicsolvents. Stability studies were done using cell-free broth obtained by centrifugation the culture at13.000 rpm for 30 minute as described in methods.

The emulsification index of the culture brothfree of cells was stable when stored at 4oC either atroom temperature (27o C). It is interesting to observethat the emulsification capacity of the biosurfactantremain stable after heating for 30 minutes up to 1200

C. The effect of thermal treatment (chilled/heated)on the activity of the biosurfactant from A.vinelandii cultivated in minimum basal medium with2% glucose as carbon source showed that noappreciable changes in emulsification capacityoccurred. Similar result reported by [18] by growingC. lipolytica on canola oil and glucose to producebiosurfactant that reach its maximum emulsification

activity at high temperature (120o

C). In contrary,liposan from C. lipolytica was found to be relativelystable between 30oC and 90oC, but lost almost 60%of its activity after boiling for 1 h [24]. The effect ofextreme temperature and pH on the emulsificationactivity of the culture broth free of cell can be seenin Figure 2. Extreme of pH could possibly transformless surface-active species into more activeemulsifiers by denaturation of proteinaceouscomponents or by increased ionization. The

effectiveness of biosurfactant produced from A.vinelandii was pH stable at pH range 2-8.

0

20

40

60

80

100

4 27 120

E24(%)

Temperature(oC)

A

50

60

70

80

90

100

2 6 8 12

E24(%)

pH

B

Fig 2 Effect of temperature (a) and pH (b) on theemulsifying activity of cell free broth ofA.vinelandii grown in fermentor with 2% (v/v) glucoseas a carbon source at 37oC.

3.3. Oil recovery from oil sludgeFor many countries like Indonesia, domestic oil

production is in decline and the likelihood ofdiscovering neither large nor new oil reserves is low.These countries must then rely on foreign imports,which can slow economic growth and employmentand aggravate trade deficits. As the oil price risedduring year 2008, attempt to recover oil from oilsludge became advantageous both from economicand environmentally point of view. Production of oilsludge as oil refinery by-product in Indonesiaexceeding 2,000 ton.day-1. Economically attractive because approximately 15% (see Table 1) of oilcould be recovered from oil sludge. Alsoenvironmentally attractive because the remaining oilsludge became less in the oil content thus make iteasier to handle with. However, the low oil pricesthat prevailed during the end of year 2008 lead to a


5/8


shift in focus from enhanced oil recovery toenhanced oil remediation processes.Table 1 showedthe properties of oil sludge and the experiments parameter. The most successful attempt in usingbiosurfactant to recover oil from oil sludge reported by [25]. The clean-up process was successful inremoving the sludge from the tank bottom, and it

also allowed the recovery of more than 90% of thehydrocarbon trapped in the sludge. The recoveredhydrocarbon had excellent properties and could besold after being blended with fresh crude. Therecovered 774 m3 (5550 barrels, at US $60/barrel)of oil is worth $333,000.

TABLE 1Properties of oil sludge and biosurfactant enhancedoil recovery from oil sludge experiment parameters.

Parameter Value

Oil sludge propertiesOil sludge density at 25oC (kg/m3)Viscosity at 25oC (centipoises)Oil content (%)

Experiment parameters

Volume of oil sludge (ml)Volume of biosurfactant (ml)Conc. of biosurfactant (g/l)Shaking time (min)Centrifuge speed (rpm)Centrifuge time (min)Oil recovered (ml)1Oil recovered (%)2

867.845030-33

559.8156000150.75

15

1

means values from triplicate measurement.2 oil in emulsion

3.4. Biosurfactant Enhanced Biodegradation ofOil Sludge

Biosurfactant are also used in emergingtechnologies like microbial remediation ofhydrocarbon and crude oilcontaminated soils.Hydrocarbon contaminants are removed from the

environment, primarily as a result of their biodegradation, which is performed by nativemicrobial populations. Such biodegradation isknown to be time-consuming and new technologieshave been developed; for example the addition of biosurfactant help to stimulate the indigenousmicrobial population to degrade hydrocarbons atrates higher than those which could be achievedthrough addition of nutrients alone. Biosurfactant isa well known surface active agent that generallyused in improving the viability of contaminant to themicrobial attack.

The biosurfactant affect the biodegradationprocess by increasing the solubility and dispersion ofthe compound [26]. There are two ways in whichbiosurfactant affect which is increasing the surfacearea of hydrophobic water insoluble substrate.Secondly is increasing the bioavailability ofhydrophobic water-insoluble substances. Alaboratory scale of biosurfactant enhancedbiodegradation of oil sludge was conducted. Effectsof addition of biosurfactant fromA. vinelandii in thebiodegradation process were shown in Table 2. Weused crude biosurfactant in the form of cell free broth directly without purifying the biosurfactant

first for the simplicity reason of the experiments.

TABLE 2TPH removal efficiency of oil sludge biodegradation in batch reactor. C1/Control 1 without addition bothPetrofilic inoculums/P and Biosurfactant/B; C2/Control 2 (P, +B); R1/Reactor 1 (+P, B); R2/Reactor 2

(+P, +B).

Biodegradation system TPH Removal Efficiency (%)1 Increased Removal Efficiency (%)

Oil sludge biodegradation

C1 12.4 6.4 (C1-C2)C2 18.8 55.6 (C1-R1)R1 68.0 20.9 (R1-R2)R2 88.9 70.1 (C2-R2)

1 means values from triplicate measurement.

The low water-solubility of many hydrocarbonsreduces their availability to microorganisms andlimits the biodegradation process. It has beenassumed that biosurfactant can be used to enhancethe bioavailability of hydrophobic compounds. Onthe other hand this low water-solubility increasessorption of compound to surface and limits their

availability to biodegrading microorganisms [27].Once again, biosurfactant can enhance growth onbound substrates by desorbing them from surfaces orby increasing their apparent water solubility. Figure3 showed the microbial growth and TPH profile ofcontrol reactor. It was noticed that changes in oilsludge environmental condition from its originally


6/8


slurry phase into more aqueous phase in the reactor,triggering the indigenous bacteria in it to grow. Forthe total plate count measurement, the CFU valuesincreased from 103.2 (CFU/ml) at day 0 and reach itsmaximum to 105.4 (CFU/ml) in the first week ofincubation. Similar pattern occurred in the controlreactor-2 (without addition of petrofilic inoculants,

added by 2% v/v of biosurfactant only).Biosurfactant addition make the oil sludge becomemore soluble in the reactor, this shown by increasein the microbial growth from 103.2 (CFU/ml) at day0 and reach its maximum to 106 (CFU/ml) in thefirst week of incubation. However, the degradationprocess of oil sludge by mean of indigenous bacteria

predicted small enough/neglect able throughout theexperiment. TPH losses in control reactors mainlydue to weathering/physical influences [28, 29] suchas temperature shift, shaking condition,volatilization of low molecular weight ofhydrocarbon [30] and photo-oxidation [31]. Thisphenomenon confirmed the results reported by [32],

who observed disappearance of diesel compound inthe absence of biological activity reached 11% after45 days of incubation. While addition of syntheticsurfactant and biosurfactant in the reactor increasedthe oil sludge degradation rate by indigenous bacteria to 12 and 21% after 60 days incubation,respectively [33].

3.0

3.5

4.0

4.5

5.0

5.5

6.0

6.5

7.0

7.5

8.0

0.0E+00

2.0E+03

4.0E+03

6.0E+03

8.0E+03

1.0E+04

1.2E+04

0 7 14 21 28 35 42 49 56 63 70

LogCFU

ml1

TPHofoilsludge

(pp

m)

Time(day)

C1 C2 BC1 BC2

Fig 3. The indigenous microbial growth (open symbol) and TPH degradation (solid symbol) profiles inbatch control reactor system with 0 (circle/C-1) and 2 % v/v (square/C-2) of biosurfactant addition at 27oC.

Figure 4 shows that after 70 days of incubation,a significant reduction of TPH (68%) occurred in thebiodegradation system supplemented with petrofilicconsortia/R-1. This positive result suggests that bio-augmented bacteria could degrade TPHsignificantly. Bioaugmentation also can be used toincrease the biodegradative capabilities of theindigenous microbial population. Compared withcontrol reactor/C1, addition of petrofilic consortia

increased the removal efficiency up to 55%. Nonbiological degradation (physical transformation) alsooccurred in the process; however the biologicaltransformation dominated the process based on thegrowth of bacteria observed during the process. Forthe total plate count measurement, the CFU values

increased from 106 (CFU/ml) at day 0 to 107.5 and106.7 (CFU/ml) in the first week and day 70respectively. The presence of biosurfactant inbiodegradation system (R-2) increased the removalefficiency up to 20% compared to those withoutaddition of biosurfactant/R-1. The present of biosurfactant also increased the microbial growthfrom 106.5 (CFU/ml) at day 0 to 108.8 (CFU/ml) inthe first week of incubation and 108.1 (CFU/ml) at

day 70. Similar result by [34], that examined theeffect of rhamnolipid biosurfactant to diesel/waterdegradation from 0 to 80 mg/l significantly increases biomass growth and diesel biodegradation percentage from 1000 to 2500 mg VSS/l and 40-100%, respectively.


7/8


6.0

6.5

7.0

7.5

8.0

8.5

9.0

0.0E+00

2.0E+03

4.0E+03

6.0E+03

8.0E+03

1.0E+04

1.2E+04

0 7 14 21 28 35 42 49 56 63 70

LogCFU

ml1

TPHofoilslud

ge

(ppm)

Time(day)

R1 R2 BR1 BR2

Fig 4 The petrofilic consortia growth (open symbol) and TPH degradation (solid symbol) profiles in batch

reactor system with 0 (circle/R-1) and 2 % v/v (square/R-2) of biosurfactant addition at 27

o

C.

Our findings show that the addition of both petrofilic consortia and biosurfactant favors the biodegradation of the oil sludge. The limitingcondition in the degradation of hydrocarbon andother PAH is their insolubility, thus decreasing theefficiency and rate of degradation. This limitationcan be overcome either by addition of surface-activecompounds surfactant to the growing culture, thusmaking hydrocarbons more water-soluble andavailable for the cell to degrade, or by production ofits own surfactant by the augmented organisms to

facilitate uptake. The presence of biosurfactant alsostimulate the catabolism of hydrocarbon by mean ofco-metabolism process since biosurfactant areorganic compound and readily degradable tomicroorganism.

4. CONCLUSIONReferring to the Indonesian Ministry of

Environmental (regulation no. 128/2003), it wasmandatory to manage a safe treatment of thesewastes (oil sludge) and also disposed off in anenvironmental friendly manner. It is regulated thatthe final concentration of TPH must less than 10.000

mg/kg. Based on the data presented in this paperindicate that the petrofilic consortia have thecapability to degrade oil sludge to comply with theregulation above.

ACKNOWLEDGMENTQ. Helmy thank S. Hidayat for contributing in partof the research. The authors also gratefullyacknowledge Prof. Naoyuki Funamizu, Laboratoryof Engineering on Sustainable Sanitation, Hokkaido

University for facilitating QH in the researchfellowship program in Japan.

REFERENCES

[1] Finnerty, W.R. 1992. Microbial enhanced oilrecovery. Genetic Eng. and BiotechnologyMonitor. 32: 36 43.

[2] Sen, R. 2008. Biotechnology in petroleum recovery:The microbial EOR. Progress in Energy andCombustion Science, Vol.34, Issue 6: 714-724

[3] Banat, I.M., R.S. Makkar and S.S. Cameotra. 2000.

Potential commercial applications of microbialsurfactants. Appl Microbiol Biotechnol, 53:495508

[4] Kanaly, R.A and S. Harayama. 2000. Biodegradationof high-molecular weight polycyclic aromatichydrocarbons by bacteria. J Bacteriol,182:20592067

[5] Marin, J.A., T. Hernandez and C. Garcia. 2005.Bioremediation of oil refinery sludge bylandfarming in semiarid conditions: Influence onsoil microbial activity. Environmental Research,Vol. 98: 185195.

[6] Barkay T, Venezi SN, Ron EZ, & Rosenberg E. 1999.Enhancement of Solubilization and

Biodegradation Of Polyaromatic Hydrocarbonsby the Bioemulsifier.J. Appl. Environ. Microbiol65: 2697-2702.

[7] Christofi, N. & I.B. Ivshina. 2002. Microbialsurfactants and their use in field studies of soilremediation.J. App. Microbiol. 93 : 915-929.

[8] Maier, R. M., and G. Soberon-Chavez. 2000.Pseudomonas aeruginosa rhamnolipids: biosynthesis and potential applications.Appl.Microbiol. Biotechnol. 54: 625-633.

[9] Ivshina, I. B., M. S. Kuyukina, and N. Christofi. 1998.Oil desorption from mineral and organic


8/8


materials using biosurfactant complexes produced by Rhodococcus species. World J.Microbiol. Biotechnol. 14: 711-717.

[10] Al-Tahhan, R. A., T. R. Sandrin, A. A. Bodour, andR. M. Maier. 2000. Rhamnolipid-inducedremoval of lipopolysaccharide fromPseudomonas aeruginosa: effect on cell surface

properties and interaction with hydrophobicsubstrates. Appl. Environ. Microbiol. 66: 3262 -3268.

[11] Ron, E. Z., and E. Rosenberg. 2001. Natural roles ofbiosurfactants.Environ. Microbiol. 3: 229-236.

[12] Xu, R and J.P. Obbard. 2004. Biodegradation of polycyclic aromatic hydrocarbons in oil-contaminated beach sediments treated withnutrient amendments. Journal Environ. Qual.33:861-867

[13] Zhang, G., Y. Wu., X. Qian and Q. Meng. 2005.Biodegradation of crude oil by Pseudomonasaeruginosa in the presence of rhamnolipids. JZhejiang Univ Sci B., 6(8): 725730.

[14] Helmy, Q., E. Kardena and Wisjnuprapto. 2009.Performance of Petrofilic Consortia and Effect ofSurfactant Tween 80 Addition in the Oil SludgeRemoval Process. J. Appl Sci Environ Sanit.Vol.4: 3

[15] Makkar, R.S. and S.S. Cameotra. 1998. Productionof biosurfactant at mesophilic and thermophilicconditions by a strain ofBacillus subtilis, J IndMicrobiol Biotechnol 20:4852.

[16] Vermani, M.V., S.M. Kelkar & M.Y. Kamat. 1995.Novel Polysaccharide Produced by A. vinelandiiIsolated from Plant Rhizosphere. Biotechnol.Letters, 17: 917-920.

[17] Batista.S.B., A.H. Mounteer, F.R. Amorim, & M.R.Totola. 2006. Isolation and characterization of

biosurfactant/ bioemulsifier-producing bacteriafrom petroleum contaminated sites. BioresourceTechnology. 97: 868875.

[18] Sarubbo, L.A., C.B.B. Farias., & G.M.C. Takaki.2007. Co-Utilization of Canola Oil and Glucoseon the Production of a Surfactant by Candidalipolytica. Current Microbiology, 54: 6873

[19] Urum, K., T. Pekdemir and M. Copur. 2005.Screening of Biosurfactants for Crude OilContaminated Soil Washing. J. Environ Eng andSci, Vol.4 No.6: 487-496

[20] Mishra, S., J. Jyot., R.C. Kuhad and B. Lal. 2001.Evaluation of inoculum addition to stimulate insitu bioremediation of oily-sludge contaminated

soil. Applied and environmental microbiology,67, (4): 1675-1681.

[21] Patel, R.M. and Desai, A.J. 1997. Biosurfactant production by Pseudomonas aeruginosa GS3from molasses.Lett. Appl. Microbiol., 25: 91-94.

[22] Park, J., C. Vipulanandan., J.W. Kim and M.H. Oh.2006. Effects of surfactants and electrolytesolutions on the properties of soil.Environ Geol,49: 977989

[23] Rosenberg, E. 1986. Production of biosurfactant tofacilitate residual oil mobilization. Crit. Rev.Biotechnol. 3: 109 132.

[24] Cirigliano, M.C and G.M. Carman. 1984. Isolation ofa bioemulsifier from Candida lipolytica. ApplEnviron Microbiol, 48:747750.

[25] Banat, I.M., N. Samarah., M. Murad., R. Horne andS. Banerjee. 1991. Biosurfactant production anduse in oil tank clean-up. World J. Microbiol.Biotechnol, 7: 80-88

[26] Desai, J.D & I.M. Banat. 1997. Microbial Productionof Surfactants and Their Commercial Potential.Microbiology and Molecular Biology Reviews, 6(1): 4764.

[27] Abalos, A., M. Vinas., J. Sabate., M.A. Manresa andA.M. Solanas. 2004. Enhanced biodegradation ofCasablanca crude oil by a microbial consortiumin presence of a rhamnolipid produced byPseudomonas aeruginosa AT10.Biodegradation15: 249-260

[28] Eweis, J., B. Ergas., J. Sarina., D, Chang., P.Y.Schroeder., D. Edward. 1998. BioremediationPrinciples. McGraw-Hill Inc. USA.

[29] Venosa, A.D and X. Zhu. 2003. Biodegradation ofCrude Oil Contaminating Marine Shorelines andFreshwater Wetlands. Spill Sci. Technol.Bulletin, 8: 163-178

[30] Swannell, R.P.J., K. Lee and M. McDonagh. 1996.Fields Evaluations of Marine Oil SpillBioremediation.Microbiology Rev. 60: 342-365.

[31] Atlas, R.M and R. Bartha. 1998. Microbial Ecology.Fundamental and Applications. BenjaminCummings Sci. Pub., CA, pp. 556-598.

[32] Gallego, J.L.R., J. Loredo., J.F. Llamas., F. Vazquezand J. Sanchez. 2001. Bioremediation of diesel-contaminated soils:Evaluation of potential in situtechnique by study of bacterial degradation.Biodegradation 12: 325-335.

[33] Rocha, C and C. Infante. 1997. Enhanced oily sludge

biodegradation by a tension-active agent isolatedfrom Pseudomonas aeruginosa USB-CSI. Appl.Microbiol. Biotechnol 47: 615-619

[34] Whang, L.M., P.W.G. Liu., C.C. Ma and S.S. Cheng.2007. Application of biosurfactant, rhamnolipid,and surfactin, for enhanced biodegradation ofdiesel-contaminated water and soil.J. HazardousMat. doi:10.1016/j.jhazmat.2007.05.063

Q. Helmy is a researcher in Institut Teknologi Bandung(ITB), Indonesia. He received his BSc in Biology (2001),Master in Environmental Engineering (2006). Currently,he is a Doctorate Student at ITB. His research areas

include environmental biotechnology. He is a member ofInternational Water Association (IWA) and theIndonesian Society for Microbiology, and receivedresearch sandwich program from Hokkaido University,Japan.

Documents

Manuscript 107501-0606 IJCEE-Biosurfactant Produced by Azotobacter Vinelandii