Bacterial biosurfactant increases ex situ biodiesel bioremediation in clayey soil

The contamination of soils by oily compounds has several environmental impacts, which can be reversed through bioremediation, using biosurfactants as auxiliaries in the biodegradation process. In this study, we aimed to perform ex situ bioremediation of biodiesel-contaminated soil using biosurfactants produced by Bacillus methylotrophicus. A crude biosurfactant was produced in a whey-based culture medium supplemented with nutrients and was later added to biodiesel-contaminated clayey soil. The produced lipopeptide biosurfactant could reduce the surface tension of the fermentation broth to 30.2 mN/m. An increase in the microbial population was observed in the contaminated soil; this finding can be corroborated by the finding of increased CO₂ release over days of bioremediation. Compared with natural attenuation, the addition of a lower concentration of the biosurfactant (0.5% w/w in relation to the mass of diesel oil) to the soil increased biodiesel removal by about 16% after 90 days. The added biosurfactant did not affect the retention of the contaminant in the soil, which is an important factor to be considered when applying in situ bioremediation technologies.

     

Simultaneous phenanthrene and cadmium removal from contaminated soil by a ligand/biosurfactant solution

Surfactants and inorganic ligands are pointed as efficient to simultaneous removal of heavy metals and hydrophobic organic pollutants from soil. However, the biosurfactants are potentially less toxic to soil organisms than other chemical agents. Thus, in this study the efficiency of combinations of iodide (I⁻) ligand and surfactants produced by different bacterial species in the simultaneous removal of cadmium (Cd²⁺) and phenanthrene in a Haplustox soil sample was investigated. Four microbial surfactants and the synthetic surfactant Triton X-100 were tested with different concentrations of ligand. Soil samples contaminated with Cd²⁺ and phenanthrene underwent consecutive washings with a surfactant/ligand solution. The removal of Cd²⁺ increased with increased ligand concentration, particularly in solutions containing biosurfactants produced by the bacterial strains Bacillus subtilis LBBMA155 (lipopeptide) and Flavobacterium sp. LBBMA168 (mixture of flavolipids) and Triton X-100. Maximum Cd²⁺ removal efficiency was 99.2% for biosurfactant produced by Arthrobacter oxydans LBBMA 201 (lipopeptide) and 99.2% for biosurfactant produced by Bacillus sp. LBBMA111A (mixed lipopeptide) in the presence of 0.336 mol iodide l⁻¹, while the maximum efficiency of Triton X-100 removal was 65.0%. The biosurfactant solutions removed from 80 to 88.0% of phenanthrene in soil, and the removal was not influenced by the presence of the ligand. Triton X-100 removed from 73 to 88% of the phenanthrene and, differently from the biosurfactants, iodide influenced the removal efficiency. The results indicate that the use of a single washing agent, called surfactant-ligand, affords simultaneous removal of organic contaminants and heavy metals.     

Physicochemical and structural characterization of biosurfactant produced by <Emphasis Type="Italic">Pleurotus djamor</Emphasis> in solid-state fermentation

Biosurfactants are amphiphilic compounds produced by several microorganisms that reduce the surface tension. Low toxicity, optimal activity in extreme conditions, biodegradability and production from several wastes are main advantages of biosurfactants as compared to synthetic surfactants. Production of biosurfactant by a white rot fungus Pleurotus djamor on sunflower seed shell in solid-state fermentation was determined by emulsification indexes, oil spreading activity and surface tension (28.82 ± 0.3mN/m) measurement. The critical micelle concentration was detected as 0.964 ± 0.09 mg/mL. Also, the chemical and physicochemical properties of the biosurfactant produced were investigated. Considering the results of the chemical contents analysis, HPLC, FT-IR and ¹H-NMR, it can be concluded that the produced biosurfactant has a complex structure. Besides, resistance of its activity to environmental factors such as temperature, pH and salt concentration, as well as its thermal stability, were investigated. Additionally, the produced biosurfactant formed stabile emulsions with different hydrocarbons. Lastly, the performance of removing waste frying oil from contaminated sand of produced biosurfactant was detected as 76.57 ± 6%. Owing to its high emulsification capacity, low surface tension and critical micelle concentration, the biosurfactant, shows great potential for use in hydrocarbon removal applications.     

Isolation and characterization of biosurfactant production under extreme environmental conditions by alkali-halo-thermophilic bacteria from Saudi Arabia

Twenty three morphologically distinct microbial colonies were isolated from soil and sea water samples, which were collected from Jeddah region, Saudi Arabia for screening of the most potent biosurfactant strains. The isolated bacteria were selected by using different methods as drop collapse test, oil displacement test, blue agar test, blood hemolysis test, emulsification activity and surface tension. The results showed that the ability of Virgibacillus salarius to grow and reduce surface tension under a wide range of pH, salinities and temperatures gives bacteria isolate an advantage in many applications such as pharmaceutical, cosmetics, food industries and bioremediation in marine environment. The biosurfactant production by V. salarius decreased surface tension and emulsifying activity (30 mN/m and 80%, respectively). In addition to reducing the production cost of biosurfactants by tested several plant-derived oils such as jatropha oil, castor oils, jojoba oil, canola oil and cottonseed oil. In this respect the feasibility to reusing old frying oil of sunflower for production rhamnolipids and sophorolipids, their use that lead to solve many ecological and industrial problems.     

Metal Removal from Contaminated Soils Through Bioleaching with Oxidizing Bacteria and Rhamnolipid Biosurfactants

The use of surfactants as a method for solubilization and removal of heavy metal contamination from soil has been reported before. Biosurfactants produced by some microorganisms are able to modify the surface of various metals and aggregate on interphases favoring the metal separation process from contaminated environments. We evaluated the feasibility of enhancing the removal of metal ions from mineral waste/contaminated soils using alternate cycles of treatment with rhamnolipid biosurfactants and bioleaching with a mixed bacterial culture of Acidithiobacillus thiooxidans and Acidithiobacillus ferrooxidans. Bioleaching alone removed 50% Zn and 19% Fe. When rhamnolipids were used at low concentration (0.4 mg/mL), 11% Fe and 25% Zn were removed, while at 1 mg/mL 19% Fe and 52% Zn removal were achieved. When using a cyclic treatment combining bioleaching and biosurfactants, metal removal reached up to 36% for Fe and 63% to 70% for Zn.     

MICROBIAL SYNTHESIS OF RHAMNOLIPIDS BY Pseudomonas aeruginosa (ATCC 10145) ON WASTE FRYING OIL AS LOW COST CARBON SOURCE

Vegetable edible oils and fats are mainly used for frying purposes in households and the food industry. The oil undergoes degradation during frying and hence has to be replaced from time to time. Rhamnolipids are produced by microbial cultivation using refined vegetable oils as a carbon source and Pseudomonas aeruginosa (ATCC 10145). The raw material cost accounts for 10–30% of the overall cost of biosurfactant production and can be reduced by using low-cost substrates. In this research, attention was focused on the preparation of rhamnolipids, which are biosurfactants, using potential frying edible oils as a carbon source via a microbial fermentation technique. The use of low-cost substrates as a carbon source was emphasized to tilt the cost of production for rhamnolipids. The yield was 2.8 g/L and 7.5 g/L from waste frying oil before and after activated earth treatment, respectively. The crude product contained mainly dirhamnolipids, confirmed by thin-layer chromatography (TLC), high-performance liquid chromatography (HPLC), liquid chromatography–mass spectroscopy (LC-MS), and ¹H-nuclear magnetic resonance (NMR). Hence, the treatment can be used to convert waste frying oil as a low-cost substrate into a cost-effective carbon source.     

Microbial biosurfactant mediated removal and/or solubilization of crude oil contamination from soil and aqueous phase: An approach with Bacillus licheniformis MTCC 5514

This study highlights the role of marine microbial biosurfactants on solubilization/removal of crude-oil contamination from four different soils in an aqueous phase. Soil of four different types, viz., sandy, fine sand soil, clay, and clay loam, were collected and saturated with crude oil. Marine isolate MTCC 5514 (Bacillus licheniformis) was chosen for the study and comparisons were made with synthetic surfactants and commercially available biosurfactant. In-situ studies were carried out with different percentages of crude oil to assess the growth and the percentage removal of oil. For ex-situ studies, soils were pre-saturated with crude oil and then treated with the chosen biosurfactant at a 10% concentration level using flask and column methods. After time intervals of 30–120 min, samples were collected and then subjected to extraction with hexane and the percentage removal was calculated. Results revealed, at 2% concentration of crude oil, that complete solubilization was achieved. With regard to ex-situ studies, clay soil absorbed the maximum percentage of oil compared to other soil types, and with regard to the removal, all the synthetic surfactants showed <60% removal irrespective of soil type. In the case of biosurfactants even at 10% concentration, >85% removal was achieved.     

泡菜水中产生物表面活性剂酵母菌株的筛选

[背景]由微生物产生的生物表面活性剂(biosurfactant,BS)具有低毒性、高效性、生物可降解性等多种特性,能在一定程度上缓解化学表面活性剂所造成的环境问题,因此筛选高产、安全的BS生产菌株备受研究者的关注.[目的]从泡菜水中筛选能代谢合成药食两用型BS的微生物菌株.[方法]运用滴崩法和排油圈法从传统发酵食品泡...     

In Situ Biosurfactant Production by Bacillus Strains Injected into a Limestone Petroleum Reservoir

Biosurfactant-mediated oil recovery may be an economic approach for recovery of significant amounts of oil entrapped in reservoirs, but evidence that biosurfactants can be produced in situ at concentrations needed to mobilize oil is lacking. We tested whether two Bacillus strains that produce lipopeptide biosurfactants can metabolize and produce their biosurfactants in an oil reservoir. Five wells that produce from the same Viola limestone formation were used. Two wells received an inoculum (a mixture of Bacillus strain RS-1 and Bacillus subtilis subsp. spizizenii NRRL B-23049) and nutrients (glucose, sodium nitrate, and trace metals), two wells received just nutrients, and one well received only formation water. Results showed in situ metabolism and biosurfactant production. The average concentration of lipopeptide biosurfactant in the produced fluids of the inoculated wells was about 90 mg/liter. This concentration is approximately nine times the minimum concentration required to mobilize entrapped oil from sandstone cores. Carbon dioxide, acetate, lactate, ethanol, and 2,3-butanediol were detected in the produced fluids of the inoculated wells. Only CO₂ and ethanol were detected in the produced fluids of the nutrient-only-treated wells. Microbiological and molecular data showed that the microorganisms injected into the formation were retrieved in the produced fluids of the inoculated wells. We provide essential data for modeling microbial oil recovery processes in situ, including growth rates (0.06 ± 0.01 h⁻¹), carbon balances (107% ± 34%), biosurfactant production rates (0.02 ± 0.001 h⁻¹), and biosurfactant yields (0.015 ± 0.001 mol biosurfactant/mol glucose). The data demonstrate the technical feasibility of microbial processes for oil recovery.     

Rhamnolipid biosurfactant production by Pseudomonas nitroreducens immobilized on Ca2+ alginate beads and under resting cell condition

Pseudomonas nitroreducens MILB-8054A isolated from petroleum-contaminated soil, immobilized on calcium alginate beads, and under resting cell condition, produced biosurfactants. Immobilized cells gave a best yield of 5.6 g rhamnolipid l^?1 using sucrose as carbon source. Time course study using resting cells showed that 2 % v/v of palm oil (preculture carbon source) and 10 % diesel (carbon source) gave the best rhamnolipid yield of 5.1 g l^?1 at pH 8 and temperature of 30 °C. Carbon utilization by resting cells was compared with that of growing cells. The best biosurfactant recovery procedure was acetone extraction.     

Bench-Scale Production of Biosurfactants and their Potential in Ex-Situ MEOR Application

The fermentative production of biosurfactants by five Bacillus strains in a bench-scale bioreactor and evaluation of biosurfactant-based enhanced oil recovery using sand pack columns were investigated. Adjusting the initial dissolved oxygen to 100% saturation, without any further control and with collection of foam and recycling of biomass, gave higher biosurfactant production. The microorganisms were able to produce biosurfactants, thus reducing the surface tension and interfacial tension to 28 mN/m and 5.8–0.5 mN/m, respectively, in less than 10 hours. The crude surfactant concentration of 0.08–1.1 g/L, and critical micelle concentration (CMC) values of 19.4–39 mg/L, corresponding to the biosurfactants produced by the different Bacillus strains, were observed. The efficiency of crude biosurfactant preparation obtained from Bacillus strains for enhanced oil recovery, by sand pack column studies, revealed it to vary from 30.22–34.19% of the water flood residual oil saturation. The results are indicative of the potential of the strains for the development of ex-situ, microbial-enhanced, oil recovery processes.     

Identification and ultra‐high‐performance liquid chromatography coupled with high‐resolution mass spectrometry characterization of biosurfactants,including a new surfactin,isolated from oil‐contaminated environments

Biosurfactant‐producing bacteria were isolated from samples collected in areas contaminated with crude oil. The isolates were screened for biosurfactant production using qualitative drop‐collapse test, oil‐spreading and emulsification assays, and measurement of their tensoactive properties. Five isolates tested positive for in the screening experiments and displayed decrease in the surface tension below 30 mN m^?1. The biosurfactants produced by these isolates were further investigated and their molecular identification revealed that they are bacteria related to the Bacillus genus. Additionally, the biosurfactants produced were chemically characterized via UHPLC‐HRMS experiments, indicating the production of surfactin homologues, including a new class of these molecules.     

Isolation and Characterization of Biosurfactant-and Bioemulsifier-Producing Bacteria from Petroleum Contaminated Sites in Western Canada

Biosurfactant-producing bacteria were isolated from two petroleum contaminated sites in western Canada. Seven potential biosurfactant/bioemulsifier-producing isolates were screened and characterized. All of the seven isolates were able to form emulsions. Emulsion-stabilizing capacity was also measured up to 48 hrs. Strain C-111-2 and C-203-2 would lead to highly reduced surface tension. For strain C-203-2, the optimum conditions that supported bacteria growth and production were investigated. The influences of carbon sources, medium pH values, and temperature were taken into account. The experimental results indicated that the crude oil and glucose were promising carbon sources for biosurfactants production; the isolated strains produced a maximum concentration of biosurfactant in a neutral pH environment and showed a higher surface activity under the temperature level of 35°C than that under 10°C. To further optimize the carbon and nitrogen source for biosurfactant production, response surface methodology (RSM) was applied to explore the favorable concentration of two carbon sources: glucose, crude oil, and one nitrogen source, NaNO₃. The optimal concentration of 8.1g/L, 4% and 3.9 g/L for glucose, crude oil, and NaNO₃, respectively, which can be obtained through RSM analysis.     

Importance of 3-Hydroxy Fatty Acid Composition of Lipopeptides for Biosurfactant Activity

Biosurfactant production may be an economic approach to improving oil recovery. To obtain candidates most suitable for oil recovery, 207 strains, mostly belonging to the genus Bacillus, were tested for growth and biosurfactant production in medium with 5% NaCl under aerobic and anaerobic conditions. All strains grew aerobically with 5% NaCl, and 147 strains produced a biosurfactant. Thirty-five strains grew anaerobically with 5% NaCl, and two produced a biosurfactant. In order to relate structural differences to activity, eight lipopeptide biosurfactants with different specific activities produced by various Bacillus species were purified by a new protocol. The amino acid compositions of the eight lipopeptides were the same (Glu/Gln:Asp/Asn:Val:Leu, 1:1:1:4), but the fatty acid compositions differed. Multiple regression analysis showed that the specific biosurfactant activity depended on the ratios of both iso to normal even-numbered fatty acids and anteiso to iso odd-numbered fatty acids. A multiple regression model accurately predicted the specific biosurfactant activities of four newly purified biosurfactants (r² = 0.91). The fatty acid composition of the biosurfactant produced by Bacillus subtilis subsp. subtilis strain T89-42 was altered by the addition of branched-chain amino acids to the growth medium. The specific activities of biosurfactants produced in cultures with different amino acid additions were accurately predicted by the multiple regression model derived from the fatty acid compositions (r² = 0.95). Our work shows that many strains of Bacillus mojavensis and Bacillus subtilis produce biosurfactants and that the fatty acid composition is important for biosurfactant activity.     

Isolation and characterization of a biosurfactant‐producing Fusarium sp. BS‐8 from oil contaminated soil

This study reports characterization of a biosurfactant‐producing fungal isolate from oil contaminated soil of Missa Keswal oil field, Pakistan. It was identified as Fusarium sp. BS‐8 on the basis of macroscopic and microscopic morphology, and 18S rDNA gene sequence homology. The biosurfactant‐producing capability of the fungal isolates was screened using oil displacement activity, emulsification index assay, and surface tension (SFT) measurement. The optimization of operational parameters and culture conditions resulted in maximum biosurfactant production using 9% (v/v) inoculum at 30°C, pH 7.0, using sucrose and yeast extract, as carbon and nitrogen sources, respectively. A C:N ratio of 0.9:0.1 (w/w) was found to be optimum for growth and biosurfactant production. At optimal conditions, it attained lowest SFT (i.e., 32 mN m^?1) with a critical micelle concentration of ≥ 1.2 mg mL^?1. During 5 L shake flask fermentation experiments, the biosurfactant productivity was 1.21 g L^?1 pure biosurfactant having significant emulsifying index (E₂₄, 70%) and oil‐displacing activity (16 mm). Thin layer chromatography and Fourier transform infrared spectrometric analyses indicated a lipopeptide type of the biosurfactant. The Fusarium sp. BS‐8 has substantial potential of biosurfactant production, yet it needs to be fully characterized with possibility of relatively new class of biosurfactants. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 30:1065–1075, 2014     

Potential application of Bacillus subtilis SPB1 biosurfactants in laundry detergent formulations: Compatibility study with detergent ingredients and washing performance

Surfactants play a very important role in laundry and household cleaning products ingredients. In this research, the application of lipopeptide biosurfactants, produced by Bacillus subtilis SPB1, in the formulation of a washing powder was investigated. The SPB1 biosurfactant was mixed with sodium tripolyphosphate as a builder and sodium sulfate as filler. The efficiency of the formulated detergent composition with different washing conditions to remove a stain from cotton fabric was examined. The results showed that the formulated detergent was effective in oil removal, with optimal washing conditions of pH, temperature, striate and time of washing system of 7, 65°C, 1000 RPM and 60 min, respectively. A comparative study of different detergent compositions (biosurfactant‐based detergent, combined biosurfactant‐commercial detergent, and a commercial detergent) for the removal of oil and tea stains, proved that the bio‐scouring was more effective (>75%) in terms of the stain removal than the commercial powders (<60%). Moreover, the results demonstrated that the biosurfactant acts additively with a commercial detergent and enhances their performance from 33 to 45% in removing oil stain and from 57 to 64% in removing tea stain. As a conclusion, in addition to the low toxicity and the high biodegradability of the microbial biosurfactants, the results of this study have shown that the future use of this lipopeptide biosurfactant as laundry detergent additive is highly promising.     

Isolation of biosurfactant producers, optimization and properties of biosurfactant produced by Acinetobacter sp. from petroleum-contaminated soil

Aims: To screen and identify biosurfactant producers from petroleum‐contaminated soil; to use response surface methodology (RSM) for medium optimization to enhance biosurfactant production; and to study the properties of the newly obtained biosurfactant towards pH, temperature and salinity. Methods and Results: We successfully isolated three biosurfactant producers from petroleum‐contaminated soil and identified them through 16S rRNA sequence analysis, which exhibit the highest similarities to Acinetobacter beijerinckii (100%), Kocuria marina (99%) and Kineococcus marinus (99%), respectively. A quadratic response model was constructed through RSM designs, leading to a 57·5% increase of the growth‐associated biosurfactant production by Acinetobacter sp. YC‐X 2 with an optimized medium: beef extract 3·12 g l^?1; peptone 20·87 g l^?1; NaCl 1·04 g l^?1; and n‐hexadecane 1·86 g l^?1. Biosurfactant produced by Acinetobacter sp. YC‐X 2 retained its properties during exposure to a wide range of pH values (5–11), high temperatures (up to 121°C) and high salinities [up to 18% (w/v) Na⁺ and Ca²⁺], which was more sensitive to Ca²⁺ than Na⁺. Conclusions: Two novel biosurfactant producers were isolated from petroleum‐contaminated soil. Biosurfactant from Acinetobacter sp. YC‐X 2 has good properties to a wide range of pH, high temperature and high salinity, and its production was optimized successfully through RSM. Significance and Impact of the Study: The fact, an increasing demand of high‐quality surfactants and the lack of cost‐competitive bioprocesses of biosurfactants for commercial utilization, motivates researchers to develop cost‐effective strategies for biosurfactant production through isolating new biosurfactant producers with special surface‐active properties and optimizing their cultural conditions. Two novel biosurfactant producers in this study will widen our knowledge about this kind of micro‐organism. This work is the first application of RSM designs for cultural optimization of biosurfactant produced by Acinetobacter genus and the first report that biosurfactant may be more sensitive to Ca²⁺ than Na⁺.     

Production of rhamnolipids by a Pseudomonas alcaligenes strain

Brazil is one of the main producers of palm oil (Ellaus guineeusis). It is a low-cost product that has some interesting industrial qualities, such as its use as the raw material for the production of glycerin and soap as well as its use in the preparation of food. Some renewable sources and agroindustrial wastes have been used extensively in research on the production of biosurfactants of the Pseudomonas strains. However, to our knowledge, no studies have been published on the use of palm oil as a substrate for the synthesis of biosurfactants by Pseudomonas alcaligenes. This paper describes the production and characterization of biosurfactants synthesized by a strain of P. alcaligenes PCL previously isolated from soil that was contaminated with crude-oil. Furthermore, the paper presents the optimization of the production of biological surface-active compounds by applying experimental design tools and their capacity to emulsify hydrocarbons.     

The Potential of Bacterial Isolates for Emulsification with a Range of Hydrocarbons

A study was undertaken to investigate the distribution of biosurfactant producing and crude oil degrading bacteria in the oil contaminated environment. This research revealed that hydrocarbon contaminated sites are the potent sources for oil degraders. Among 32 oil degrading bacteria isolated from ten different oil contaminated sites of gasoline and diesel fuel stations, 80% exhibited biosurfactant production. The quantity and emulsification activity of the biosurfactants varied. Pseudomonas sp. DS10‐129 produced a maximum of 7.5 ± 0.4 g/l of biosurfactant with a corresponding reduction in surface tension from 68 mN/m to 29.4 ± 0.7 mN/m at 84 h incubation. The isolates Micrococcus sp. GS2‐22, Bacillus sp. DS6‐86, Corynebacterium sp. GS5‐66, Flavobacterium sp. DS5‐73, Pseudomonas sp. DS10‐129, Pseudomonas sp. DS9‐119 and Acinetobacter sp. DS5‐74 emulsified xylene, benzene, n‐hexane, Bombay High crude oil, kerosene, gasoline, diesel fuel and olive oil. The first five of the above isolates had the highest emulsification activity and crude oil degradation ability and were selected for the preparation of a mixed bacterial consortium, which was also an efficient biosurfactant producing oil emulsifying and degrading culture. During this study, biosurfactant production and emulsification activity were detected in Moraxella sp., Flavobacterium sp. and in a mixed bacterial consortium, which have not been reported before.     

Application of Biosurfactants Produced by Pseudomonas aeruginosa SP4 for Bioremediation of Soils Contaminated by Pyrene

Biosurfactants are considered to facilitate PAHs dissolution in soil slurries for bioremediation applications. In this work, the carbon and nitrogen sources, pH, C/N ratio, and salinity, were considered for optimization of biosurfactant production by Pseudomonas aeruginosa SP4 isolate to enhance pyrene removal from the contaminated soil. Analysis of ANOVA indicated that the carbon source was the most effective factor, followed by pH, nitrogen source, C/N ratio, and salinity. Taguchi experimental design proposed the optimum operating conditions of olive oil, NH₄NO₃, C/N ratio of 5, salinity of 0.5%, and pH 7. Applying the conditions determined by Taguchi design led to a production yield of 452 mg L^?1 (13% improvement) at the optimum conditions. The main characteristics of produced biosurfactant included the critical micelle concentration (CMC) of 60 mg L^?1 and liquid medium surface tension of 29.5 mN m^?1. Produced biosurfactant was used for bioremediation of soil artificially contaminated with 500 mg kg^?1 of pyrene. Following the addition of 250 mg L^?1 biosurfactant, the pyrene removal of 84.6% was obtained compared to 59.8% for control sample without any surfactant.