Aerobic biodegradation and inhibition kinetics of poly‐aromatic hydrocarbons (PAHs) in a petrochemical industry wastewater in the presence of biosurfactants

BACKROUND: In Izmir, Turkey, wastewaters from the petrochemical industry are treated using conventional activated sludge systems. A significant proportion of poly‐aromatic hydrocarbons (PAHs) with high‐molecular weights remains in this treatment system and inhibits the biological activity. Biosurfactants increase PAHs degradation by enhancing the solubility of the petroleum components. The aerobic inhibition kinetics of PAHs has not previously been investigated in the presence of biosurfactants for a real petrochemical industry wastewater. RESULTS: Among the kinetic models used (Monod‐type, zero, first‐order and second‐order) it was found that the Monod kinetic was effective for describing the biodegradation of PAHs in petrochemcal industry wastewater in the presence of three biosurfactants, namely Rhamnolipid (RD), Surfactine (SR) and Emulsan (EM) in an aerobic activated sludge reactor (AASR). The maximum PAH removal rate (R_max) and specific growth rate of PAH degrading bacteria (µ_max) increased, while the half saturation concentration of PAH (K_s) decreased at 15 mg L^?1 RD concentration compared with the control without biosurfactant at a sludge retention time (SRT) of 25 days. CONCLUSION: PAH oxidation is typified by competitive inhibition at RD concentrations > 15 mg L^?1 resulting in increases in K_s values with PAH accumulation. Low inhibition constant (K_ID) values reflect difficulties in the metabolizability of PAHs. Metabolite production decreased at RD = 25 mg L^?1 in the PAHs indeno (1,2,3‐cd) pyrene (IcdP), flourene (FLN), phenanthrene (PHE) and benzo(a)pyrene (BaP). Copyright © 2011 Society of Chemical Industry     

Statistical optimization of fermentation conditions for the improved production of poly-β-hydroxybutyrate from Bacillus subtilis

Poly-β-hydroxybutyrate (PHB) has been an effective biodegradable plastic obtained by microbial fermentation. Batch fermentation of Bacillus subtilis features an attractive system for the production of PHB. Identification of appropriate media components and cultivation conditions are extremely important for the optimal production of biomass and/or PHB production. Statistical media design was utilized for the optimization of different fermentation variables (glucose, peptone, sodium chloride, K₂HPO₄, KH₂PO₄, ammonium sulfate, ammonium chloride, sodium sulfate, temperature, inoculum size, and pH). The optimized media predicted the optimal dry cell weight of 7.54?g?L^?1 and PHB production of 77.2?mg?L^?1 at 1?g?L^?1 of peptone, 1.46?g?L^?1 sodium sulfate, and pH 6.8 in 24?h. Glucose utilization, batch growth, and PHB production kinetics of B. subtilis were determined experimentally. The effect of substrate inhibition on specific growth rate was also determined experimentally for B. subtilis. The values of kinetic and substrate inhibition parameters obtained from this study shall be utilized to develop a mathematical model for PHB production for further improving the production of PHB.     

Economic production of Bacillus subtilis SPB1 biosurfactant using local agro‐industrial wastes and its application in enhancing solubility of diesel

BACKGROUND: The present work aimed to optimize a new economic medium for lipopeptide biosurfactant production by Bacillus subtilis SPB1 for application in the environmental field as an enhancer of diesel solubility. Statistical experimental designs and response surface methodology were employed to optimize the medium components. RESULTS: A central composite design was applied to increase the production yield and predict the optimal values of the selected factors. An optimal medium, for biosurfactant production of about 4.5 g L^?1, was found to be composed of sesame peel flour (33 g L^?1) and diluted tuna fish cooking residue (40%) with an inoculum size of 0.22. Increased inoculum size (final OD₆₀₀) significantly improved the production yield. The emulsifier produced was demonstrated to be an alternative to chemically synthesized surfactants since it shows high solubilization efficiency towards diesel oil in comparison with SDS and Tween 80. CONCLUSION: Optimization studies led to a strong improvement in production yield. The emulsifier produced, owing its high solubilization capacity and its large tolerance to acidic and alkaline pH values and salinity, shows great potential for use in bioremediation processes to enhance the solubility of hydrophobic compounds. © 2012 Society of Chemical Industry     

Effect of nutrients on optimal production of biosurfactants by <Emphasis Type="Italic">Pseudomonas putida</Emphasis>—A gujarat oil field isolate

Nutritional requirements for maximal production of biosurfactant by an oil field bacterium Pseudomonas putida were determined. The optimal concentrations of nitrogen, phosphate, sulfur, magnesium, iron, potassium, sodium, calcium, and trace elements for maximal production of biosurfactants were ascertained, and a new “Pruthi and Cameotra” salt medium was formulated. Data show that maximal biomass (2.4 g L⁻¹) and biosurfactant production (6.28 g L⁻¹) takes place after 72 h of growth on 2% hexadecane. The biosurfactant was produced optimally over pH and temperature ranges of 6.4–7.2 and 30–40°C, respectively. That the highest biosurfactant yield was obtained during late log phase of growth indicates that the biosurfactant is a secondary metabolite.     

Culture Medium Evaluation Using Low-Cost Substrate for Biosurfactants Lipopeptides Production by Bacillus amyloliquefaciens in Pilot Bioreactor

Total biosurfactant production by Bacillus amyloliquefaciens IT45 was evaluated with different substrates concentrations in a culture medium. A central composite design (CCD) was developed to evaluate the influence of variables, including glucose syrup, yeast extract, and calcium chloride, on surface tension (ST), total biosurfactant production, and residual sugar (RS). As a result, the best observed results for ST, RS, and total biosurfactant production were 30 mN m⁻¹, 31%, and 5.5 g L⁻¹, respectively, after 48 h of fermentations carried out in batch operation process. Characterization of the biosurfactant identified the presence of surfactin. To validate the CCD experiments, fermentations were conducted in a 40 L pilot bioreactor. For this fermentation, the cellular growth was 3.0 × 10⁹ CFU mL⁻¹, surfactin production was 0.55 g L⁻¹, and RS was 28%. The results demonstrate that B. amyloliquefaciens IT45 has the potential to produce biosurfactants and does not require high concentrations of carbon and nitrogen sources for its development.     

Batch production of biosurfactant with foam fractionation

Methods of producing the biosurfactant surfactin from cultures of Bacillus subtilis (BBK006) have been investigated. A reactor with integrated foam fractionation was designed and used in batch mode, and the performance compared with that of the same culture in shaken flasks. In the batch reactor, significant foaming occurred between 12.5 h and 14.5 h of culture time. During this period, the foam was routed through the foam fractionation column to a mechanical foam breaker, and a biosurfactant‐enriched foamate was collected. Concentration of surfactin in the foamate product was around 50 times greater than that in the culture medium. Using the integrated reactor, 136 mg L^?1 of surfactin was produced, significantly more than was achieved in shaken flasks (92 mg L^?1). The foam fractionation method allowed a real‐time measurement of the rate of surfactin production during growth. This showed that the maximum rate of production occurred at the interphase between log and stationary modes of growth, in contrast to previous work showing that surfactin is exclusively a secondary metabolite. The high value of surfactin yield in relation to biomass (Y_P/x = 0.262) indicated that surfactin was produced very efficiently by Bacillus subtilis (BBK006) in this integrated bioreactor. Copyright © 2006 Society of Chemical Industry     

Kinetic study on ethanol production using Saccharomyces cerevisiae ITV‐01 yeast isolated from sugar cane molasses

BACKGROUND: Bio‐ethanol production from renewable sources, such as sugar cane, makes it a biofuel that is both renewable and environmentally friendly. One of the strategies to reduce production costs and to make ethanol fuel economically competitive with fossil fuels could be the use of wild yeast with osmotolerance, ethanol resistance and low nutritional requirements. The aim of this work was to investigate the kinetics of ethanol fermentation using Saccharomyces cerevisiae ITV‐01 yeast strain in a batch system at different glucose and ethanol concentrations, pH values and temperature in order to determine the optimum fermentation conditions. RESULTS: This strain showed osmotolerance (its specific growth rate (µ_max) remained unchanged at glucose concentrations between 100 and 200 g L^?1) as well as ethanol resistance (it was able to grow at 10% v/v ethanol). Activation energy (Ea) and Q₁₀ values calculated at temperatures between 27 and 39 °C, pH 3.5, was 15.6 kcal mol^?1 (with a pre‐exponential factor of 3.8 × 10¹² h^?1 (R² = 0.94)) and 3.93 respectively, indicating that this system is biologically limited. CONCLUSIONS: The optimal conditions for ethanol production were pH 3.5, 30 °C and initial glucose concentration 150 g L^?1. In this case, a maximum ethanol concentration of 58.4 g L^?1, ethanol productivity of 1.8 g L^?1 h^?1 and ethanol yield of 0.41 g g^?1 were obtained. Copyright © 2010 Society of Chemical Industry     

Modeling of Rhamnolipid Biosurfactant Production: Estimation of Kinetic Parameters by Genetic Algorithm

Studies about kinetics and modeling of production parameters for biosurfactants are essential to the development of efficient processes from an economic point of view. In this sense, this work evaluated the performance of four nonstructured models to explain the experimental data for biomass growth, substrate consumption, and rhamnolipid production using glycerol as carbon source and a Pseudomonas aeruginosa strain. The kinetic parameters of each model were estimated using a global search method known as genetic algorithm and numerical discretization of differential equations by the Runge–Kutta 4th order method. The main result of this study showed that the Monod model best represented the experimental data, with μ_max values of 0.06 h⁻¹, K_S of 50.8 g L⁻¹, Y_X/S of 0.43 g g⁻¹, and Y_P/X equal to 0.017 g g⁻¹.     

Utilization of molasses for biosurfactant production by two Bacillus strains at thermophilic conditions

Traditionally, biosurfactants have been produced from hydrocarbons. Some possible substitutes for microbial growth and biosurfactant production include urban wastes, peat hydrolysate, and agro-industrial by-products. Molasses, a nonconventional substrate (agro-industrial by-product) can also be used for biosurfactant production. It has been utilized by two strains of Bacillus subtilis (MTCC 2423 and MTCC1427) for biosurfactant production and growth at 45°C. As a result of biosurfactant accumulation, the surface tension of the medium was lowered to 29 and 31 dynes/cm by the two strains, respectively. This is the first report of biosurfactant production by strains of B. subtilis at 45°C. Potential application of the biosurfactant in microbial enhanced oil recovery is also presented.     

Characterization of Biosurfactant Produced by a Novel Strain of Pseudomonas aeruginosa,Isolate ADMT1

Several biosurfactant‐producing bacterial strains were isolated from petroleum‐contaminated soil. The isolate ADMT1, identified as a new strain of Pseudomonas aeruginosa, was selected for further studies on the basis of oil displacement test and emulsification index (E24). The optimal parameters for production, determined by employing Box–Behnken design, were temperature 36.5 °C and pH 7. The environmental isolate ADMT1 produced significant amount of biosurfactant (1.7 g L^?1 in 72 h) in minimal salt medium (MSM) using dextrose as the sole carbon source. The E24 value and critical micelle concentration (CMC) of the biosurfactant was 100% and 150 mg L^?1, respectively. At CMC, the surface tension of water was reduced to 28.4 mN m^?1. The biosurfactant exhibited hemolytic activity and antibacterial activity against 8 reference strains of pathogenic bacteria, including 2 methicillin‐resistant Staphylococcus aureus strains (MRSA ATCC 562 and MRSA ATCC 43300), with minimum inhibitory concentration (MIC) of 0.4 and 0.2 mg mL^?1, respectively. The structure of biosurfactant was characterized by FTIR, ¹H, and ¹³C NMR spectroscopy. 7 di‐rhamnolipid (RL) congeners were identified in the biosurfactant by ultraperformance liquid chromatography–mass spectrometry analysis. The major congeners, which constituted 67% of the RL mixture, included Rha‐Rha‐C₁₀‐C₁₀, Rha‐Rha‐C₁₂‐C₁₀, and Rha‐Rha‐C_12:1‐C₁₀. The minor congeners were Rha‐Rha‐C₁₀‐C₈, Rha‐Rha‐C_10:1‐C₁₀, Rha‐Rha‐C₁₀‐C_14:1, and Rha‐Rha‐C₁₀‐C₁₄. The congener Rha‐Rha‐C₁₀‐C₁₄ is being reported for the first time from any species of Pseudomonas. The high surface activity and E24 value make the ADMT1‐RL a potential candidate for its use in detergents, environmental bioremediation, and as an emulsifier in the food industry.     

Conversion efficiency of glucose/xylose mixtures for ethanol production using Saccharomyces cerevisiae ITV01 and Pichia stipitis NRRL Y‐7124

BACKGROUND: Efficient conversion of glucose/xylose mixtures from lignocellulose is necessary for commercially viable ethanol production. Oxygen and carbon sources are of paramount importance for ethanol yield. The aim of this work was to evaluate different glucose/xylose mixtures for ethanol production using S. cerevisiae ITV‐01 (wild type yeast) and P. stipitis NRRL Y‐7124 and the effect of supplying oxygen in separate and co‐culture processes. RESULTS: The complete conversion of a glucose/xylose mixture (75/30 g L^?1) was obtained using P. stipitis NRRL Y‐7124 under aerobic conditions (0.6 vvm), the highest yield production being Y_p/s = 0.46 g g^?1, volumetric ethanol productivity Q_pmax = 0.24 g L^?1 h^?1 and maximum ethanol concentration P_max = 34.5 g L^?1. In the co‐culture process and under aerobic conditions, incomplete conversion of glucose/xylose mixture was observed (20.4% residual xylose), with a maximum ethanol production of 30.3 g L^?1, ethanol yield of 0.4 g g^?1 and Q_pmax = 1.26 g L^?1 h^?1. CONCLUSIONS: The oxygen present in the glucose/xylose mixture promotes complete sugar consumption by P. stipitis NRRL Y‐7124 resulting in ethanol production. However, in co‐culture with S. cerevisiae ITV‐01 under aerobic conditions, incomplete fermentation occurs that could be caused by oxygen limitation and ethanol inhibition by P. stipitis NRRL Y‐7124; nevertheless the volumetric ethanol productivity increases fivefold compared with separate culture. Copyright © 2011 Society of Chemical Industry     

Production of L‐lactic acid with very high gravity distillery wastewater from ethanol fermentation by a newly isolated Enterococcus hawaiiensis

BACKGROUND: A great amount of wastewater with high contents of chemical oxygen demand (COD) are produced by ethanol production. It would be useful to utilize distillery wastewater to produce L‐lactic acid, which could be a high additional value byproduct of ethanol production. The fermentation process of L‐lactic acid production by a newly isolated Enterococcus hawaiiensis CICIM‐CU B0114 is reported for the first time. RESULTS: The strain produced 56 g L^?1 of L‐lactic acid after cultivation for 48 h in optimized medium consisting of (g L^?1) 80 glucose, 10 peptone, 10 yeast extract, 1.5 Na₂HPO₄ and 0.2 MgSO₄. E. hawaiiensis CICIM‐CU B0114 was isolated and purified by subculture for growing and producing L‐lactic acid in distillery wastewater of very high gravity (VHG) from ethanol fermentation. L‐lactic acid fermentation was further studied with distillery wastewater substrate in 7 L and 15 L fermentors. The results showed that L‐lactic acid concentrations of 52 g L^?1 and 68 g L^?1 was achieved in 7 L and 15 L fermentors with the initial sugar concentrations of 67 g L^?1 and 87 g L^?1, respectively. CONCLUSION: The production of L‐lactic acid by the newly isolated E. hawaiiensis CICIM‐CU B0114 was carried out and the fermentation medium was optimized by orthogonal experimental design. This new strain holds the promise of L‐lactic acid production utilizing distillery wastewater from VHG ethanol fermentation. Copyright © 2010 Society of Chemical Industry     

Mathematical modelling of the alcoholic fermentation of glucose by Zymomonas mobilis mobilis

The kinetics of alcoholic fermentation of a strain of Zymomonas mobilis, isolated from sugarcane juice, has been studied with the objective of determining the constansts of a non-structured mathematical model that represents the fermentation process. Assays in batch and in continuous culture have been carried out with different initial concentrations of glucose. The final concentrations of glucose, ethanol and biomass were determined. The following kinetic parameters were obtained: μ_max, 0·5 h^?1; K_s, 4·64 g dm^?3; P_max, 106 g dm^?3; Y_x/s, 0·0265 g g^?1; m, 1·4 g g^?1 h^?1; α, 17·38 g g^?1; β, 0·69 g g^?1 h^?1.     

Catalysis of an Essential Step in Vitamin B2 Biosynthesis by a Consortium of Broad Spectrum Hydrolases

An enzyme catalysing the essential dephosphorylation of the riboflavin precursor, 5‐amino‐6‐ribitylamino‐2,4(1H,3H)‐pyrimidinedione 5′‐phosphate ( 6 ), was purified about 800‐fold from a riboflavin‐producing Bacillus subtilis strain, and was assigned as the translation product of the ycsE gene by mass spectrometry. YcsE is a member of the large haloacid dehalogenase (HAD) superfamily. The recombinant protein was expressed in Escherichia coli. It catalyses the hydrolysis of 6 (v_max, 12 μmol mg^?1 min^?1; K_M, 54 μm ) and of FMN (v_max, 25 μmol mg^?1 min^?1; K_M, 135 μm ). A ycsE deletion mutant of B. subtilis was not riboflavin dependent. Two additional proteins (YwtE, YitU) that catalyse the hydrolysis of 6 at appreciable rates were identified by screening 13 putative HAD superfamily members from B. subtilis. The evolutionary processes that have resulted in the handling of an essential step in the biosynthesis of an essential cofactor by a consortium of promiscuous enzymes require further analysis.     

Continuous production of biosurfactant with foam fractionation

A glucose‐limited chemostat was used to determine the growth parameters of BBK006 for continuous production of the biosurfactant surfactin. The continuous cultivation exhibited low maintenance metabolism (m = 0.39 mmol_glucose g_bacteria^?1 h^?1) and high molar growth yield ( g_bacteria mol_glucose^?1). It was found that the surfactin production rate in continuous culture was not only a function of dilution rate but also varied with the initial concentration of glucose in the feed. A high steady state concentration of surfactin (18 mg L^?1) was maintained in the culture at a dilution rate of 0.2 h^?1 when glucose concentration in the feed was 0.25 g L^?1. This is the first demonstration of continuous surfactin production and recovery using glucose as a carbon source. The production of surfactin is known to be related to the age of the microorganisms and a simple mathematical model has been constructed to show how the age‐related production can be quantified. Copyright © 2006 Society of Chemical Industry     

Performance evaluation of batch and unsteady state fed‐batch reactor operations for the production of a marine microbial surfactant

BACKGROUND: Biosurfactants are microbially derived surface‐active and amphipathic molecules produced by various microorganisms. These versatile biomolecules can find potential applications in food, cosmetics, petroleum recovery and biopharmaceutical industries. However, their commercial use is impeded by low yields and productivities in fermentation processes. Thus, an attempt was made to enhance product yield and process productivity by designing a fed‐batch mode reactor strategy. RESULTS: Biosurfactant (BS) production by a marine bacterium was performed in batch and fed‐batch modes of reactor operation in a 3.7 L fermenter. BS concentration of 4.61 ± 0.07 g L^?1 was achieved in batch mode after 22 h with minimum power input of 33.87 × 10³ W, resulting in maximum mixing efficiency. The volumetric oxygen flow rate (K_La) of the marine culture was about 0.08 s^?1. BS production was growth‐associated, as evident from fitting growth kinetics data into the Luedeking‐Piret model. An unsteady state fed batch (USFB) strategy was employed to enhance BS production. Glucose feeding was done at different flow rates ranging from 3.7 mL min^?1 (USFB‐I) to 10 mL min^?1 (USFB‐II). USFB‐I strategy resulted in a maximum biosurfactant yield of 6.2 g l^?1 with an increment of 35% of batch data. The kinetic parameters of USFB‐I were better than those from batch and USFB‐II. CONCLUSION: Comparative performance evaluation of batch and semi‐continuous reactor operations was accomplished. USFB‐I operation improved biosurfactant production by about 35% over batch mode. USFB‐I strategy was more kinetically favorable than batch and USFB‐II. © 2012 Society of Chemical Industry     

Impinging-Jet Ozone Bubble Column Modeling: Hydrodynamics,Gas Hold-up,Bubble Characteristics,and Ozone Mass Transfer

A transient back flow cell model was used to model the hydrodynamic behaviour of an impinging-jet ozone bubble column. A steady-state back flow cell model was developed to analyze the dissolved ozone concentration profiles measured in the bubble column. The column-average overall mass transfer coefficient, k_La (s^?1), was found to be dependent on the superficial gas and liquid velocities, u_G (m.s^?1) and u_L (m.s^?1), respectively, as follows: k_La?=?55.58 · u_G ^1.26· u_L ^0.08 . The specific interfacial area, a (m^?1), was determined as a = 3.61 × 10³ · u_G ^0.902 · u_L ^?0.038 by measuring the gas hold-up (ε _G?=?4.67 · u_G ^1.11 · u_L ^?0.05 ) and Sauter mean diameter, d_S (mm), of the bubbles (d_S?=?7.78 · u_G ^0.207 · u_L ^{? 0.008} ). The local mass transfer coefficient, k_L (m.s^?1), was then determined to be: k_L?=?15.40 · u_G ^0.354 · u_L ^0.118 .     

Utilization of Oil Industry Residues for the Production of Rhamnolipids by Pseudomonas indica

In the present work, a new strain Pseudomonas indica MTCC 3714 was studied for the production of biosurfactants using various rice‐bran oil industry residues viz. rice‐bran, de‐oiled rice‐bran, fatty acids and waxes. Among all the carbon sources, a maximum reduction in surface tension (26.4 mN/m) was observed when the media were supplemented with rice‐bran and the biosurfactant was recovered using the ultrasonication technique as one of the steps in the extraction process. Biosurfactants were obtained in yields of about 9.6 g/L using rice‐bran as the carbon source. The structure of the biosurfactants as characterized by FT‐IR, NMR (¹H and ¹³C) and LC–MS analysis revealed that the majority of the biosurfactants were di‐rhamnolipids. The biosurfactants produced were able to emulsify various hydrocarbons and showed excellent potential in microbial enhanced oil recovery, as it was able to recover kerosene up to 70 % in a sandpack test.     

Factors affecting biosurfactant production using Pseudomonas aeruginosa CFTR-6 under submerged conditions

Studies on the biosurfactants produced by Pseudomonas aeruginosa CFTR-6 revealed that they consisted of glycolipids R-1 and R-2. A time course study of fermentation indicated that the appearance of glycolipids in the fermentation broth coincided with the exhaustion of nitrogen and the commencement of the stationary phase with respect to biomass. The effect of variation of the media components such as carbon, nitrogen, phosphate and metal ions has been investigated. The following values were found to be optimum for biosurfactant production: glucose, 20 g dm^?3; carbon to nitrogen ratio, 38; phosphate, 30 mmol dm^?3; MgSO₄.7H₂O, 100 mg dm^?3. Addition of iron to the medium significantly diminished the glycolipid yield.     

Cells Were a More Important Foaming Factor than Free Rhamnolipids in Fermentation of Pseudomonas aeruginosa E03-40 for High Rhamnolipid Production

Rhamnolipids are among the best‐known biosurfactants. Severe foaming occurs in aerobic rhamnolipid fermentation and negatively affects operation and economics of the biosurfactant production. In this study the foaming properties were examined with samples taken along a Pseudomonas aeruginosa fermentation that produced 55 g l^?1 rhamnolipids with a maximum volumetric productivity of 0.080 g l^?1 h^?1 and a maximum specific productivity of 0.013 g g^?1 h^?1. For a better understanding of the process, the broth samples were also centrifuged to prepare cell‐free supernatants and cell suspensions in water, and all samples were evaluated under fixed foaming conditions. In addition to the time profiles of foam rise, the initial foaming rates and maximum foam volumes were determined. Contrary to the general assumption, the cells, not rhamnolipids, were the main foaming agents in the fermentation. Soluble components including rhamnolipids had secondary roles. Supernatant foaming was higher after the culture entered the rhamnolipid‐producing stationary phase; however, the foaming appeared to decrease with increasing rhamnolipid concentrations at high concentrations (>15 g l^?1). The pH effects on foaming of broths, supernatants, and cell suspensions were also studied. Broth foaming was 55 and 80 % less at pH 5.5 and 5.0, respectively, compared to that at pH 6.5. Cell growth and rhamnolipid production at lower pH should be included in future studies. In addition, strain selection or genetic engineering and medium modification to reduce cell hydrophobicity are suggested as useful strategies to address the foaming issue of rhamnolipid fermentation.