Evaluation of production of lipopeptide biosurfactants and surfactin micelles by native Bacillus of Iran,for a broader application range

Surfactin is one of the most important lipopeptide biosurfactants obtained by biocatalysts of Bacillus subtilis. The aim of this study was to isolate surfactin-producing bacilli from native soils of the country (Iran), investigate their properties, convert surfactin to surfactin micelles, determine the properties of surfactin micelles and investigate the effect of starch-coated Fe⁰ and Fe³⁺ nanoparticles on the production of surfactin from B. subtilis. To do so, 20 bacilli were isolated from the native soil sample by heat shock method and genome sequenced by SrRNA16 method. The samples with strong β-hemolysis activity were screened as surfactant-producing strains. Two species of 61 and 63 (B. subtilis subspecies. Inaquosorum) were selected and examined by quantitative and qualitative screening tests such as hemolysis activity, surfactin production, droplet aggregation, emulsifying activity, and surface tension reduction in MSM medium containing Fe⁰ and Fe³⁺ nanoparticles. Surfactin was converted to surfactin micelles by sonication and confirmed by SEM. The antimicrobial and emulsifying activity and surface tension reduction of surfactin micelles were investigated. According to the results, the surface tension reduction of surfactin micelles was greater than that of surfactin. The strain 61 (99.7%) culture in 5 L bioreactors containing Fe³⁺ nanoparticles produced more surfactin than culture of the same strain without nanoparticles. This study presents an efficient method to increase the biosynthesis of microbial metabolites.     

Surfactant Effects on Aeration Performance of Stirred Tank Reactors

The effect of surfactants on aeration performance in stirred tank reactors (STR) at high rates of foaming is studied. The volumetric oxygen transfer coefficient (k_La) and foaming activity estimated as foaming height (H_f) were determined. Biotechnology of lipopeptide biosurfactants from aerobic organisms, e.g., Bacillus subtilis were addressed. Using model solutions of known foam‐generating properties, high‐molecular weight surfactin and low‐molecular weight sodium dodecyl sulphate (SDS), as well as impellers of different types, with flat and fluid‐foil blades, clues on the concentration dependence of STR oxygen transfer and foaming as well as options for foam reduction in the presence of biosurfactant were sought. In response to a two‐fold decrease of surface tension by surfactin, k_La values decreased up to 30 % but remained within the range expected for the mixing system in water; the experiments with SDS showing stronger dependence on surfactant concentration and surface tension. Mixing of surfactant media by a standard six‐blade disc turbine (RT) imposed rate limitations on gassing. A low‐shear impeller Narcissus (NS) could be used to avoid bulk foam outflow, while preserving k_La values that remained unchanged. The ‘power per unit volume' correlation of k_La in stirred tanks is tested in the presence of surfactin.     

表面活性素集成生产过程(Ⅲ)吸附柱的模型

The study is focused on modeling of separation process and optimization.An adsorption separation process is simulated.The surfactin production process by Bacillus subtilis ATCC 21332 followed by surfactin adsorption in a fixed-bed column packed with commercial active carbon is studied in laboratory.The adsorption column achieves high surfactin recovery(94%)by up-flow methanol elution at 25°C.The adsorption column is simulated with a complex one-dimensional plug flow dispersion model coupled with nonlinear adsorption equilibrium,based on the assumption that the adsorption of surfactin is monomolecular layer and no micelle is formed.The molecular diffusion coefficient of surfactin in water solution with electric neutrality is estimated to be 0.428×10 -5 cm 2 ·s -1 by molecular dynamics simulation.The model developed can describe the complex interplay of adsorption kinetics,fluid dynamics,and mass-transfer phenomena based on the assumption of no radial temperature and concentration gradients,and is of adequate precision.The work involved in this paper is valuable for the optimization of the production process of surfactin.     

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     

Effects of surface active agents on hydrodynamics and mass transfer characteristics in a split-cylinder airlift bioreactor with packed bed

The effects of three types of surface active agents (containing SDS, HCTBr and Tween 40) with various concentrations (0–5 ppm) on the hydrodynamic and oxygen mass transfer characteristics in a split-cylinder airlift bioreactor with and without packing were investigated. It was observed that in the surfactant solutions, surface tension of the liquid decreased and smaller bubbles were produced in comparison with pure water. So, surfactants presence strongly enhanced mixing time and gas hold-up although oxygen mass transfer coefficient and the liquid circulation velocity reduced. Furthermore, the packing installation enhanced the overall gas–liquid volumetric mass transfer coefficient by increasing flow turbulency and Reynolds number compared to an unpacked column. The packing increased gas hold-up and decreased bubbles size and liquid circulation velocity.     

Scale-up of a cyclone bioreactor

The operation of a cyclone bioreactor differs from conventional stirred tanks since the agitation is accomplished by means of a pumped reciculation loop. Oxygen transfer can occur across the swirling gas—liquid interface in the cyclone or from bubbles entrained in the recirculation loop. A cyclone bioreactor was scaled-up from a 1 dm³ bench top unit to a 75 dm³ Process Development Unit (PDU). A reduction in the aspect ratio was compensated for by extending the length of the recirculation loop and providing additional aeration. Performance of the two reactors for the production of microbial poly-β-hydroxybutyrate (PHB) was compared under various operation conditions. The culture used for PHB production was Alcaligenes eutrophus DSM 545, grown on a mineral salts medium limited by the supply of nitrogen. The levels of dissolved oxygen obtained in the PDU were strongly dependent on the location at which the air was introduced into the reactor. However, with aeration balanced between two injection points and a similar level of power input, 17 J s^?1 dm^?3, the PDU was able to provide at least as much oxygen transfer capability as the laboratory-scale reactor. Under all conditions tested, the PHB accumulation by A. eutrophus was in excess of 80% of the biomass dry weight, although the yield on glucose was lower in the PDU than in the laboratory-scale reactor.     

Removal of mercury by foam fractionation using surfactin, a biosurfactant

The separation of mercury ions from artificially contaminated water by the foam fractionation process using a biosurfactant (surfactin) and chemical surfactants (SDS and Tween-80) was investigated in this study. Parameters such as surfactant and mercury concentration, pH, foam volume, and digestion time were varied and their effects on the efficiency of mercury removal were investigated. The recovery efficiency of mercury ions was highly sensitive to the concentration of the surfactant. The highest mercury ion recovery by surfactin was obtained using a surfactin concentration of 10 × CMC, while recovery using SDS required < 10 × CMC and Tween-80 >10 × CMC. However, the enrichment of mercury ions in the foam was superior with surfactin, the mercury enrichment value corresponding to the highest metal recovery (10.4%) by surfactin being 1.53. Dilute solutions (2-mg L(-1) Hg(2+)) resulted in better separation (36.4%), while concentrated solutions (100 mg L(-1)) enabled only a 2.3% recovery using surfactin. An increase in the digestion time of the metal solution with surfactin yielded better separation as compared with a freshly-prepared solution, and an increase in the airflow rate increased bubble production, resulting in higher metal recovery but low enrichment. Basic solutions yielded higher mercury separation as compared with acidic solutions due to the precipitation of surfactin under acidic conditions.     

Oxygen mass transfer in bubble columns

The mass transfer of oxygen between air and water has been studied in a bubble column over wide ranges of liquid and gas velocity. An oxygen probe was used to map the steady-state liquid phase concentration of oxygen throughout the column.At any given point in the column, the oxygen concentration increased with gas velocity. Minima were observed in plots of concentration against liquid velocity.Two distinct absorption regions were observed. Close to the distributor the concentration decreased rapidly with height and volumetric mass transfer coefficients ranged from about 0.2 to 2.1 s^?1. These high values were attributed to enhanced mass transfer due to turbulence induced by the liquid and gas jets in the grid region. In the bulk of the column, axial concentration gradients were much smaller and the mass transfer coefficients were up to two orders of magnitude lower than in the grid region.     

Pilot-scale production of bacterial cellulose by a spherical type bubble column bioreactor using saccharified food wastes

Bacterial cellulose (BC) was produced by Acetobacter xylinum KJ1 in a modified airlift-type bubble column bioreactor, which had a low shear stress and high oxygen transfer rate (k _L a). Saccharified food wastes (SFW) were used as the BC production medium due to its low cost. An aeration rate of 1.2 vvm (6 L/min) was tentatively determined as the optimal aeration condition in a 10 L spherical type bubble column bioreactor, by analysis of the oxygen transfer coefficient. When 0.4% agar was added, the BC production reached 5.8 g/L, compared with 5.0 g/L in the culture without the addition of agar. The BC productivity was improved by 10% in the addition of 0.4% agar into the SFW medium. Then, by conversion of a linear velocity of 0.93 cm/sec, from the relationship between the linear velocity and oxygen transfer rate, 1.0 vvm (30 L/min) was determined as an optimal aeration condition in a 50 L spherical type bubble column reactor. Using SFW medium, with the addition of 0.4% agar, and air supplied of 1.0 vvm, 5.6 g/L BC was produced in the 50 L spherical type bubble column bioreactor after 3 days of cultivation, which was similar to that produced in the 10 L bioreactor. In conclusion, the addition of agar, a viscous polysaccharide, into SFW medium is effective for the production of BC, and this scale-up method is very useful for the mass production in a 50 L spherical type bubble column bioreactor by decreasing the shear stress and increasing the k _L a.     

Mixing of Surfactin,an Anionic Biosurfactant,with Alkylbenzene Sulfonate,a Chemically Synthesized Anionic Surfactant,at the n-Decane/Water Interface

Biosurfactants show synergic effects with synthesized surfactant in reducing hydrophobic/hydrophilic interfacial tension, while the understanding of the synergistic mechanism is limited. In the present work, mixed monolayers of surfactin and branched alkylbenzene sulfonate at the n-decane/water interface were studied using atomistic molecular dynamics simulations, and the presence of surfactin affecting the microstructure and dynamic properties of the mixed monolayer was evaluated at molecular level. The density distributions of the surfactants along the direction normal to the interface, radial distribution functions of the surfactant head groups, hydrophobic contacts between surfactants, translational activities of both surfactants and counterions, and the dynamics of the hydrogen bonds formed between surfactant and water were calculated. The results suggested that the structure of the mixed monolayers was more compact than that of the individual system of alkylbenzene sulfonate and the interfacial tension was more efficiently reduced, and the translational activities of both surfactants within the mixed monolayers were much lower. The results implied that biosurfactant surfactin and alkylbenzene sulfonate mixed well at the n-decane/water interface, though they were both anionic surfactants.     

Effect of surfactants on α‐amylase production in a solid substrate fermentation process

Solid substrate fermentation (SSF) is a process where the substrate is a moist solid, which is insoluble in water but not suspended in water. In this study SSF of Bacillus subtilis (ATCC 21556) was used to produce an enzyme of commercial importance, α‐amylase, using as a substrate potato peel. To enhance the production of this enzyme, two nonionic synthetic surfactants were used, Tween 80 and Tween 20, one anionic surfactant, SDS at concentrations of 0.05% and 0.10% (v/w) and a biosurfactant produced by Bacillus subtilis (ATCC 21332), known as surfactin, at concentrations of 0.003%, 0.007%, 0.013% and 0.03% (w/w). The results have shown that surfactants significantly increase the production of α‐amylase. Tween 80 at 0.10% and surfactin at 0.013% provided the highest enzyme activity when compared with the control. © 1999 Society of Chemical Industry     

Mass transfer in a recirculating bubble column

The mass transfer of oxygen between air and water in a recirculating bubble column has been studied, at gas and water superficial velocities of up to 0.23 m s^-1 and 0.68 m s^-1 respectively. Experiments show that the assumption of plug flow for the gas phase is reasonable, eliminating a possible source of error identified by other workers in calculating the mass transfer rates. The results obtained are consistent with other published work. It is also shown that for the air-water system breaking up the gas bubbles to increase the mass transfer does not have a large effect, due to rapid recoalescence of the bubbles.     

Design and characterization of a multistage,mechanically stirred column absorber

A multistage, mechanically stirred column absorber has been designed and built with a modular construction, based on preliminary experiments with a test column. The column has been characterized as a gas-liquid contactor by its gas holdup, gas and liquid axial dispersion, mixing times, oxygen transfer coefficients and power consumptions, determined as a function of gas velocity, liquid velocity and impeller speed for one and two impellers per stage.Gassed power was correlated with ungassed power, gas rate and impeller speed. The gas phase axial mixing was essentially plug flow and the liquid phase axial mixing varied between 5 and 12 equivalent stages.Oxygen transfer coefficients were correlated with power consumptions and aeration rates by the equation K_La γ (P/V)^a(υ_sg)^b. The oxygen transfer coefficients with single stiffer stages were 25% above those for the double stirrer stages for equal power consumption and gas rates. Except for the low aeration and high power consumption extremes, the column showed superior oxygen transfer performance. in comparison to tubular loop and tank fermenters.     

Surfactin生物表面活性剂及其驱油研究进展

表面活性素（surfactin）是一类由革兰氏阳性的枯草芽孢杆菌产生的脂肽（lipopeptide）型生物表面活性剂,因其具有优于化学合成表面活性剂的若干优点,如低毒性、高生物降解性、更好的环境相容性,且在极端环境下稳定性好,在提高石油采收率方面有较好的应用潜力,但是目前只有少数的生物表面活性剂可以大规模生产实现工业化应用。本文介绍了surfactin生物表面活性剂的化学结构和生物合成机制,并对其发酵生产过程的影响因素进行分析,为提高其生产经济性探索不同的策略,例如使用更便宜的原材料、优化培养基组分、优化反应器等,系统论述了surfactin生物表面活性剂的驱油机理和其与化学合成表面活性剂的复配研究,同时针对其应用时的不足之处提出研究新思路。     

Surfactin from Bacillus velezensis H2O-1: Production and Physicochemical Characterization for Postsalt Applications

The use of surfactin, a powerful biosurfactant, is generally hampered by poor production yield. Consequently, identification of new producers and the study of operational parameters are essential. We identify Bacillus sp. H2O-1 as Bacillus velezensis, a species previously not investigated for its biosurfactant production. Among the nitrogen sources we tested, (NH₄)₂SO₄ and NH₄NO₃ were the most appropriate for surfactin production, reaching 608.5 and 659.5 mg L⁻¹, respectively. Only temperature affected the production, whereas rotation and the C/N ratio did not. Biosurfactants can be used in enhanced oil recovery (EOR) in reservoirs located in the presalt and postsalt layers, where conditions of temperature, pressure, and salinity are quite varied, requiring a study of the stability of these molecules under these conditions. We found the surfactin produced by B. velezensis to be stable at different temperatures, pH, and ionic strengths. We evaluated the concurrent effects of different salinity, temperature, and pressure conditions on surface and interfacial activities of this surfactin. Overall, we found the surfactin produced by B. velezensis H2O-1 to have considerable potential for industrial applications, mainly due to the stability of its physical and chemical characteristics when subjected to different temperatures, pressures, and salinities, in addition to its low toxicity.     

Low-Toxic and Nonirritant Biosurfactant Surfactin and its Performances in Detergent Formulations

Developing eco-friendly, nonirritant, low-toxic, and high-efficient surface active ingredients for detergents is an ongoing challenge in the detergent field. Surfactin is one of the representative lipopeptides produced by microorganisms. In this article, we report the surfactin isolated from cell-free broth of Bacillus subtilis HSO121 and purified by reversed-phase high-performance liquid chromatography for detergent formulations. The biodegradability, acute dermal irritation, acute oral toxicity (LD₅₀ and LC₅₀), surface activity, washing efficiency, and compatibility with hard water of the purified biosurfactant surfactin have been studied to explore the feasibility for applications of the surfactin in detergents. Acute oral toxicity tests (LD₅₀ > 5000 mg kg⁻¹, LC₅₀ > 1000 mg kg⁻¹) and skin irritation tests (PII = 0) indicate that the surfactin is a low-toxic and nonirritant ingredient for detergent formulation. Moreover, the surfactin shows excellent surface and interfacial properties of emulsification and wettability, high compatibility, and stability in a wide range of temperatures, pH, and hard water and acceptable properties in biodegradability and foaming ability, which suggests that the biosurfactant surfactin is a promising ingredient for detergent formations in our daily life and for industrial applications.     

A comparison of oxygen mass transfer into sodium sulphite solution and a biological system

To explain previously reported discrepancies between oxygen uptake rates in biological systems and the sodium sulphite test system a comparison was made between oxygen mass transfer into a sodium sulphite solution catalysed by copper(II) ions, and into a biological system, the bacterium E. coli in a mineral medium. A stirred transfer cell, with a relatively low rate of physical mass transfer per unit area, and a bubble column, with a high physical mass transfer rate per unit area, were used to make this comparison; in both cases, the areas of gas/liquid contact could be determined. In the transfer cell the gas/liquid mass transfer coefficient for the biological system was only 10–25% of that for the sulphite oxidation system (for which the absorption rate was increased by chemical reaction). In the bubble column mass transfer into both systems was controlled by physical absorption and the mean mass transfer coefficients were similar for both systems. However, the mass transfer coefficient for the biological system increased with E. coli concentration, probably due to physical effects of the small particles.     

Comparison of Ozone and Oxygen Mass Transfer in a Laboratory and Pilot Plant Operation

Mass transfer of ozone and oxygen to water was investigated both in pilot plant countercurrent bubble column and in a Rushton type laboratory stirred reactor supplied with a variable speed turbine agitator. A comparison was made for different hydrodynamic conditions with the main task of developing an engineering approach for determination of the physical volumetric mass transfer coefficient (K_L ^oa), specific interfacial area (a), and physical masstransfer coefficient (K_LO). The mass transfer characteristics of ozone and oxygen can be determined quickly in a pilot plant or laboratory apparatus, and employed to optimize the performance ofthe full scale water treatment plant.     

Evaluation of a novel self-aerating,oscillating baffle column

We describe the operation of a pilot scale oscillating baffle column using a self-aeration system for oxygenation of water. The top baffle has a high constriction ratio and is sufficiently near to the surface of the water such that gas bubbles are generated. This aeration plate is coupled with a series of equally spaced low constriction orifice baffles, which lead to uniform dispersion of entrained air bubbles throughout the liquid volume. Flow visualisation experiments using video and still photography were used to identify two mechanisms for bubble generation: bubble formation under the water surface by surface vortex suction, and bubble generation from the collapse of a surface fountain and subsequent entrainment of bubbles into the bulk. Mass transfer measurements have shown that under most conditions a uniform oxygen concentration could be obtained throughout the column as a result of efficient mixing, and that the sole limitation to mass transfer performance was determined by the aeration mechanism. Initial comparison on the basis of aeration efficiency with other devices reveals a modest oxygen transfer rate, but with low specific power consumption of order 0.3 kW/m³.     

Enhanced Wettability Alteration by Surfactants with Multiple Hydrophilic Moieties

The main production mechanism during water flooding of naturally fractured oil reservoirs is the spontaneous imbibition of water into matrix blocks and resultant displacement of oil into the fracture system. This is an efficient recovery process when the matrix is strongly water-wet. However, in mixed- to oil-wet reservoirs, secondary recovery from water flooding is often poor. Oil production can be improved by dissolving low concentrations of surfactants in the injected water. The surfactant alters the wettability of the reservoir rock, enhancing the spontaneous imbibition process. Our previous study revealed that the two main mechanisms responsible for the wettability alteration are ion-pair formation and adsorption of surfactant molecules through interactions with the adsorbed crude oil components on the rock surface. Based on the superior performance of surfactin, an anionic biosurfactant with two charged groups on the hydrophilic head, it was hypothesized that the wettability alteration process might be further improved through the use of dimeric or gemini surfactants, which have two hydrophilic head groups and two hydrophobic tails. We believe that when ion-pair formation is the dominant wettability alteration mechanism, wettability alteration in oil-wet cores can be improved by increasing the charge density on the head group(s) of the surfactant molecule since the ion-pair formation is driven by electrostatic interactions. At a concentration of 1.0 mmol L⁻¹ a representative anionic gemini surfactant showed oil recoveries of up to 49% original oil-in-place (OOIP) from oil-wet sandstone cores, compared to 6 and 27% for sodium laureth sulfate and surfactin, respectively. These observations are consistent with our hypothesis.