Surfactin Recovery Research Articles

Fouling Analysis in One-Stage Ultrafiltration of Precipitation-Treated Bacillus subtilis Fermentation Liquors for Biosurfactant Recovery.

Primary recovery of surfactin from precipitation-pretreated fermentation broths of Bacillus subtilis ATCC 21332 culture by one-stage dead-end and cross-flow ultrafiltration (UF) was studied. Dead-end experiments were first performed to select suitable conditions, including the amount of added ethanol-a micelle-destabilizing solvent (0-70 vol%), type (polyethersulfone, polyacrylonitrile, poly(vinylidene fluoride)) and molecular-weight cut-off (MWCO, 30-100 kDa) of the membrane in the surfactin concentration range of 0.25-1.23 g/L. Then, the cross-flow UF experiments were conducted to check the recovery performance in the ranges of feed surfactin concentration of 1.13-2.67 g/L, flow velocity of 0.025-0.05 m/s, and transmembrane pressure of 40-100 kPa. The Hermia model was also used to clarify membrane fouling mechanisms. Finally, three cleaning agents and two in situ cleaning ways (flush and back-flush) were selected to regain the permeate flux. As for the primary recovery of surfactin from the permeate in cross-flow UF, a polyethersulfone membrane with 100-kDa MWCO was suggested, and the NaOH solution at pH 11 was used for membrane flushing.

A prospection on membrane-based strategies for downstream processing of surfactin

Abstract Surfactin is one of the most powerful biosurfactants (surface activity, antiviral, antibiotic, and antitumor properties). Synthetic surfactants are derived from the petroleum industry, a finite source. However, the production costs of surfactin, mainly downstream processing, hinder its application at the industrial scale. Two-step ultrafiltration (UF) using polymeric membranes reaches a high recovery yield (>67%) and purity (up to 96%). This review aimed to critically discuss the current state of the art and future trends on surfactin recovery by membrane strategies, in particular the unexplored ceramic membranes. Although ceramic membranes have been successfully used in the separation of phospholipids and other biocompounds, only polymeric membranes have been studied in the downstream processing of surfactin. In addition to their proven potential in the separation of biocompounds, ceramic membranes still have higher chemical resistance that allows the use of organic solvents, such as those used in the purification of surfactin by two-stage ultrafiltration; and also allow for more aggressive chemical cleaning, longer lifetime, and consequently lower maintenance costs, which makes them more attractive economically, in particular for a long-term operation.

Influence of liquid phase hydrodynamics on biofilm formation on structured packing: Optimization of surfactin production from Bacillus amyloliquefaciens

This work gives a useful insight of hydrodynamic aspects within single-species biofilm reactor with metal structured packing and proposes an original methodology based on high energy X-ray tomography to characterize liquid and biofilm distribution. It has been observed that efficiency of initial liquid distribution controls the biofilm distribution within the packing. This parameter mainly depends on the liquid distributor design. The local liquid velocity within the packing element, depending of the recirculation flow rate, the structural properties of the packing and the bed void fraction distribution, have an effect on initial cell attachment and subsequent biofilm growth. However, once started the process efficiency depends mainly on the biofilm spreading on the packing, affecting further liquid distribution. The liquid flow rate is involved in a physicochemical effect leading to foam formation. The latter decreases surfactin recovery in the liquid phase and provokes biofilm detachment of the packing. Thus, the best surfactin yield production, up to 300mg/L, occurs for a liquid load of 17.6L/m2min supplied by a multiple point source distributor. From a technical point of view, these operating conditions are particularly interesting since foam formation is completely suppressed.

An efficient method for separation of surfactin from Bacillus amyloliquefaciens fmb50 broth by flocculation

An efficient method for removing microbial cells and macromolecular impurities and purification of surfaction from fermentation broth produced by Bacillus amyloliquefaciens fmb50 was carried out in this study. Among three inorganic flocculants and a macromolecular flocculants, the combination use of CaCl2 and Na2HPO4 was the most effective separation process. Addition of 50% ethanol into fermentation broth could not only disrupt the surfactin micells, but also promote the permeating of surfactin in filtration. The flocculation condition was optimized by an L9 (34) orthogonal design. The light transmittance, surfactin recovery rate, protein removal rate and filtration flux could reach to 96.3%, 95.31%, 56.59% and 3204.41Lm−2h−1 respectively, the surfactin purity reached to 79.5% and the residual protein was 8.1% in separated product under the optimal flocculation condition (flocculants dosage 0.5%, pH 5.0, and temperature 35°C). Validation test also demonstrated stable results under the optimal conditions. Due to higher efficiency, lower cost and scale-up more easily of flocculation and filtration processes, it is feasible to separate surfactin from fermentation broth.

Integrated Process for Production of Surfactin (III) Modeling of Adsorption Column

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.

Biosurfactants production by immobilized cells of Bacillus subtilis ATCC 21332 and their recovery by pertraction

Microbial production and isolation of biosurfactants was studied. The production of lipopeptides surfactin and fengycin was performed by free and immobilized aerobic cells of Bacillus subtilis ATCC 21332. After preliminary tests with 5 polymer materials, the particles of polypropylene foamed with powder activated carbon (PPch) were selected for lipopeptides production for their thermal and mechanical stability and for the high colonizing effect. To avoid foaming during biosurfactant production, biofilm grown on solid floating support was aerated by air injected over the surface of cultural medium. The synthesis of both lipopeptides and especially of the fengycin was greatly enhanced by the immobilization. The relationship between support wettability, colonization of the cells, and lipopeptide production was discussed. Extraction behaviour of the lipopeptides into alkanes was studied. The distribution ratio of surfactin was found to be higher than this of fengycin at the same conditions and the n-heptane was more efficient solvent for both lipopeptides. Kinetics of surfactin recovery from fermentation broth applying batch pertraction in a rotating discs contactor was studied. Lipopeptide was successfully extracted (more than 75% in the first hour) using n-heptane as liquid membrane and a 0.2 mol L −1 phosphate buffer solution (pH ∼ 7.3) as receiving solution. However, the stripping of the organic liquid and surfactin accumulation into the receiving phase were less efficient.

A further study of the recovery and purification of surfactin from fermentation broth by membrane filtration

Surfactin is a bacterial lipopeptide produced by Bacillus subtilis and is a powerful surfactant, having also antiviral, antibacterial and antitumor properties. The recovery and purification of surfactin from complex fermentation broths is a major obstacle to its commercialization; therefore, a two-step membrane filtration process was developed using a lab scale tangential flow filtration (TFF) unit with 10 kDa MWCO regenerated cellulose (RC) and polyethersulfone (PES) membranes at three different transmembrane pressure (TMP) of 1.5 bar, 2.0 bar and 2.5 bar. Two modes of filtrations were studied, with and without cleaning of membranes prior to UF-2. In a first step of ultrafiltration (UF-1), surfactin was retained effectively by membranes at above its critical micelle concentration (CMC); subsequently in UF-2, the retentate micelles were disrupted by addition of 50% (v/v) methanol solution to allow recovery of surfactin in the permeate. Main protein contaminants were effectively retained by the membrane in UF-2. Flux of permeates, rejection coefficient ( R) of surfactin and protein were measured during the filtrations. Overall the three different TMPs applied have no significant effect in the filtrations and PES is the more suitable membrane to selectively separate surfactin from fermentation broth, achieving high recovery and level of purity. In addition this two-step UF process is scalable for larger volume of samples without affecting the original functionality of surfactin, although membranes permeability can be affected due to exposure to methanolic solution used in UF-2.

Liquid membrane extraction of bio-active amphiphilic substances: Recovery of surfactin

The interest of application of liquid membrane (pertraction) processes for recovery of biosurfactants from aqueous media was demonstrated. Transport of pure surfactin in three-liquid-phase system was studied. Surfactin was successfully extracted from slightly acid media (pH 5.65–6.05) applying batch pertraction in a rotating discs contactor and using n-heptane as liquid membrane. The process efficiency was found to be strongly affected by the feed solution acidity (83% at pHF 6.05 and 97% at pHF 5.65 after 4 h pertraction). An atypical pH effect was observed when the behaviour of surfactin extraction from aqueous media by non-polar solvents (n-heptane and n-octane) was studied. The obtained high extraction degrees from both acid and basic media and the clearly reduced degree of extraction from neutral media could be attributed to the different conformations of surfactin in these media.

Purification of surfactin in pretreated fermentation broths by adsorptive removal of impurities

An attempt was made to further improve the purity of surfactin in the fermentation broth of Bacillus subtilis ATCC 21332 culture by adsorption or ion exchange after the broth had been treated by a two-stage ultrafiltration process. The commonly used neutral adsorption resin (macroporous XAD-7) and charged ion-exchange resin (AG1-X4) were adopted. Under the conditions tested the impurities in the treated broth such as macromolecules (proteins, polysaccharides, peptides) or smaller molecules (glycine, serine, threonine, alanine) were adsorbed or exchanged more quickly and preferably within 5 h of operation, purification of surfactin was hence achieved by adsorption removal of the impurities. It was shown that XAD-7 resins gave higher purity of surfactin than AG1-X4 resins. The recovery of surfactin from the remaining solution after adsorption using XAD-7 resin exceeded 95% at pH 6.5 and the purity increased from 76% (the feed) to 88%; however, the purity of surfactin reached up to 80% only after ion exchange using AG1-X4 resin. Finally, the breakthrough phenomena of surfactin solutions in a XAD-7 column were tested and discussed.

Flux decline and membrane cleaning in cross-flow ultrafiltration of treated fermentation broths for surfactin recovery

Recovery of surfactin from the fermentation broths with culture of Bacillus subtilis ATCC (American Type Culture Collection) 21332 by cross-flow ultrafiltration (UF) process was experimentally examined, in which the broth was pretreated by acid precipitation and the precipitate was then dissolved in NaOH. Two types of membranes, polyethersulfone (PES) and cellulose ester (CE), with a molecular weight cut-off (MWCO) of 100 kDa were compared, and the effect of the presence of micelle-destabilizing solvent ethanol on the performance was studied. Experiments were performed at different initial concentrations of surfactin (1.13–4.57 g/L), transmembrane pressures (TMP, 20–100 kPa), and cross-flow velocities (0.16–0.48 m/s). Under the conditions investigated, the flux increased with increasing cross-flow velocity, but decreased with increasing the initial surfactin concentration and TMP. Finally, four cleaning agents and two cleaning ways (flushing and back-flushing) were screened to recover the flux. From the viewpoint of maximizing surfactin recovery, flushing with NaOH solution at pH 11 was suggested to clean the fouled PES membrane in this cross-flow UF process.

Integrated process for production of surfactin: Part 2. Equilibrium and kinetic study of surfactin adsorption onto activated carbon

Previous work has presented kinetics of pure surfactin adsorption onto activated carbon. Being an efficient biosurfactant, the lipopeptide surfactin has been produced in a bioprocess supported by the strain Bacillus subtilis ATCC 21332. This work is aimed at studying surfactin recovery directly from the culture medium. A thermodynamic study is carried out. Referring to adsorption capacity, the thermodynamic study confirmed that the adsorption of pure surfactin is an exothermic process. The capacity of surfactin adsorption from culture media shows that the activated carbon could be used as efficient adsorbent for surfactin recovery in an integrated process. The study shows the importance of the temperature for process control. Aimed at fixed bed column design, surfactin adsorption modelling on a single microporous pellet is demonstrated.

Recovery of surfactin from fermentation broths by a hybrid salting-out and membrane filtration process

Surfactin biosurfactant was produced by fermentation method with culture of Bacillus subtilis ATCC 21332. Downstream processing for surfactin recovery by ammonium sulfate salting-out, ultrafiltration (UF), nanofiltration (NF), and their hybrid process was studied. Experiments were performed at different initial concentrations of surfactin (210–3620 mg/L), the concentrations of added ammonium sulfate (0–46%, w/v) and micelle-destabilizing solvent ethanol (0–44%, v/v), and membrane molecular-weight cut-off (MWCO, 1–300 kDa) were investigated. When 23% (w/v) ammonium sulfate and 33% (v/v) ethanol were added to the broths, it was shown that surfactin micelles could be efficiently destroyed and other protein macromolecules could be removed. On the other hand, the UF membrane with MWCO less than 100 kDa was found to be suitable for the retention of surfactin micelles, and the NF membrane with a MWCO less than 1 kDa was found to be suitable for the retention of surfactin monomers. Finally, the hybrid salting-out and membrane filtration (UF or NF) process was proposed and tested. Comparisons among these single and hybrid processes demonstrated that the hybrid process not only enhanced the recovery yield but also improved the purity of surfactin.

Recovery and separation of surfactin from pretreated fermentation broths by physical and chemical extraction

The recovery of surfactin from fermentation broths with the culture of Bacillus subtilis ATCC 21332 by physical and chemical extraction was studied, in which the broths were pretreated by acid precipitation and, if necessary, the precipitate was further dissolved in NaOH solution. The physical solid–liquid and liquid–liquid extractions were performed with different organic solvents (ethyl acetate, n-hexane) and at different times of extraction run. It was shown that better extraction was obtained with ethyl acetate than n-hexane. The extraction could be improved by increasing the times of extraction run under a given volume of the organic solution. For chemical liquid–liquid extraction of surfactin with Aliquat 336 (5–200 mM) in n-hexane, the efficiency was improved. The amounts of inorganic salt in the strip solution were also optimized. Surfactin recovered after physical or chemical extraction was finally characterized by mass and NMR spectroscopies. The results of chemical extraction showed that surfactin would readily bind with the quaternary ammonium cations of Aliquat 336.

Separation of surfactin from fermentation broths by acid precipitation and two-stage dead-end ultrafiltration processes

Separation and recovery of surfactin from fermentation broths with the culture of Bacillus subtilis ATCC 21332 by two-stage ultrafiltration (UF) or nanofiltration (NF) processes was studied, in which the broths were pre-treated by acid precipitation and then the precipitate was dissolved in NaOH at pH 11. Experiments were performed at different initial concentrations of surfactin (210–3620 mg/L), concentrations of added micelle-destabilizing solvent ethanol (0–44%, v/v), membrane molecular-weight cut-off (MWCO, 1–300 kDa), and transmembrane pressures (86.184–517.106 × 10 3 Pa (12.5–75 psi)). Under the concentration ranges tested, surfactin micelles could be efficiently destroyed when more than 33% (v/v) ethanol was added to the broth. The UF membranes with MWCO less than 100 kDa were found to be suitable for the retention of surfactin micelles, and the NF membrane with a MWCO less than 1 kDa was suitable for the retention of surfactin monomers. Finally, the separation strategy involving two-stage membrane filtration (UF or NF) processes was proposed. Depending on the forms of the final product, it was shown that more than 72% of recovery yield and more than 83% of purity could be achieved.

Recovery and purification of surfactin from fermentation broth by a two-step ultrafiltration process

Surfactin is a bacterial lipopeptide produced by Bacillus subtilis and it is a powerful surfactant, having also antiviral, antibacterial and antitumor properties. The recovery and purification of surfactin from complex fermentation broths is a major obstacle to its commercialization; therefore, two-step membrane filtration processes were evaluated using centrifugal and stirred cell devices while the mechanisms of separation were investigated by particle size and surface charge measurements. In a first step of ultrafiltration (UF-1), surfactin was retained effectively by membranes at above its critical micelle concentration (CMC); subsequently in UF-2, the retentate micelles were disrupted by addition of 50% (v/v) methanol solution to allow recovery of surfactin in the permeate. Main protein contaminants were effectively retained by the membrane in UF-2. Ultrafiltration was carried out either using centrifugal devices with 30 and 10 kDa MWCO regenerated cellulose membranes, or a stirred cell device with 10 kDa MWCO polyethersulfone (PES) and regenerated cellulose (RC) membranes. Total rejection of surfactin was consistently observed in UF-1, while in UF-2 PES membranes had the lowest rejection coefficient of 0.08 ± 0.04. It was found that disruption of surfactin micelles, aggregation of protein contaminants and electrostatic interactions in UF-2 can further improve the selectivity of the membrane based purification technique.

Integrated process for production of surfactin: Part 1: Adsorption rate of pure surfactin onto activated carbon

The work reported in this paper is aimed at studying the adsorption of surfactin from aqueous solution onto activated carbon. Among the factors, agitation rate, activated carbon particle-size, pH, temperature, initial adsorbate concentration, adsorbent amount and ionic strength of the solution were studied. Both adsorption equilibrium and kinetics showed that activated carbon acted as a suitable adsorbent for surfactin recovery. Two mechanisms represented by different kinetic models were examined, namely, the intraparticle diffusion one and the one involving chemisorption accompanied by surface coverage (conforming to the Elovich concept).

Characterization of concentration and purification parameters and operating conditions for the small-scale recovery of surfactin

Batch recovery including concentration and purification of surfactin from the cell-free fermentation broth was carried out using an ultrafiltration membrane in a magnetically stirred cell system. The filtration characteristics of surfactin on passing through the polymeric membrane, such as permeate flux, rejection coefficient, concentration factor, solute yield or purification factor and recovery factor were evaluated. The effects of the trans-membrane pressure, initial surfactin concentration and the pH of the surfactin solution on the permeate flux were also determined. A maximum of 166-fold concentration of surfactin was rapidly achieved under the optimum trans-membrane pressure and solution pH. The feed surfactin concentration of about 0.5 g L −1 resulted in the formation of a gel polarized layer which was difficult to remove by magnetic stirring, thereby decreasing the permeate flux and the solute yield. The present investigations also qualitatively compare the purity of the final product obtained through solvent extraction and ultrafiltration. The purity of surfactin obtained by ultrafiltration was found to be 70%, as calculated from critical micelle concentration value (CMC) of 0.017 g L −1 against that of 0.013 g L −1 for surfactin (Sigma, 98% pure), used as the reference standard. Thus, an understanding of ultrafiltration characteristics and operating conditions from the analyses of the present study constitutes a very important milestone for process optimization and scale-up.

Surfactin Production from Potato Process Effluent by <I>Bacillus subtilis</I> in a Chemostat

The biosurfactant surfactin has potential to aid in the recovery of energy resources (oil recovery) or subsurface organic contaminants (environmental remediation). However, high medium and purification costs limit its use in these high-volume applications. In previous work, we showed that surfactin could be produced from an inexpensive low-solids potato process effluent with minimal amendments or pretreatments. Previous research has also shown that surfactin can be both produced in Bacillus subtilis cultures and recovered by foam fractionation in an airlift reactor. Results using both purified potato starch and unamended low-solids potato process effluent as substrates for surfactin production indicate that the process is oxygen limited and that recalcitrant indigenous bacteria in the potato process effluent hamper continuous surfactin production. The research reported here features the use of a chemostat operated in batch mode for producing surfactin with concomitant use of antifoam to prevent surfactant loss. The antifoam did not interfere with surfactin recovery by acid precipitation or its efficacy. Initial trials took about 48 h to produce 0.9 g/L of surfactin from potato process effluent. Increasing the oxygen mass transfer by increasing the stirring speed and adding a baffle decreased production time to 12-24 h and produced about 0.6 g/L of surfactin from two different potato-processing facilities.

The application of foaming for the recovery of Surfactin from B. subtilis ATCC 21332 cultures.

Foaming, a proficient method for the recovery of surface active solutes from dilute solutions, was successfully applied for the concentration of the lipopeptide biosurfactant Surfactin from B. subtilis ATCC 21332 cell culture broths. Foaming was only partially successful in concentrating Surfactin when applied as a separate semi-batch unit downstream of the cell culture stage. Surfactin partitioned strongly into the foam during the latter stages of the semi-batch process, where enrichments of over 50 could be obtained. However, simultaneous high enrichments and recoveries of Surfactin could not be obtained as the majority of Surfactin (around 70% of the total recovered) was produced at a low concentration during the early stages of foaming. Foam fractionation was considered for both cell free and cell containing broths; the presence of cells increased the foamability of the solution and therefore yielded more dilute Surfactin preparations. More favourable recovery and enrichment of Surfactin occurred when foaming was integrated with the cell culture stage. The use of low stirrer speeds was essential in producing foam at a controlled rate. By collecting fractions of the foam produced between 10 and 30 hours, from systems stirred at 166 and 146 rpm, a highly concentrated Surfactin extract could be obtained. The Surfactin concentration in the foam was 1.22 and 1.67 g l(-1) respectively, which represented enrichments and percent recoveries of over 60. This study points to the utility of foaming as a method for the recovery of surface-active fermentation products, particularly when used in an integrated production/recovery system.