Influence of fermentation conditions and microfiltration processes on membrane fouling during recovery of glucuronane polysaccharides from fermentation broths.

We have investigated the recovery of exopolysaccharides produced by Sinorhizobium meliloti M5N1 CS bacteria from fermentation broths using different membrane filtration processes: cross-flow filtration with a 7 mm i.d. tubular ceramic membrane of 0.5-microm pores under fixed transmembrane pressure or fixed permeate flux and dynamic filtration with a 0.2 microm nylon membrane using a 16-cm rotating disc filter. With the tubular membrane, the polysaccharide mass flux was mainly limited by polymer transmission that decayed to 10% after 90 min. The mass flux of polymer produced under standard fermentation conditions (70 h at 30 degrees C) stabilized after 70 min to 15 g/h/m(2). This mass flux rises to 36 g/h/m(2) when the mean stirring speed during fermentation is increased and to 123 g/h/m(2) when fermentation is extended to 120 h. In both cases, the mean molecular weight of polysaccharides drops from 4.0 10(5) g/mol under standard conditions to 2.7 10(5) g/mol. A similar reduction in molecular weight was observed when the fermentation temperature was raised to 36 degrees C without benefit to the mass flux. These changes in fermentation conditions have little effect on stabilized permeate flux, but raise significantly the sieving coefficient, due probably to molecular weight reduction and the filamentous aspect of the polymer as observed from SEM photographs. The polymer-mass flux was also increased by reducing transmembrane pressure (TMP) and raising the shear rate by inserting a rod in the membrane lumen. Operation under fixed permeate flux instead of constant TMP inhibited fouling during the first 4 h, resulting in higher sieving coefficients and polymer mass fluxes. The most interesting results were obtained with dynamic filtration because it allows operation at high-shear rates and low TMP. Sieving coefficients remained between 90 and 100%. With a smooth disc, the polysaccharide mass flux remained close to 180 g/h/m(2) at 1500 rpm and cell concentrations from 1 to 3 g/L. When radial rods were glued to the disc to increase wall shear stress and turbulence, the mass flux rose to 275 g/h/m(2) at the same speed and cell concentration.     

Protein recovery from cell debris using rotary and tangential crossflow microfiltration

Protein recovery from a bacterial lysate was accomplished using microfiltration membranes in a flat crossflow filter and in a cylindrical rotary filter. Severe membrane fouling yielded relatively low long-term permeate flux values of 10(-4)-10(-3) cm/s (where I cm/s = 3.6 x 10(4) L/m(2) - h). The permeate flux was found to be nearly independent of transmembrane pressure and to increase with increasing shear rate and decreasing solids concentration. The flux increased with shear to approximately the one-third power or greater for the flat filter and the one-half power or greater for the rotary filter; the stronger dependence for the rotary filter is thought to result from Taylor vortices enhancing the back transport of debris carried to the membrane surface by the permeate flow. The average protein transmission or sieving coefficient was measured at approximately 0.6, but considerable scatter in the transmission data was observed. The largest sieving coefficients were obtained for dilute suspensions at high shear rate. The rotary filter provided higher fluxes than did the flat filter for dilute suspensions, but not for concentrated suspensions. (c) 1995 John Wiley & Sons, Inc.     

Modulation of transmembrane pressure in manufacturing scale tangential flow filtration N-1 perfusion seed culture

Mammalian cells were grown to high density in a 3,000 L culture using perfusion with hollow fibers operated in a tangential flow filtration mode. The high-density culture was used to inoculate the production stage of a biomanufacturing process. At constant permeate flux operation, increased transmembrane pressures (TMPs) were observed on the final day of the manufacturing batches. Small scale studies suggested that the filters were not irreversibly fouled, but rather exposed to membrane concentration polarization that could be relieved by tangential sweeping of the hollow fibers. Studies were undertaken to analyze parameters that influence the hydrodynamic profile within hollow fibers; including filter area, cell density, recirculation flow rate, and permeate flow rate. Results indicated that permeate flow rate had the greatest influence on modulating TMP. Further evaluation showed a significant decrease in TMP when permeate flow was reduced, and this occurred without any negative effect on cell growth or viability. Hence, a 30% reduction of permeate flow rate was implemented at manufacturing scale. A stable operation was achieved as TMP was successfully reduced by 75% while preserving all critical factors for performance in the perfusion bioreactor.     

Protein transmission during Dean vortex microfiltration of yeast suspensions.

Substantially higher rates of protein and fluid volume transport for microfiltration of yeast suspensions were possible with improved hydrodynamics using centrifugal fluid instabilities called Dean vortices. Under constant permeate flux operation with suspended yeast cells, a helical module exhibited 19 times the filtration capacity of a linear module. For feed containing both BSA and beer yeast under constant transmembrane pressure with diafiltration, about twice as much protein (BSA and other proteins from cell lysis) was transported out of the feed by the helical module as compared with the linear module. The volumetric permeation flux improvements for the helical over the linear module ranged from 18 to 43% for yeast concentrations up to 4.5 dry wt %.     

A model for high-pressure ultrafiltration of blood

A semi-analytic model to predict the permeate flux during high-pressure ultrafiltration of blood with highly permeable membranes is proposed. This model explicitly considers the hydraulic resistance of the retained particles that limits the flux. An empirically derived relationship between particle surface concentration and hydraulic resistance is used. This model incorporates the axial variations in blood cell and solute surface concentrations (or concentration polarization), shear-induced diffusion coefficient for the blood cells, effective diffusion coefficient for the blood solutes, hydraulic (lumen) pressure, and flow rate. This model agrees well with experimental results in the pressure-independent filtration flux region.     

Lysozyme separation by hollow-fibre ultrafiltration

This paper discusses the purification of lysozyme from chicken egg white using hollow-fibre ultrafiltration (30kDa MWCO, polysulphone membrane). Lysozyme is preferentially transmitted through the membrane while the membrane largely retains other egg white proteins. Improvement in system hydrodynamics resulted in an increase in permeate flux while lysozyme transmission remained unaffected, leading to higher productivity. The percentage purity of lysozyme obtained was generally insensitive to system hydrodynamics. The permeate flux and productivity increased with increase in transmembrane pressure (TMP) before levelling off around 0.7bar. However, the TMP did not have any pronounced effect on the transmission and the purity of lysozyme. Experiments carried out in the diafiltration mode showed that moderately pure lysozyme (80-90%) could be obtained in an extended operation.     

Membrane separation of protein precipitates: Studies with cross flow in hollow fibers

Hollow fiber ultrafiltration and microfiltration membranes are examined for the processing of isoelectric soya protein precipitate suspensions. A model based on the various resistances to permeate flux is used to describe membrane performance. The main resistance to permeate flux is due to the interaction between the active membrane and the soluble and precipitated protein; that is, as compared with resistances due to the active membrane itself or the membrane support structure, or arising from concentrated soluble or precipitated protein layers over the membrane surface. Soluble protein rejection and precipitate mean particle diameter are correlated with observed values of this main resistance.In contract to the ultrafiltration of soluble proteins, the flux rates observed when processing protein precipitate suspensions under a similar range of operating conditions do not approach a limiting value with increased transmembrane pressure. At high protein concentrations, greater flux rates may be achieved for precipitated as compared with soluble proteins. The use of a microfiltration membrane does not give further improvement in flux rate; this may be attributed to problems of pore fouling with precipitate particles.     

Flux enhancement for membrane filtration of bacterial suspensions using high-frequency backpulsing

A promising method for reducing membrane fouling during crossflow microfiltration of biological suspensions is backpulsing. Very short backpulses (0.1-1.0 s) have been used to increase the net flux for washed bacterial suspensions and whole bacterial fermentation broths. The net fluxes under optimum backpulsing conditions for the washed bacteria are approximately 10-fold higher than those obtained during normal crossflow microfiltration operation, whereas only a 2-fold improvement in the net flux is achieved for the fermentation broths. A theory is presented that is based on external fouling during forward filtration and nonuniform cleaning of the membrane during reverse filtration. The model contains an adjustable parameter which is a measure of the cleaning efficiency during backpulsing; the cleaning efficiency found by fitting the model to the experiments increases with increasing frequency and duration of the backpulses. The theory predicts an optimum backpulsing frequency, as was observed experimentally. An economic analysis shows that crossflow microfiltration with backpulsing has lower costs than centrifugation, rotary vacuum filtration, and crossflow microfiltration without backpulsing.     

Air slugs entrapped cross-flow filtration of bacterial suspensions

A novel cross-flow technique for membrane filtration of bacterial cell suspensions was established. This is an air slugs entrapped cross-flow method in which air slugs were generated by introducing air into the cross-flow stream. As air slugs moved along with cross-flow, the disturbance of cell sublayer formation on membrane surface was enhanced. As a consequence, filtration flux was improved and stabilized. The effect of air slugs on improving filtration flux was more pronounced in filtering gram-negative Escherichia coli cell than grampositive Brevibacterium flavum cell. Moreover, air slug was about 50% more effective on reducing filtration resistance using ultrafiltration (UF) membrane of 300,000 molecular weight cutoff (MWCO) than microfiltration (MF) membrane of 0.2 mum. (c)1993 John Wiley & Sons, Inc.     

Plasma fractionation by dead-end membrane filtration: effects of membrane properties and plasma flux

Plasma fractionation by membrane filtration permits the reinfusion of the patient with his own albumin. In this study, the influence of membrane nature and plasma flux on plasma fractionation in dead-end mode is investigated with acetate hollow fiber filters. It is found that transmembrane pressure TMP rises exponentially with time, the rate of increase being proportional to plasma flux. The faster TMP rises, the faster the drop in sieving coefficient SC. It is also found that albumin SC is a function of TMP and not of plasma flux. Theoretical analysis of the dead-end filtration was performed. This theoretical model indicates that the observed variation of TMP with time is consistent with the assumptions that pore volume decreases proportionally to the filtrate plasma volume.     

Viability of an ectomycorrhizal fungus during cross-flow filtration

Factors affecting the viability and infectivity of an ectomycorrhizal fungus during moderate concentration by cross-flow filtration were determined. Mycelial suspensions were concentrated with three commercial membrane filters (Prostak Millipore Co., M14 Tech-Sep Co. and Ceraflo Norton Co.) under aseptic conditions. Medium components may reduce the filtration rate due to their low solubility. An antifoam agent did not reduce the average flux rates as much as did the malt extract. Clear unobstructed channels (I.D. 6mm) of the tubular modules (Tech-Sep) gave the best results both in terms of performance (filtration rate) and cell viability. Shear stresses caused by pumping and flow through narrow retentate channels were probably responsible for lowering viability and infectivity. There was no linear relationship between permeate fluxes and cell concentration. There is an optimum pore size both in terms of performance (filtration rate) and cell viability. Physical blockage of large pores by hyphae could explain lower permeate flux rates than those obtained with lower pore sizes membranes.     

Two-phase bioconversion product recovery by microfiltration I. Steady state studies

Recovery of an aqueous bioconversion product from complex, two-phase Pseudomonas putida broths containing 20% (v/v) soybean oil presents a significant challenge for downstream processing. Although not used before in multiple-phase separation for complex biotech products, crossflow filtration employing ceramic filters is one of the most attractive options which allow the design of integrated, continuous bioconversion processes. As a first attempt, we studied multichannel, monolithic ceramic membranes of different nominal pore sizes and lumen diameters under steady-state conditions. The best performance was obtained with 0.2-microm-pore/3-mm-lumen membrane, which completely rejected both cells and oil droplets from the permeate, creating a clear aqueous product stream. Although the same separation was achieved, the 50K molecular weight cut-off (MWCO) ultrafilter showed greater irreversible but similar reversible resistance, in addition to an order-of-magnitude higher membrane resistance. Larger nominal pore microfilters, such as 0.45 and 1.0 microm, experienced both cell and oil leakage even at low transmembrane pressure (10 psig). Attributed to greater shear at the same recirculation rate, smaller lumen filters did provide greater permeate flux. However, for practical purposes, the 0. 2-microm-pore/4-mm-lumen ceramic membrane was chosen for further evaluation. Transmembrane pressures up to 50 psig provided only marginal gains in filtration performance, whereas increasing shear rate resulted in linear increases in steady-state flux, presumably due to formation of shear-sensitive, complex gel/oil/cell layer near the membrane surface. A nominal shear rate of 9200 s-1 and 20 psig transmembrane pressure were chosen as optimal operating conditions. Additional studies in a clean system revealed that as low as 5% (v/v) soybean oil in deionized (DI) water resulted in an order-of-magnitude decline in steady-state permeate flux. Breakthrough of oil droplets occurred at 35 psig transmembrane pressure. The severe fouling and breakthrough phenomena disappeared in the presence of washed cells for transmembrane pressure up to 43 psig, implying an oil/cell layer coating the membrane surface, thus preventing oil penetration. Serious membrane fouling was also experienced in microfiltration of oil-free, cell-free supernatant and oil-free whole broth. Consequently, soluble proteins/surfactants were suspected to be the major membrane foulants. Interestingly, soybean oil up to 30% (v/v) enhanced the flux, presumably through complicated interactions with the major foulants. Regeneration of membrane was best achieved with protease and hot caustic/bleach treatments, supporting the hypothesized fouling mechanisms mentioned above. This work provides process and system information for batch microfiltration runs in the future, to be reported elsewhere as Part II of this work.     

The Effect of Gas Sparging in Cross‐Flow Microfiltration of 2,3‐Butanediol Fermentation Broth

Recovery of 2,3‐butanediol from a fermentation broth entails the separation of cells and other suspended solids as the initial step for subsequent separation stages. The aim of this work was to study the cross‐flow filtration of broth in the fermentation of 2,3‐butanediol from blackstrap molasses by Klebsiella oxytoca (NRRL B‐199). A plate type laboratory scale cross‐flow microfiltration unit with a 0.2‐μm cellulose acetate membrane was employed for this purpose. Preliminary results showed that the permeate flux would decline rapidly due to fouling caused by the natural impurities of blackstrap molasses, and modifications of the conventional cross‐flow filtration would be essential to achieve a filtration rate appropriate for practical purposes. In this work, the permeate flux was enhanced by air sparging, which scoured the membrane surface of colloidal deposits and allowed a practical filtration rate to be maintained. The average permeate flux increased by 39 % and 54 % for an air sparging rate of 0.5 L/min and 1.0 L/min respectively, in the case of an initial biomass concentration of 4.66 g/L. For an initial biomass concentration of 14.2 g/L, the flux increased by 105 % and 146 % for the gas rate of 0.5 and 1.0 L/min, respectively. It may be concluded that gas sparging is beneficial in cross‐flow filtration of thick suspensions like a fermentation broth.     

Enhanced COD and nitrogen removals for the treatment of swine wastewater by combining submerged membrane bioreactor (MBR) and anaerobic upflow bed filter (AUBF) reactor

In order to enhance performances of organics removal and nitrification for the treatment of swine wastewater containing high concentration of organic solids and nitrogen than conventional biological nitrogen removal process, a submerged membrane bioreactor (MBR) was followed by an anaerobic upflow bed filter (AUBF) reactor in this research (AUBF–MBR process). The AUBF reactor is a hybrid reactor, which is the combination of an anoxic filter for denitrification and upflow anaerobic sludge blanket (UASB) for acid fermentation. In the AUBF–MBR process, it showed a considerable enhancement of the effluent quality in terms of COD removal and nitrification. The submerged MBR could maintain more than 14,000 mg VSS/L of the biomass concentration. Total nitrogen (T-N) removal efficiency represented 60% when internal recycle ratio was three times of flow-rate (Q), although the nitrification occurred completely. Although the volatile fatty acids produced in AUBF reactor can enhance denitrification rate, but the AUBF–MBR process showed reduction of overall removal efficiency of the nitrogen due to the reduction of carbon source by methane production in the AUBF reactor compared to that of theoretical nitrogen removal efficiency.

Long-term operation of the submerged MBR showed that the throughputs of the submerged MBR were respectively 74, 63, and 31 days at 10, 15, and 30 L/m² h (LMH) of permeate flux. Resistance to filtration by rejected solid is the primary cause of fouling, however the priority of cake resistance (R_c) and fouling resistance (R_f) with respect to filtration phenomenon was different according to the amount of permeate flux. The submerged MBR, here, achieved a steady-state flux of 15 LMH at 0.4 atm. of trans-membrane pressure (TMP) but the flux can be enhanced in the future because shear force by tangential flow will be greater when multi-layer sheets of membrane were used.     

Comparison of the performance of submerged membrane bioreactor (SMBR) and submerged membrane adsorption bioreactor (SMABR)

This study focuses on comparing the performance of submerged membrane bioreactor (SMBR) and submerged membrane adsorption bioreactor (SMABR) over a period of 20 days at a hydraulic retention time (HRT) of 3.1h. The effects of PAC on critical flux and membrane fouling were also investigated. The SMABR exhibited better results in terms of mixed liquor suspended solids (MLSS) growth, DOC removal (over 96%), COD removal (over 95%), transmembrane pressure (TMP) and oxygen uptake rate. Nearly 100% of bacteria and 100% of total coliforms were removed in both systems. The addition of PAC could maintain the critical flux at a lower TMP value (7.5 kPa), while irreversible fouling caused by PAC occurred when the filtration flux exceeded critical flux.     

Understanding and modeling alternating tangential flow filtration for perfusion cell culture

Alternating tangential flow (ATF) filtration has been used with success in the Biopharmaceutical industry as a lower shear technology for cell retention with perfusion cultures. The ATF system is different than tangential flow filtration; however, in that reverse flow is used once per cycle as a means to minimize fouling. Few studies have been reported in the literature that evaluates ATF and how key system variables affect the rate at which ATF filters foul. In this study, an experimental setup was devised that allowed for determination of the time it took for fouling to occur for given mammalian (PER.C6) cell culture cell densities and viabilities as permeate flow rate and antifoam concentration was varied. The experimental results indicate, in accordance with D'Arcy's law, that the average resistance to permeate flow (across a cycle of operation) increases as biological material deposits on the membrane. Scanning electron microscope images of the post‐run filtration surface indicated that both cells and antifoam micelles deposit on the membrane. A unique mathematical model, based on the assumption that fouling was due to pore blockage from the cells and micelles in combination, was devised that allowed for estimation of sticking factors for the cells and the micelles on the membrane. This model was then used to accurately predict the increase in transmembane pressure during constant flux operation for an ATF cartridge used for perfusion cell culture. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 30:1291–1300, 2014     

Effect of Permeate Drag Force on the Development of a Biofouling Layer in a Pressure-Driven Membrane Separation System

The effect of permeate flux on the development of a biofouling layer on cross-flow separation membranes was studied by using a bench-scale system consisting of two replicate 100-molecular-weight-cutoff tubular ultrafiltration membrane modules, one that allowed flow of permeate and one that did not (control). The system was inoculated with Pseudomonas putida S-12 tagged with a red fluorescent protein and was operated using a laminar flow regimen under sterile conditions with a constant feed of diluted (1:75) Luria-Bertani medium. Biofilm development was studied by using field emission scanning electron microscopy and confocal scanning laser microscopy and was subsequently quantified by image analysis, as well as by determining live counts and by permeate flux monitoring. Biofilm development was highly enhanced in the presence of permeate flow, which resulted in the buildup of complex three-dimensional structures on the membrane. Bacterial transport toward the membrane by permeate drag was found to be a mechanism by which cross-flow filtration contributes to the buildup of a biofouling layer that was more dominant than transport of nutrients. Cellular viability was found to be not essential for transport and adhesion under cross-flow conditions, since the permeate drag overcame the effect of bacterial motility.     

Separation of lactic acid-producing bacteria from fermentation broth using a ceramic microfiltration membrane with constant permeate flow

The influence of several operating parameters on the critical flux in the separation of lactic acid-producing bacteria from fermentation broth was studied using a ceramic microfiltration membrane equipped with a permeate pump. The operating parameters studied were crossflow velocity over the membrane, bacterial cell concentration, protein concentration, and pH. The influence of the isoelectric point (IEP) of the membrane was also investigated. In the interval studied (5.3-10.8 m/s), the crossflow velocity had a marked effect on the critical flux. When the crossflow velocity was increased the critical flux also increased. The bacterial cells were retained by the membrane and the concentration of bacterial cells did not affect the critical flux in the interval studied (1.1-3.1 g/L). The critical flux decreased when the protein concentration was increased. It was found that the protein was adsorbed on the membrane surface and protein retention occurred even though the conditions were such that no filter cake was present on the membrane surface. When the pH of the medium was lowered from 6 to 5 (and then further to 4) the critical flux decreased from 76 L/m(2)h to zero at both pH 5 and pH 4. This was found to be due to the fact that the lowering in pH had affected the physiology of the bacterial cells so that the bacteria tended to adhere to the membrane and to each other. The critical flux, for wheat flour hydrolysate without particles, was much lower (28 L/m(2)h) when using a membrane with an IEP of 5.5 than the critical flux of a membrane with an IEP at pH 7 (96 L/m(2)h). This was found to be due to an increased affinity of the bacteria for the membrane with the lower IEP.     

Comparing membrane and spacer biofouling by Gram-negative Pseudomonas aeruginosa and Gram-positive Anoxybacillus sp. in forward osmosis

Bacteria of different Gram-types have inherently different outer cell structures, influencing cell surface properties and bacterial attachment. Dynamic biofouling experiments were conducted over four days in a bench-scale forward osmosis (FO) system with Gram-negative Pseudomonas aeruginosa or Gram-positive Anoxybacillus sp. Biofouling resulted in ～10% decline in FO permeate water flux and was found to be significant for Anoxybacillus sp. but not for P. aeruginosa. Additionally, a stronger permeate water flux decline for P. aeruginosa in experiments with a superhydrophilic feed spacer demonstrated that mitigation methods require testing with different bacterial Gram-types. It was found that although permeate water flux decline can be affected by bacterial Gram-type the stable performance under enhanced biofouling conditions highlights the potential of FO for wastewater reclamation.     

Crossflow filtration of Saccharomyces cerevisiae using an unsteady jet

This paper deals with the influence of a new flow unsteadiness on the permeate flux in crossflow filtration of microbial suspension during fermentation. A pneumatically controlled valve generates intermittent jets from the main flow, leading to the formation of large vortices moving downstream along the tubular membrane. The unsteadiness does not affect the cell behaviour during fermentation and the resulting permeate flux is found twice higher than for usual filtration process.