Syngas fermentation of Clostridium carboxidivoran P7 in a hollow fiber membrane biofilm reactor: Evaluating the mass transfer coefficient and ethanol production performance

Gasification followed by syngas fermentation is a unique hybrid process for converting lignocellulosic biomass into fuels and chemicals. Current syngas fermentation faces several challenges with low gas–liquid mass transfer being one of the major bottlenecks. The aim of this work is to evaluate the performance of hollow fiber membrane biofilm reactor (HFM-BR) as a reactor configuration for syngas fermentation. The volumetric mass transfer coefficient (K_La) of the HFM-BR was determined at abiotic conditions within a wide range of gas velocity/flowrate passing through the hollow fiber lumen and liquid velocity/flowrate passing through the membrane module shell. The K_La values of the HFM-BR were higher than most reactor configurations such as stir tank reactors and bubble columns. A continuous syngas fermentation of Clostridium carboxidivorans P7 was implemented in the HFM-BR system at different operational conditions, including the syngas flow rate, liquid recirculation between the module and reservoir, and the dilution rate. It was found that the syngas fermentation performance such as syngas utilization efficiency, ethanol concentration and productivity, and ratio of ethanol to acetic acid depended not only on the mass transfer efficiency but also the characteristics of biofilm attached on the membrane module (biofouling or abrading of the biofilm). The HFM-BR results in a highest ethanol concentration of 23.93 g/L with an ethanol to acetic acid ratio of 4.79. Collectively, the research shows the HFM-BR is an efficient reactor system for syngas fermentation with high mass transfer.     

A new dynamic model for highly efficient mass transfer in aerated bioreactors and consequences for kLa identification

Gas–liquid mass transfer is often rate‐limiting in laboratory and industrial cultures of aerobic or autotrophic organisms. The volumetric mass transfer coefficient k_La is a crucial characteristic for comparing, optimizing, and upscaling mass transfer efficiency of bioreactors. Reliable dynamic models and resulting methods for parameter identification are needed for quantitative modeling of microbial growth dynamics. We describe a laboratory‐scale stirred tank reactor (STR) with a highly efficient aeration system (k_La ≈ 570 h^?1). The reactor can sustain yeast culture with high cell density and high oxygen uptake rate, leading to a significant drop in gas concentration from inflow to outflow (by 21%). Standard models fail to predict the observed mass transfer dynamics and to identify k_La correctly. In order to capture the concentration gradient in the gas phase, we refine a standard ordinary differential equation (ODE) model and obtain a system of partial integro‐differential equations (PIDE), for which we derive an approximate analytical solution. Specific reactor configurations, in particular a relatively short bubble residence time, allow a quasi steady‐state approximation of the PIDE system by a simpler ODE model which still accounts for the concentration gradient. Moreover, we perform an appropriate scaling of all variables and parameters. In particular, we introduce the dimensionless overall efficiency κ, which is more informative than k_La since it combines the effects of gas inflow, exchange, and solution. Current standard models of mass transfer in laboratory‐scale aerated STRs neglect the gradient in the gas concentration, which arises from highly efficient bubbling systems and high cellular exchange rates. The resulting error in the identification of κ (and hence k_La) increases dramatically with increasing mass transfer efficiency. Notably, the error differs between cell‐free and culture‐based methods of parameter identification, potentially confounding the determination of the “biological enhancement” of mass transfer. Our new model provides an improved theoretical framework that can be readily applied to aerated bioreactors in research and biotechnology. Biotechnol. Bioeng. 2012; 109: 2997–3006. © 2012 Wiley Periodicals, Inc.     

Hydrodynamics and mass transfer coefficient in activated sludge aerated stirred column reactor: experimental analysis and modeling

The aerated stirred reactor (ASR) has been widely used in biochemical and wastewater treatment processes. The information describing how the activated sludge properties and operation conditions affect the hydrodynamics and mass transfer coefficient is missing in the literature. The aim of this study was to investigate the influence of flow regime, superficial gas velocity (U(G)), power consumption unit (P/V(L)), sludge loading, and apparent viscosity (mu(ap)) of activated sludge fluid on the mixing time (t(m)), gas hold-up (epsilon), and volumetric mass transfer coefficient (k(L)a) in an activated sludge aerated stirred column reactor (ASCR). The activated sludge fluid performed a non-Newtonian rheological behavior. The sludge loading significantly affected the fluid hydrodynamics and mass transfer. With an increase in the U(G) and P/V(L), the epsilon and k(L)a increased, and the t(m), decreased. The epsilon, k(L)a, and t(m), were influenced dramatically as the flow regime changed from homogeneous to heterogeneous patterns. The proposed mathematical models predicted the experimental results well under experimental conditions, indicating that the U(G), P/V(L), and mu(ap) had significant impact on the t(m), epsilon, and k(L)a. These models were able to give the t(m), epsilon, and k(L)a values with an error around +/-8%, and always less than +/-10%.     

Measurement of local mass transfer coefficient in biofilms

Local mass transfer rates for an electrochemically formed microsink in an aerobic biofilm was measured by a mobile microelectrode using limiting current technique. Mass transfer coefficients varied both horizontally and vertically in the biofilm. The results implied the existence of an irregular biofilm structure consisting of microbial cell clusters surrounded by tortuous water channels. An unexpected increase of the local mass transfer coefficient just above the biofilm surface suggested the existence, of local flow instability in this region. As expected, the influence of bulk flow velocity on the local mass transfer rate decreased with increasing depth into the biofilm. Mass transfer coefficients fluctuated significantly inside microbial cell clusters, suggesting the existence of internal channels through which liquid could flow. A new conceptual model of biofilm microbial cluster structure is proposed to account for such biofilm microstructure irregularities. (c) 1995 John Wiley & Sons, Inc.     

Prediction of gas-liquid mass transfer coefficient in sparged stirred tank bioreactors

Oxygen mass transfer in sparged stirred tank bioreactors has been studied. The rate of oxygen mass transfer into a culture in a bioreactor is affected by operational conditions and geometrical parameters as well as the physicochemical properties of the medium (nutrients, substances excreted by the micro-organism, and surface active agents that are often added to the medium) and the presence of the micro-organism. Thus, oxygen mass transfer coefficient values in fermentation broths often differ substantially from values estimated for simple aqueous solutions. The influence of liquid phase physicochemical properties on kLa must be divided into the influence on k(L) and a, because they are affected in different ways. The presence of micro-organisms (cells, bacteria, or yeasts) can affect the mass transfer rate, and thus kLa values, due to the consumption of oxygen for both cell growth and metabolite production. In this work, theoretical equations for kLa prediction, developed for sparged and stirred tanks, taking into account the possible oxygen mass transfer enhancement due to the consumption by biochemical reactions, are proposed. The estimation of kLa is carried out taking into account a strong increase of viscosity broth, changes in surface tension and different oxygen uptake rates (OURs), and the biological enhancement factor, E, is also estimated. These different operational conditions and changes in several variables are performed using different systems and cultures (xanthan aqueous solutions, xanthan production cultures by Xanthomonas campestris, sophorolipids production by Candida bombicola, etc.). Experimental and theoretical results are presented and compared, with very good results.     

Comparison of oxygen mass transfer coefficient in simple and extractive fermentation systems

Aeration and agitation are important variables to ensure effective oxygen transfer rate during aerobic bioprocesses; therefore, the knowledge of the volumetric mass transfer coefficient (k_La) is required. In view of selecting the optimum oxygen requirements for extractive fermentation in aqueous two-phase system (ATPS), the k_La values in a typical ATPS medium were compared in this work with those in distilled water and in a simple fermentation medium, in the absence of biomass. Aeration and agitation were selected as the independent variables using a 2² full factorial design. Both variables showed statistically significant effects on k_La, and the highest values of this parameter in both media for simple fermentation (241 s⁻¹) and extractive fermentation with ATPS (70.3 s⁻¹) were observed at the highest levels of aeration (5 vvm) and agitation (1200 rpm). The k_La values were then used to establish mathematical correlations of this response as a function of the process variables. The exponents of the power number (N³D²) and superficial gas velocity (V_s) determined in distilled water (α = 0.39 and β = 0.47, respectively) were in reasonable agreement with the ones reported in the literature for several aqueous systems and close to those determined for a simple fermentation medium (α = 0.38 and β = 0.41). On the other hand, as expected by the increased viscosity in the presence of polyethylene glycol, their values were remarkably higher in a typical medium for extractive fermentation (α = 0.50 and β = 1.0). A reasonable agreement was found between the experimental data of k_La for the three selected systems and the values predicted by the theoretical models, under a wide range of operational conditions.     

Analysis of mass transfer for immobilized cells in an extractive lactic acid fermentation

The use of immobilization in extractive lactic acid fermentation by Lactobacillus delbrueckii is preferred. In this article, the mathematical simulations to examine the influences of substrate and product transport were performed to assess the overall performance. The simulations showed that transport of the substrate in k-carrageenan beads was not a rate limiting factor. However, the model observed significant buildup of inhibitory product in large beads. The model was validated through comparisons with the experimental results. Finally, the model was used to predict the performance of the extractive fermentation under different operating strategies.     

Effect of fermentation broth and biosurfactants on mass transfer during liquid-liquid extraction

Mass transfer rates in liquid-liquid extraction processes can be seriously affected by the presence of surface-active contaminants. This is especially true of applications of a biotechnological origin, where the microorganism used in the process may produce the surface-active contaminants. An investigation into the effects of soluble and insoluble fermentation broth components on mass transfer using chloramphenicol extraction into octanol as the model system was conducted. Soluble components produced during fermentation were found to adsorb to the interface, where they reduced the overall mass transfer coefficient by up to 70%. After fractionation it was found that components in the weight range from 10-30 kDa had the greatest effect on mass transfer. Protein and phospholipid compounds of similar size were found to reduce the overall mass transfer coefficient to a similar extent to the broth components at concentrations around 0.001mg/l. The biomass produced during the fermentation also reduced mass transfer substantially, and it is likely that this was due to physical blockage of the interface.     

Determination of the volumetric mass transfer coefficient (k(L)a) using the dynamic "gas out-gas in" method: Analysis of errors caused by dissolved oxygen probes

There are many dynamic methods for measuring the volumetric mass transfer coefficient. The "gas out-gas in" method can directly determine the volumetric mass transfer coefficient in a bioreactor system and provide estimates of the volumetric microbial oxygen uptake rate and the average oxygen saturation concentration at the gas-liquid interface. The errors on these parameters are large if the dissolved oxygen probe response time is not considered. For reliable measurements, deconvolution of the oxygen probe measurements must be made. (c) 1995 John Wiley & Sons, Inc.     

Prediction of mass transfer coefficients in non-Newtonian fermentation media using first-principles methods

Bubble flows in non-Newtonian fluids were analyzed using first-principles methods with the aim to compute and predict mass transfer coefficients in such fermentation media. The method we used is a Direct Numerical Simulation (DNS) of the reactive multiphase flow with deformable boundaries and interfaces. With this method, we are able for the first time to calculate mass transfer coefficients in non-Newtonian liquids of different rheologies without any experimental data. In the current article, shear-thinning fluids are considered. However, the results provide the basis for further investigations, such as the study of viscoelastic fluids.     

Oxygen transfer effects in serine alkaline protease fermentation by Bacillus licheniformis: use of citric acid as the carbon source

The effects of oxygen transfer on serine alkaline protease (SAP) production by Bacillus licheniformis on a defined medium with C_c = 9.0 kg m⁻³ citric acid as sole carbon source were investigated in 3.5 dm³ batch bioreactor systems. The concentrations of the product (SAP) and by-products, i.e., neutral protease, amylase, amino acids, and organic acids were determined in addition to SAP activities. At Q_o/V = 1 vvm air flow rate, the effect of agitation rate on DO concentration, pH, product, and by-product concentrations and SAP activity were investigated at N = 150, 500, and 750 min⁻¹; these are named as low-(LOT), medium-(MOT), and high oxygen transfer (HOT) conditions. LOT conditions favor biomass concentration; however, substrate consumption was highest at HOT conditions. MOT was optimum for maximum SAP activity which was 441 U cm⁻³ at t = 37 h. The total amino acid concentration was maximum in LOT and minimum in MOT conditions; lysine had the highest concentration under all oxygen transfer conditions. Among organic acids, acetic acid had the highest concentration and its concentration increased with oxygen transfer rate. The oxygen transfer coefficient increases with the agitation rate and the oxygen consumption rate increased almost linearly with the biomass concentration.     

Mathematical analysis of solid-liquid mass transfer in a batch reactor for the enzymatic transformation of testosterone to 4AD

A previous mathematical analysis of mass transfer in a two-phase (solid-liquid) batch reactor for enzymatic transformation of testosterone to 4AD (Pereira et al., 1987) is extended to incorporate the effect of convective mixing. The results of the analysis showed that for a given enzyme loading, the mass transfer resistance in the solid (a function of the bead size) and the intensity of convective mixing (as embodied in the mass transfer coefficient) are two parameters that can be varied such that the overall mass transfer rate from the solid to the liquid phase ensures optimal reactor performance.     

Application of equilibrium and mass transfer models to dynamic removal of Cr(VI) ions by Chitin in packed column reactor

The dynamic removal of hexavalent chromium by chitin flakes was studied in a packed column reactor. The values of column parameters were predicted as a function of flow rate, bed depth, particle size and inlet metal ion concentration. On evaluating the breakthrough curves, sorption isotherms were obtained and modelled according to the Langmuir, the Redlich–Peterson and the Freundlich models. Kinetic and mass transfer aspects of the dynamic removal of Cr(VI) ions by chitin were investigated using several mathematical models. Column studies showed a good correlation between the experimental data and the calculated breakthrough curves obtained by the Adams–Bohart or the Wolborska models and the Clark model. The simulation of the whole breakthrough curve was effective with the Clark model, but the breakthrough was best predicted by Adams–Bohart, or related derived models.     

Using CFD simulations and statistical analysis to correlate oxygen mass transfer coefficient to both geometrical parameters and operating conditions in a stirred-tank bioreactor

Optimization of a bioreactor design can be an especially challenging process. For instance, testing different bioreactor vessel geometries and different impeller and sparger types, locations, and dimensions can lead to an exceedingly large number of configurations and necessary experiments. Computational fluid dynamics (CFD), therefore, has been widely used to model multiphase flow in stirred-tank bioreactors to minimize the number of optimization experiments. In this study, a multiphase CFD model with population balance equations are used to model gas–liquid mixing, as well as gas bubble distribution, in a 50 L single-use bioreactor vessel. The vessel is the larger chamber in an early prototype of a multichamber bioreactor for mammalian cell culture. The model results are validated with oxygen mass transfer coefficient (k_La) measurements within the prototype. The validated model is projected to predict the effect of using ring or pipe spargers of different sizes and the effect of varying the impeller diameter on k_La. The simulations show that ring spargers result in a superior k_La compared to pipe spargers, with an optimum sparger-to-impeller diameter ratio of 0.8. In addition, larger impellers are shown to improve k_La. A correlation of k_La is presented as a function of both the reactor geometry (i.e., sparger-to-impeller diameter ratio and impeller-to-vessel diameter ratio) and operating conditions (i.e., Reynolds number and gas flow rate). The resulting correlation can be used to predict k_La in a bioreactor and to optimize its design, geometry, and operating conditions.     

An on-line sampling system for fermentation monitoring using membrane inlet mass spectrometry (MIMS): application to phenoxyacetic acid monitoring in penicillin fermentation

A sampling system for on-line monitoring of organic compounds of low volatility in complex fermentation media uses membrane inlet mass spectrometry (MIMS). A Syringe pump draws a continuous flow of microfiltered broth from the reactor and circulates it after acidification through a membrane inlet, in which a membrane is the only interface between the sample and the high vacuum of a mass spectrometer. All operations run automatically, i.e., sampling, acidification measurement, and calibration. The on-stream acidification enables MIMS monitoring of carboxylic acids, as they must be undissociated in order to pass the hydrophobic membrane. The performance of the monitoring system was tested by measurements of standard solutions of phenoxyacetic acid (POAA, the sie chain precursor of penicillin-V) as well as on POAA during 200 h penicillin-V fermentation. During the entire fermentation POAA was monitored n low millimolar concentrations with high accuracy and fast response to step changes in POAA concentration. Tandem mass spectrometry (MS/MS) allowed direct identification of peaks in the mass spectrum of the broth that were not accounted for by POAA. These peaks were identified as SO(2) and SCO. (c) 1994 John Wiley & Sons, Inc.     

Determination of the interparticular effective diffusion coefficient for CO(2) and O(2) in solid state fermentation

A simple experimental diffusion controlled fermentor (DCF), coupled with the use of a mathematical model based on mass balance, is proposed to measure the variation of the gas (CO(2) and O(2)) diffusion coefficients in solid state fermentation. The DCF was packed with an ion-exchange resin impregnated with a nutritive medium and inoculated with Aspergillus niger. The growth conditions in the DCF were very similar to those found in equipment operated with convective oxygen supply. The diffusion coefficient was shown to be very dependent on the biomass concentration within the solid state fermentor, and attained values of less than 5% of the molecular diffusion in air when the biomass in the fermentor reached 27 mg dry/g dry support.     

Implementation of proton transfer reaction‐mass spectrometry (PTR‐MS) for advanced bioprocess monitoring

We report on the implementation of proton transfer reaction‐mass spectrometry (PTR‐MS) technology for on‐line monitoring of volatile organic compounds (VOCs) in the off‐gas of bioreactors. The main part of the work was focused on the development of an interface between the bioreactor and an analyzer suitable for continuous sampling of VOCs emanating from the bioprocess. The permanently heated sampling line with an inert surface avoids condensation and interaction of volatiles during transfer to the PTR‐MS. The interface is equipped with a sterile sinter filter unit directly connected to the bioreactor headspace, a condensate trap, and a series of valves allowing for dilution of the headspace gas, in‐process calibration, and multiport operation. To assess the aptitude of the entire system, a case study was conducted comprising three identical cultivations with a recombinant E. coli strain, and the volatiles produced in the course of the experiments were monitored with the PTR‐MS. The high reproducibility of the measurements proved that the established sampling interface allows for reproducible transfer of volatiles from the headspace to the PTR‐MS analyzer. The set of volatile compounds monitored comprises metabolites of different pathways with diverse functions in cell physiology but also volatiles from the process matrix. The trends of individual compounds showed diverse patterns. The recorded signal levels covered a dynamic range of more than five orders of magnitude. It was possible to assign specific volatile compounds to distinctive events in the bioprocess. The presented results clearly show that PTR‐MS was successfully implemented as a powerful bioprocess‐monitoring tool and that access to volatiles emitted by the cells opens promising perspectives in terms of advanced process control. Biotechnol. Bioeng. 2012; 109: 3059–3069. © 2012 Wiley Periodicals, Inc.     

A simple method to estimate the contribution of biological floc and reactor-solution to mass transfer of oxygen in activated sludge processes

In this study, the mass transfer coefficient of biological floc (K(L)a(bf)) was estimated from the mass transfer coefficient of the mixed-liquor (K(L)a(f)) and the reactor-solution (K(L)a(e)). The biological floc resistance (BFR) and reactor-solution resistance (SR) were defined as the reciprocal of K(L)a(bf) and K(L)a(e), respectively, by applying the concept of serial-resistance originally presented in two-film theory (Lewis and Whitman (1924) Ind Eng Chem 16:1215-1220). The specific biological floc resistance (SBFR) was defined as biological floc resistance per unit biomass concentration. The data indicated that an activated sludge process yielding low BFR/MLR and BFR/SR tended to produce higher oxygen transfer efficiency. Surprisingly, the reactor-solution posed the same level of resistance as clean water in all experiments, except in a 5-day SRT, non-nitrifying, completely mixed activated sludge (CMAS) process run. Furthermore, SBFR successfully represented biological floc and showed a positive correlation to sludge volume index (SVI). In addition, SBFR/SR and oxygen transfer efficiency (OTE(f)) followed an exponential relationship for the complete data set. The method of separating the mixed-liquor into biological floc and reactor-solution improved the understanding of oxygen transfer under process conditions, without resorting to intrusive techniques or direct handling of fragile biological floc.     

Mass transfer studies on the reduction of Cr(VI) using calcium alginate immobilized Bacillus sp. in packed bed reactor

This study presents the external mass transfer effects on the reduction of hexavalent chromium (Cr(VI)) using calcium alginate immobilized Bacillus sp. in a re-circulated packed bed batch reactor (RPBR). The effect of flow rate on the reduction Cr(VI) was studied. Theoretically calculated rate constants for various flow rates were analyzed using external film diffusion models and compared with experimental values. The external mass transfer coefficients for the bioconversion of Cr(VI) were also investigated. The external mass transfer effect was correlated with a model of the type J_D = K Re⁻⁽¹⁻ⁿ⁾. The model was tested with various K values and the mass transfer correlation J_D = 5.7 Re^−0.70 was found to predict the experimental data accurately. The proposed model would be useful for the design of industrial reactor and scale up.