Surface energetics to assess biomass attachment onto hydrophobic interaction adsorbents in expanded beds

Cell-to-support interaction and cell-to-cell aggregation phenomena have been studied in a model system composed of intact yeast cells and Phenyl-Streamline adsorbents. Biomass components and beaded adsorbents were characterized by contact angle determinations with three diagnostic liquids and zeta potential measurements. Subsequently, free energy of interaction vs. distance profiles between interacting surfaces was calculated in the aqueous media provided by operating mobile phases. The effect of pH and ammonium sulphate concentration within the normal operating ranges was evaluated. Calculation indicated that moderate interaction between cell particles and adsorbent beads can develop in the presence of salt. Cell-to-cell aggregation was suspected to occur at high salt concentration and neutral pH. Predictions based on the application of the XDLVO approach were confirmed by independent experimental methods like biomass deposition experiments and laser diffraction spectroscopy. Understanding biomass attachment onto hydrophobic supports can help in alleviating process limitations normally encountered during expanded bed adsorption of bioproducts.     

The influence of complex biological feedstock on the fluidization and bed stability in expanded bed adsorption

The stability of expanded bed adsorption systems (EBA) was studied in biomass containing culture broth by residence time distribution (RTD) experiments, using pulse inputs of fluorescent molecules as tracers. Different commercial adsorbents (Streamline DEAE, SP, Phenyl, Chelating, and AC) were tested at various biomass concentrations (2.5–12 %, wet weight) of whole (Saccharomyces cerevisiae) yeast, yeast cell homogenate, and Escherichia coli homogenate. Analyzing the RTD according to the PDE model (PDE: axially dispersed plug‐flow exchanging mass with stagnant zones) allowed the calculation of three parameters: the number of transfer units for mass exchange between mobile and stagnant fraction (N), the Peclet number for overall axial dispersion (P), and the mobile fraction of the liquid in axially dispersed plug flow (φ). When fluidization was performed in particle‐free buffer the normalized response signal (after perfect input pulse) was symmetric (N:0; P: 50–100; φ: 1), thus, demonstrating the formation of a homogeneous fluidized (expanded) bed. Upon application of suspended biomass the RTD was skewed, depending on the adsorbent used and the type and level of biomass present in the sample. This situation leads to three different characteristic pictures: the well‐fluidized system (N: ≥ 7–10; P: ≥ 40; φ: 0.80–0.90), the system exhibiting bottom channeling (N: < 1–2; P: ≥ 40; φ: 0.5–0.7) and, the system where extensive agglomeration develops (N: 4–7; P: 20–40; φ: < 0.5). These results demonstrate that changes in the hydrodynamics of EBA already take place in the presence of moderate concentrations of biomass. Furthermore, those changes can be quantitatively described mainly in terms of the fraction of stagnant zones in the system, which are formed due to the interaction of biomass and adsorbent. The technique described here can be used to evaluate a certain combination of adsorbent and biomass with regard to its suitability for expanded bed adsorption from whole broth. © 1999 John Wiley & Sons, Inc. Biotechol Bioeng 64: 484–496, 1999.     

Surface energetics to assess biomass attachment onto immobilized metal-ion chromatography adsorbents in expanded beds

Cell-to-support interaction and cell-to-cell agglomeration phenomena have been studied in a model system composed of intact yeast cells and Chelating-Streamline™ adsorbents. Biomass components and beaded adsorbents were mainly characterized by contact angle determinations with three diagnostic liquids. Complementarily, zeta potential measurements were performed. These experimental values were employed to calculate free energy of interaction versus distance profiles in aqueous media. The effect of immobilized metal-ion type and buffer pH on the interaction energy was evaluated. Calculations indicated that moderate interaction between cell particles and adsorbent beads can develop due to the presence of Cu²⁺ ions onto the solid phase. The strength of interaction increased with buffer pH, within the range 6.0 to 8.3 e.g. secondary energy pockets increased from |15| to |60| kT. Cell-to-cell secondary energy minimum was ≥ |14| kT showing low-to-moderate tendencies to aggregate, particularly at pH ≥ 8. Extended DLVO predictions were generally confirmed by biomass deposition experiments. However, an exception was found when working with immobilized Cu²⁺ at pH 8 since yeast cells were able to sequestrate such immobilized ions. Therefore, lower-than-expected values for the depositions coefficient (α) were observed. Understanding biomass attachment onto Chelating supports can help in better design and operate expanded bed adsorption of bioproducts.     

The influence of consumer diversity and indirect facilitation on trophic level biomass and stability

The relationship between species diversity and the stability and production of trophic levels continues to receive intense scientific interest. Though facilitation is commonly cited as an essential underlying mechanism, few studies have provided evidence of the impact that indirect facilitation may have on diversity–ecosystem functioning relationships. In this laboratory study, we examined the effect of zooplankton species diversity on trophic structure (total algal and zooplankton biomass) and temporal stability of total zooplankton biomass. We utilized four species of pond zooplankton grown in either monoculture or in polyculture. When comparing responses in polycultures with responses averaged across monocultures, a positive effect of diversity on total zooplankton biomass was observed. This occurred as a result of positive facilitative effects among competing zooplankton. Daphnia pulex , a biomass dominant in monoculture, was negatively affected by the presence of interspecific competitors. In contrast, Diaphanosoma brachyurum , a species that performed poorly in monoculture, was strongly and positively affected by the presence of interspecific competitors, driving positive diversity effects on total zooplankton biomass. Positive temporal covariances among zooplankton were detected in several polyculture replicates, increasing temporal variability of total zooplankton biomass. However, this destabilizing effect was weak relative to effects of high biomass yields in polyculture which caused temporal biomass variability (as measured by the coefficient of variation) to be lower in polyculture relative to monocultures. Zooplankton diversity effects on total algal biomass were not detected. However, increased zooplankton diversity significantly altered the size structure of algae, increasing the relative abundance of large, grazer-resistant algae.     

Protein adsorption and hydrodynamic stability of a dense, pellicular adsorbent in high-biomass expanded bed chromatography

A dense, pellicular UpFront adsorbent (ϱ=1.5 g/cm³, UpFront Chromatography, Cophenhagen, Denmark) was characterized in terms of hydrodynamic properties and protein adsorption performance in expanded bed chromatography. Cibacron Blue 3GA was immobilised into the adsorbent and protein adsorption of bovine serum albumin (BSA) was selected to test the setup. The Bodenstein number and axial dispersion coefficient estimated for this dense pellicular adsorbent was 54 and 1.63×10⁻⁵ m²/s, respectively, indicating a stable expanded bed. It could be shown that the BSA protein was captured by the adsorbent in the presence of 30% (w/v) of whole-yeast cells with an estimated dynamic binding capacity (C/C ₀=0.01) of approximately 6.5 mg/mL adsorbent.     

Microbial biomass and productivity in seagrass beds

This research focused on whether bacteria living in aerobic environments where Fe is often a limiting nutrient could access Fe associated with the clay mineral kaolinite. Kaolinite is one of the most abundant clays at the Earth's surface, and it often contains trace quantities of Fe as surface precipitates, accessory minerals, and structural substitutions. We hypothesized that aerobic bacteria may enhance kaolinite dissolution as a means of obtaining associated Fe. To test this hypothesis, we conducted microbial growth experiments in the presence of an aerobic Pseudomonas mendocina     

Stability of expanded beds during the application of crude feedstock

Expanded bed adsorption is an integrated technology that allows the introduction of a particle containing feedstock without the risk of blocking the bed. Provided a perfectly classified fluidized bed (termed expanded bed) is formed in the crude feed, a sorption performance comparable to packed beds is found. During the application of biomass containing samples to stable expanded beds an increase in bed expansion due to the higher density and viscosity of the feed is encountered. In this article it is investigated whether the expanded bed condition is also fulfilled during the transition in bed expansion from lower to higher density (i.e., from an equilibration buffer to a biomass containing feedstock). Residence time distribution analyses were performed by using model systems and a yeast suspension during this transition phase. It is shown that in systems in which the biomass does not interact with the fluidized stationary phase, the perfectly classified fluidization is maintained also during this transition phase regardless of the type of feedstock. Additional bed expansion takes place in an "ordered" manner without compromising bed stability. In case of biomass/adsorbent interactions, a deterioration in bed stability is found directly when the crude feed is loaded.     

Microbial biomass and productivity in seagrass beds

Different methods for measuring the rates of processes mediated by bacteria in sediments and the rates of bacterial cell production have been compared. In addition, net production of the seagrass Zostera capricorni and bacterial production have been compared and some interrelationships with the nitrogen cycle discussed. Seagrass productivity was estimated by measuring the plastochrone interval using a leaf stapling technique. The average productivity over four seasons was 1.28 +/- 0.28 g C m-2 day-1 (mean +/- standard deviation, n = 4). Bacterial productivity was measured five times throughout a year using the rate of tritiated thymidine incorporated into DNA. Average values were 33 +/- 12 mg C m-2 day-1 for sediment and 23 +/- 4 for water column (n = 5). Spatial variability between samples was greater than seasonal variation for both seagrass productivity and bacterial productivity. On one occasion, bacterial productivity was measured using the rate of 32P incorporated into phospholipid. The values were comparable to those obtained with tritiated thymidine. The rate of sulfate reduction was 10 mmol SO4(-2) m-2 day-1. The rate of methanogenesis was low, being 5.6 mg CH4 produced m-2 day-1. A comparison of C flux measured using rates of sulfate reduction and DNA synthesis indicated that anaerobic processes were predominant in these sediments. An analysis of microbial biomass and community structure, using techniques of phospholipid analysis, showed that bacteria were predominant members of the microbial biomass and that of these, strictly anaerobic bacteria were the main components. Ammonia concentration in interstitial water varied from 23 to 71 micromoles. Estimates of the amount of ammonia required by seagrass showed that the ammonia would turn over about once per day. Rapid recycling of nitrogen by bacteria and bacterial grazers is probably important.     

Influence of the extent of disruption of Bakers' yeast on protein adsorption in expanded beds

Expanded bed adsorption chromatography is used to capture the protein product of interest from a crude biological suspension directly, thereby eliminating the need for the removal of the cell debris. While this technique may replace three or four unit operations in a typical downstream process for biological product recovery, the adsorption process is influenced by the interaction between the microbial cells or cell debris and the adsorbent as well as the presence of contaminating solutes. The influence of the extent and nature of disruption of Bakers' yeast on the adsorption of the total soluble protein and alpha-glucosidase was investigated in this study. Two different techniques were used for cell disruption: high pressure homogenisation and hydrodynamic cavitation. Two different adsorbents were chosen: anionic Streamline DEAE and cationic Streamline SP. The settled bed height and the superficial velocity were constant across all experiments. The feedstock was characterised in terms of viscosity, pH, conductivity, particle size distribution of the cell debris and the extent of protein and alpha-glucosidase released. The performance of the adsorption process was found to be influenced by the electrostatic interactions of cell debris with the anionic adsorbent Streamline DEAE and the intraparticle diffusional resistance inside the pores of the adsorbent matrix. The increase in the intensity of disruption resulted in an increase in the dynamic binding capacity (10% feed) of both the total soluble protein and the alpha-glucosidase. However, the increase in the DBC of protein and alpha-glucosidase were not proportional. The amount of protein that could be adsorbed per ml of adsorbent from the samples subjected to a lower intensity of disruption was found to exceed that obtained at a higher disruption intensity on increasing the volume of feed suggesting multilayer adsorption. In this case, selective adsorption of the model protein alpha-glucosidase was reduced, illustrating the compromise of maximising protein recovery through non-specific binding. The study illustrates the need for an interrogation of the intensity of disruption needed and a rigorous understanding of the influence of cell debris and adsorbent-protein interaction, in optimising the selective recovery of intracellular products by EBA.     

The influence of vessel height and top-section size on the hydrodynamic characteristics of airlift fermentors

Fermentations of the yeast Saccharomyces cerevisiae were carried out in a 90 to 250-L working volume concentric tube airlift fermentor. Measurements of liquid circulation velocity, gas hold-up, and liquid mixing were made under varying conditions of gas flowrate, vessel height, and top-section size. Both liquid circulation velocity and mixing time increased with vessel height. Liquid velocity varied approximately in proportion to the square root of column height, supporting a theoretically based relationship. The effect of vessel height on gas hold-up was negligible. The height of the top-section had a significant effect on liquid mixing. Mixing time decreased with increasing size of the top-section up to a critical height. As the top-section was expanded beyond this height, little improvement in mixing was seen. This indicated the presence of a two-zone flow pattern in the top-section. Liquid velocity and gas hold-up were essentially independent of top-section height. (c) 1994 John Wiley & Sons, Inc.     

The influence of particle size distribution and operating conditions on the adsorption performance in fluidized beds

The influence of matrix properties and operating conditions on the performance in fluidized-bed adsorption has been studied using Streamline diethyl-aminoethyl (DEAE), an ion exchange matrix based on quartz-weighted agarose, and bovine serum albumin (BSA) as a model protein. Three different particle size fractions (120-160 mum, 120-300 mum, and 250-300 mum) were investigated. Dispersion in the liquid phase was reduced when particles with a wide size distribution were fluidized compared to narrow particle size distributions. When the mean particle diameter was reduced, the breakthrough capacities during frontal adsorption were enlarged due to a shorter diffusion path length within the matrix. At small particle diameters the effect of film mass transfer became more relevant to the adsorption performance in comparison to larger particles. Therefore matrices designed for fluidized-bed adsorption should have small particle diameter and increased mean particle density to ensure small diffusion path length in the particle and a high interstitial velocity to improve film mass transfer. Studies on the influence of sedimented matrix height on axial mixing showed an increased Bodenstein number with increasing bed length. Higher breakthrough capacities were also found for longer adsorbent beds due to reduced dispersion and improved fluid and particle side mass transfer. With increasing bed height the influence of flow rate on breakthrough capacity was reduced. For a settled bed height of 50 cm breakthrough capacities of 80% of the equilibrium capacity for flow rates varying from 3 to 9 cm/min could be achieved. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 54-64, 1997.     

The influence of periphyton biomass and density on grazing in Physella virgata

We studied how differences in periphyton colonization interval and snail density affected grazing rates in Physella virgata, and whether snails controlled periphyton biomass. Both egestion rates and incorporation rates of ¹⁴C labeled periphyton were estimated in laboratory experiments. Periphyton biomass increased with field colonization interval in all experiments, but did not consistently influence estimates of grazing rate. However, increased periphyton abundance in one of the experiments could still explain higher grazer rates in that year, although larger snail body size is a confounding explanation. Increased snail density also resulted in decreased grazing rates, as observed in earlier studies with this snail species, as well as in studies with other snail grazers. Our results suggest grazing rates and resulting impacts may change seasonally with variation in either periphyton biomass, grazer life-history stage or population density.     

The influence of biomass temperature on biostabilization-biodrying of municipal solid waste

A laboratory study was carried out to obtain data on the influence of biomass temperature on biostabilization-biodrying of municipal solid waste (initial moisture content of 410 g kg wet weight (w.w.)(-1)). Three trials were carried out at three different biomass temperatures, obtained by airflow rate control (A = 70 degrees C, B = 60 degrees C and C = 45 degrees C). Biodegradation and biodrying were inversely correlated: fast biodrying produced low biological stability and vice versa. The product obtained from process A was characterized by the highest degradation coefficient (166 g kg TS0(-1); TS0(-1) = initial total solid content) and lowest water loss (409 g kg W0(-1); W0 = initial water content). Due to the high reduction of easily degradable volatile solid content and preservation of water, process A produced the highest biological stability (dynamic respiration index, DRI = 141 mg O2 kg VS(-1); VS = volatile solids) but the lowest energy content (EC = 10,351 kJ kg w.w.(-1)). Conversely, process C which showed the highest water elimination (667 g kg W0(-1)), and lowest degradation rate (18 g kg TS0(-1)) was optimal for refuse-derived fuel (RDF) production having the highest energy content (EC = 14,056 kJ kg w.w.(-1)). Nevertheless, the low biological stability reached, due to preservation of degradable volatile solids, at the end of the process (DRI = 1055 mg O2 kg VS(-1)), indicated that the RDF should be used immediately, without storage. Trial B showed substantial agreement between low moisture content (losses of 665 g kg W0(-1)), high energy content (EC = 13,558 kJ kg w.w.(-1)) and good biological stability (DRI = 166 mg O2 kg VS(-1)), so that, in this case, the product could be used immediately for RDF or stored with minimum pollutant impact (odors, leaches and biogas production).     

The influence of homogenisation conditions on biomass-adsorbent interactions during ion-exchange expanded bed adsorption

Expanded bed adsorption (EBA) is an integrative step in downstream processing allowing the direct capture of target proteins from cell-containing feedstocks. Extensive co-adsorption of biomass, however, may hamper the application of this technique. The latter is especially observed at anion exchange processes as cells or cell debris are negatively charged under common anion exchange conditions. The restrictions observed under these conditions are, however, directly related to processing steps prior to fluidised bed application. In this study, it could be shown that the effective surface charge of cell debris obtained during homogenisation is closely related to the debris size and thus to the homogenisation method and conditions. The amount and thus effect of cells binding to the adsorbent could be significantly decreased when optimising the homogenisation step not only towards optimal product release but towards a reduction of debris size and charge. The lower size and charge of the debris results not only in a reduced retention probability but also, in a lower collision probability between debris and adsorbent. The applicability was shown in an example where the homogenisation conditions of E. coli were optimised towards EBA applications. In a previous report (Reichert et al., 2001) studying the suitability of EBA for the capture of formate dehydrogenate from E. coli homogenate the pseudo affinity resin Streamline Red was identified as the only suitable adsorbent. The new approach, however, led to a system where anion exchange as capture step became possible, however, to the cost of binding capacity.     

The influence of prey biomass on activity and consumption rates of brook trout

The objectives of this work were (1) to assess the influence of zooplankton biomass on activity and consumption rates of young-of-the-year brook trout ( Salvelinus fontinalis ) and (2) to validate an in situ enclosure approach to quantify energy allocation patterns in fish. These objectives were attained by directly estimating fish growth, consumption and activity rates on 10 occasions characterized by different levels of zooplankton biomass (0.005 to 0.100 mg dry weight 1⁻¹). One enclosure was used to estimate growth and activity rates and five additional enclosures were used to estimate consumption rates. Among-experiment variations of activity rates (sum for five trout = 2.4 to 33.5 calories day⁻¹) were proportionally more important than variations of consumption rates (sum for five trout = 59.5 to 112.7 calories day⁻¹). The results support the existence of a significant positive relationship between fish activity rates and zooplankton biomass. No significant relationship was found between consumption rates and prey biomass. Final size of fish inside the enclosure was within 7.6% of the value estimated using experimentally derived activity and consumption rates. This situation, together with the stability of among-enclosure activity and feeding schedules, suggested that the experimental design was appropriate to estimate fish energy allocation patterns. Combination of our observations with those of a previously published work indicated that small variations of fish size or zooplankton biomass can cause a two-fold variation of fish activity costs.     

Factors affecting biomass attachment during startup and operation of anaerobic fluidized beds

Three anaerobic fluidized bed reactors at 37 degrees C were utilized to observe the effects of startup and operational procedures on biomass attachment. Using a meat-based synthetic waste and stepped-loading regime, the influences of synthetic polymer addition and maintenance of anaerobiosis during startup were investigated. Subsequently, increasing bed expansions were applied to assess shear effects. Synthetic polymer addition enhanced biomass retention but did not improve process performance. Maintenance of a reduced environment ameliorated fluctuating process parameters during start up and aided biomass retention and substrate removal. A bed expansion of 5% was detrimental to biomass attachment and COD removal but system stability was maintained at expansions between 10% and 30%. Startup was achieved in 56 days. Anaerobiosis appeared to enhance the initial evolution of a stable, well-adapted microbial population, whereas polymer addition interfered with this. Moderate bed expansions had negligible effects on attachment and performance.     

Comparative hydrodynamic stability of brachiopod shells on current-scoured arenaceous substrates

Selected brachiopod specimens were placed on a well-sorted medium grained sand (0.5 mm) in a recirculating flume and subjected to a constant unidirectional current of 26–27 cm/sec. The specimens were placed in each of six feasible life-orientations. In three orientations, the valves were reclining with either the lateral, anterior or posterior profile upcurrent. In three orientations, the commissural plane was perpendicular to the substrate with either the pedicle or brachial valve upcurrent, or a lateral margin upcurrent. Destabilization indices were calculated for each specimen in each orientation based on the elapsed time until the specimen reoriented. Nonstrophic biconvex brachiopods are most stable with the valves reclining on the substrate. Among orientations with the valves erect, the pedicle valve upcurrent and the brachial valve upcurrent are the most and least stable orientations, respectively. Shell stability increased among nonstrophic specimens that were more equi-biconvex. Biconvex strophic brachiopods are also more stable if the valves were equi-biconvex, cither compressed (biplanate) or inflated, provided that the specimen was propped up on its hingeline with a lateral margin upcurrent. Catacline interareas afforded greater stability than procline, apsacline or orthocline interareas when the valves were perpendicular to the substrate. Accentuated plication reduced shell stability. Alate biconvex forms were often more stable if the valves were vertical to the substrate rather than reclining. Alate biconvex geometries are more stable than nonstrophic and strophic biconvex forms if the commissural plane is perpendicular to the substrate and parallel to the current. Broad catacline interareas increased stability in all orientations. Plano-convex and dorsi-biconvex alate forms are usually less stable than equi-biconvex or ventri-biconvex alate geometries. Concavo-convex geometries are stable in all orientations except with the valves nearly vertical to the substrate and parallel to the current. Spines greatly retard sediment-scour and maintain concavo-convex specimens in orientations with the valves elevated above the substrates. D Brachiopoda, shell shape, hydrodynamic stability.     

Evaluation of aboveground and belowground biomass recovery in physically disturbed seagrass beds

Several studies addressed aboveground biomass recovery in tropical and subtropical seagrass systems following physical disturbance. However, there are few studies documenting belowground biomass recovery despite the important functional and ecological role of roots and rhizomes for seagrass ecosystems. In this study, we compared the recovery of biomass (g dry weight m(-2)) as well as the biomass recovery rates in ten severely disturbed multi-species seagrass meadows, after the sediments were excavated and the seagrasses removed. Three sites were located in the tropics (Puerto Rico) and seven in the subtropics (Florida Keys), and all were originally dominated by Thalassia testudinum. Total aboveground biomass reached reference values at four out of ten sites studied, two in the Florida Keys and two in Puerto Rico. Total belowground biomass was lower at the disturbed locations compared to the references at all sites, apart from two sites in the Florida Keys where the compensatory effect of opportunistic species (Syringodium filiforme and Halodule wrightii) was observed. The results revealed large variation among sites in aboveground and belowground biomass for all species, with higher aboveground recovery than belowground for T. testudinum. Recovery rates for T. testudinum were highly variable across sites, but a general trend of faster aboveground than belowground recovery was observed. Equal rates between aboveground and belowground biomass were found for opportunistic species at several sites in the Florida Keys. These results indicate the importance of belowground biomass when assessing seagrass recovery and suggest that the appropriate metric to assess seagrass recovery should address belowground biomass as well as aboveground biomass in order to evaluate the full recovery of ecological services and functions performed by seagrasses. We point out regional differences in species composition and species shifts following severe disturbance events and discuss ecological implications of gap dynamics in multi-species seagrass meadows.