Cell recovery by continuous flotation

Summary Cell recovery by means of continuous flotation of the Hansenula polymorpha cultivation medium without additives was investigated as a function of the cultivation conditions as well as of the flotation equipment construction and flotation operational parameters. The cell enrichment and separation is improved at high liquid residence times, high aeration rates, small bubble sizes, increasing height of the aerated column, and diameter of the foam column. Increasing cell age and cultivation with nitrogen limitation reduce the cell separation.Symbols C_P cell mass concentration in medium g·l^–1 - C_R cell mass concentration in residue g·l^–1 - C_S cell mass concentration in foam liquid g·l^–1 - V equilibrium foam volume cm³ - V gas flow rate through the aerated liquid column cm³·s^–1 - V_F feed rate to the flotation column ml/min - ¹ V _S/V foaminess s - mean liquid residence time in the column s     

Hydrodynamics and mass transfer in miniaturized bubble column bioreactors

Miniaturized bubble columns (MBCs) have different hydrodynamics in comparison with the larger ones, but there is a lack of scientific data on MBCs. Hence, in this study, the effect of gas hold-up, flow regimes, bubble size distribution on volumetric oxygen mass transfer coefficient at different pore size spargers and gas flow rates in MBCs in the presence and absence of microorganisms were investigated. It was found that flow regime transition occurred around low gas flow rates of 1.18 and 0.85 cm/s for small (16–40 µm) and large (40–100 µm) pore size spargers, respectively. Gas hold-up and K_La in MBC with small size sparger were higher than those with larger one, with an increasing effect in the presence of microorganisms. A comparison revealed that the wall effect on the flow regime and gas hold-up in MBCs was greater than bench-scale bubble columns. The K_La values significantly increased up to tenfold using small pore size sparger. In the MBC and stirred tank bioreactors, the maximum obtained cell concentrations were OD₆₀₀ of 41.5 and 43.0, respectively. Furthermore, it was shown that in MBCs, higher K_La and lower turbulency could be achieved at the end of bubbly flow regime.

     

Incorporating hydrodynamics into spatiotemporal metabolic models of bubble column gas fermentation

Gas fermentation has emerged as a technologically and economically attractive option for producing renewable fuels and chemicals from carbon monoxide (CO) rich waste streams. LanzaTech has developed a proprietary strain of the gas fermentating acetogen Clostridium autoethanogenum as a microbial platform for synthesizing ethanol, 2,3-butanediol, and other chemicals. Bubble column reactor technology is being developed for the large-scale production, motivating the investigation of multiphase reactor hydrodynamics. In this study, we combined hydrodynamics with a genome-scale reconstruction of C. autoethanogenum metabolism and multiphase convection–dispersion equations to compare the performance of bubble column reactors with and without liquid recycle. For both reactor configurations, hydrodynamics was predicted to diminish bubble column performance with respect to CO conversion, biomass production, and ethanol production when compared with bubble column models in which the gas phase was modeled as ideal plug flow plus axial dispersion. Liquid recycle was predicted to be advantageous by increasing CO conversion, biomass production, and ethanol and 2,3-butanediol production compared with the non-recycle reactor configuration. Parametric studies performed for the liquid recycle configuration with two-phase hydrodynamics showed that increased CO feed flow rates (more gas supply), smaller CO gas bubbles (more gas–liquid mass transfer), and shorter column heights (more gas per volume of liquid per time) favored ethanol production over acetate production. Our computational results demonstrate the power of combining cellular metabolic models and two-phase hydrodynamics for simulating and optimizing gas fermentation reactors.     

Penicillin recovery from aqueous solutions by continuous foam flotation

Summary Penicillin G recovery is investigated in a continuous flotation column in the presence of different collectors which form a complex with penicillin. The performance of the penicillin recovery was investigated as a function of the mole ratio () of collector-to-penicillin and the aliphatic chain length of the collector. At =1 and low penicillin concentrations (e.g., 20 mg·1^-1), high foam liquid concentrations (680 mg·l^-1), low residue concentrations (12 mg·l^-1) and high penicillin separation (56) can be attained. At =4 the separation increases to 150, and 95% of the penicillin can be recovered.Symbols C_p penicillin concentration in feed (mg·l^-1) - C_R penicillin concentration in outlet liquid (mg·l^-1) - C_S penicillin concentration in foam liquid (mg·l^-1) - C_S/C_P penicillin enrichment (-) - C_S/C_R penicillin separation (-) - % Pen in S penicillin yield in foam liquid (%) - VV}_S foam liquid volume flow (ml·min^-1) - VV}_P feed (ml·min^-1) - VV_{N 2} nitrogen flow rate (ml·s^-1) - temperature     

A mechanism for internal reflux in foam fractionation

Multiple equilibrium stages can be engendered in foam fractionation, a process used for the enrichment of streams of proteins, by returning some of the foamate stream to the top of the column as external reflux liquor. However, it was recognised, 40 years ago that reflux could be autogenously created through the coalescence of bubbles in fractionation columns. By invoking the hydrodynamic theory of rising foam, we suggest a mechanism for the creation of internal reflux in foam fractionation. This method can give internal reflux rate as a function of bubble size. However, since the bubble size profile in a rising foam cannot be estimated, we cannot yet estimate how internal reflux varies with position in the column.     

The influence of polymeric membrane gas spargers on hydrodynamics and mass transfer in bubble column bioreactors

Gas sparging performances of a flat sheet and tubular polymeric membranes were investigated in 3.1 m bubble column bioreactor operated in a semi batch mode. Air–water and air–CMC (Carboxymethyl cellulose) solutions of 0.5, 0.75 and 1.0 % w/w were used as interacting gas–liquid mediums. CMC solutions were employed in the study to simulate rheological properties of bioreactor broth. Gas holdup, bubble size distribution, interfacial area and gas–liquid mass transfer were studied in the homogeneous bubbly flow hydrodynamic regime with superficial gas velocity (U _G) range of 0.0004–0.0025 m/s. The study indicated that the tubular membrane sparger produced the highest gas holdup and densely populated fine bubbles with narrow size distribution. An increase in liquid viscosity promoted a shift in bubble size distribution to large stable bubbles and smaller specific interfacial area. The tubular membrane sparger achieved greater interfacial area and an enhanced overall mass transfer coefficient (K _La) by a factor of 1.2–1.9 compared to the flat sheet membrane.     

Protein recovery by continuous flotation

Summary Bovine serum albumin (BSA) was recovered from aqueous solutions by foam flotation. The protein concentrations in foam liquid C _S, in feed C _Pand in residue C _Rwere determined. The protein enrichment C _S/C_Pand the separation C _S/C_Ras well as the protein fraction in the foam liquid % BSA and foam liquid volume flow were determined as functions of the medium properties, operational conditions, and equipment parameters as well as concentrations of solid particles. At low protein concentrations in feed (e.g., C _P=40 mg · l^-1), and at 40° C, high performance was attained (C _X=2,000 mg · l^-1, C _R=4.4 mg · l^-1, C _S/C_P=50, C _S/C_R=450, 90% BSA. Continuous foam flotation is an efficient procedure for the recovery of low concentrations of proteins from liquid cultures.Abbreviations BSA bovine serum albumine - C _P protein concentration in feed (mg · l^-1) - C _R protein concentration in residue (mg · l^-1) - C _S protein concentration in foam liquid (mg · l^-1) - C _S/C_R protein separation (-) - C _S/C_P protein enrichment (-) - V _P feed rate (ml · min^-1) - V _R residue flow rate (ml · min^-1) - V _S foam liquid volume flow (ml · min^-1) - N number of theoretical stages in an ideal cascade (-) - temperature (° C) - mean residence time (min)     

Bubble column bioreactors

Summary The photographic and electrical conductivity methods to measure the structure of two phase flow, especially bubble size, bubble frequency, local gas hold-up and, for the latter, the bubble velocity are described.Symbols specific interfacial area - a gas/liquid interfacial area - B constant in Eq. (4) - d diameter of the bubbles - d mean diameter of the bubbles - d_S Sauter diameter - E_G relative gas hold up - I current - k_L mass transfer coefficient across the gas/liquid interface - k_L local k_L - LT^–1 - LT^–1 - 1 longitudinal distance between the start and stop sensors - 1_B pierced length of the bubble - t time - t₁ length of the square-wave signal at the start sensor - t₂ length of the square-wave signal at the stop sensor - t₁₂ time delay between start and stop signals - V volume of the bubbling layer - V_L volume of the bubble free layer - V_B bubble volume - v_B bubble velocity     

Change in body size of juvenile marbled sole <Emphasis Type="Italic">Pseudopleuronectes yokohamae</Emphasis> after preservation in ethanol

We evaluated the extent of shrinkage in body size of juvenile marbled sole Pseudopleuronectes yokohamae after preservation in 70% ethanol for 6, 12, 24, or 48 h, or 4 weeks. Standard length (SL, range of the analyzed specimens: 22.82–53.01 mm) decreased by 5.6% after 12 h and body weight (BW 0.174–2.964 g) decreased 27.8% after 24 h preservation in 70% ethanol. There was no further decrease in SL or BW after 12 or 24 h of preservation, respectively. The original body size could be estimated based on the size after preservation in 70% ethanol for 4 weeks using the following equations: SL _original = 1.05 SL _preserved + 0.37 and BW _original = 1.36 BW _preserved + 0.04. The condition factor calculated using the body size of preserved individuals was 16.0 and 15.8% lower than that calculated from the original body size and the body size back-calculated using the linear regression equations described above, respectively. Thus, our data suggest that the shrinkage of body size due to preservation in ethanol can cause errors in estimation of the condition factor. Our results can be used to improve the accuracy of estimating size-related biological parameters based on juvenile marbled sole that are preserved in ethanol.     

Improvement of carbon dioxide biofixation in a photobioreactor using Anabaena sp. PCC 7120

The biological photosynthetic process is useful and environmentally benign compared with other carbon dioxide (CO₂) mitigation processes. In the present study, Anabaena sp. PCC 7120 was utilized for carbon dioxide mitigation. A customized airlift photobioreactor was found to provide higher light utilization efficiency and a higher rate of CO₂ biofixation compared with that of a bubble column. The maximum biomass concentrations were 0.71 and 1.13 g L^?1 in the bubble column and airlift photobioreactor, respectively, using BG11₀ medium under aerated conditions. A lower mixing time in the airlift photobioreactor compared with that of the bubble column resulted in improved mass transfer. The CO₂ biofixation rate of Anabaena sp. PCC 7120 was determined using different phosphate concentrations at a light intensity of 120 μE m^?2 s^?1 and 5% (v/v) CO₂-enriched air in the airlift photobioreactor. However, it was observed that the specific growth rate was independent at higher light intensity. In addition, it was observed that increased light intensity, phosphate and CO₂ concentrations could enhance the CO₂ biofixation efficiency to a greater extent.     

Correlation of liquid dispersion and oxygen transfer in bubble column

Summary In steady state, attained by continuous aeration after oxygen saturation of water in a bubble column, vertical composition distribution of liquid and gas phases has been determined. It has been assumed that, as a result of absorption at the bottom of the column, desorption in the upper section and vertical dispersion of dissolved oxygen flux, a closed oxygen circulation is created. Determination of the axial dispersion coefficient from hydrodynamic and oxygen transfer data verifies the mathematical model proposed. The results allow conclusions to be drawn about supersaturation and desorption and other phenomena expected in biological systems.Abbreviations C[-] Dimensionless oxygen concentration Unit=0.21 bar oxygen partial pressure or dissolved oxygen level in equilibrium with latter - E[m²/s] Axial dispersion coefficient - F[m²] Horizontal cross-section area - k _L a[s^-1] Overall oxygen transfer coefficient - u; u ₂[m/s; cm/s] Superficial velocity: related to state of bubbles leaving the sparger - x; x _atm[-] Signal registered in the experiment; signal recorded in O₂ saturated water, or water vapor saturated air stream, at temperature identical to the experiment under atmospheric pressure - y[m] Water column height - [s^-1] Dimensionless oxygen flux Indices a asorption - d desorption - g gas - l liquid - k dispersion - m measured value/in the case of hydrodynamically measured E/ Dedicated to Professor Dr. H. J. Rehm on the occasion of his 60th birthday     

High efficiency production of astaxanthin by autotrophic cultivation of Haematococcus pluvialis in a bubble column photobioreactor

Haematococcus pluvialis was cultivated under photoautotrophic conditions in a bubble column with fed-batch addition of nutrients, especially nitrate, and a cell number above 5 × 10⁶ cells mL⁻¹ was attained after 300 h.The reduction of nutrient concentrations accompanied by dilution of the fermentation broth and an increase in the light intensity enhanced accumulation of astaxanthin. The final astaxanthin concentration of 390 mg L⁻¹ was several times higher than ever reported. This combination of fed-batch addition of nutrients and dilution of broth for nutrient deficiency is a promising method for attainment of high cell and astaxanthin concentrations in a bubble column photobioreactor.     

Synthesis of closo-Dodecaboryl Lipids and their Liposomal Formation for Boron Neutron Capture Therapy

High accumulation and selective delivery of boron into tumor tissues are the most important requirements to achieve efficient neutron capture therapy of cancers. We focused on liposomal boron delivery system to achieve a large amount of boron delivery to tumor. We succeeded in the synthesis of the double-tailed boron cluster lipids 4a–c and 5a–c, which has a B₁₂H₁₁S-moiety as a hydrophilic function, by S-alkylation of B₁₂H₁₁SH with bromoacetyl and chloroacetocarbamate derivatives of diacylglycerols. Size distribution of liposomes prepared from the boron cluster lipid 4b, dimyristoylphosphatidylcholine, polyethyleneglycol-conjugated distearoylphosphatidylethanolamine, and cholesterol was determined as 100 nm in diameter by an electrophoretic light scattering spectrophotometer. Calcein-encapsulation experiments revealed that these boronated liposomes are stable at 37 °C in fetal bovine serum solution for 24 h.     

Analysis of carvedilol enantiomers in human plasma using chiral stationary phase column and liquid chromatography with tandem mass spectrometry

Carvedilol is an antihypertensive drug available as a racemic mixture. (?)‐(S)‐carvedilol is responsible for the nonselective β‐blocker activity but both enantiomers present similar activity on α₁‐adrenergic receptor. To our knowledge, this is the first study of carvedilol enantiomers in human plasma using a chiral stationary phase column and liquid chromatography with tandem mass spectrometry. The method involves plasma extraction with diisopropyl ether using metoprolol as internal standard and direct separation of the carvedilol enantiomers on a Chirobiotic T® (Teicoplanin) column. Protonated ions [M + H]⁺ and their respective ion products were monitored at transitions of 407 > 100 for the carvedilol enantiomers and 268 > 116 for the internal standard. The quantification limit was 0.2 ng ml^?1 for both enantiomers in plasma. The method was applied to study enantioselectivity in the pharmacokinetics of carvedilol administered as a single dose of 25 mg to a hypertensive patient. The results showed a higher plasma concentration of (+)‐(R)‐carvedilol (AUC^0–∞ 205.52 vs. 82.61 (ng h) ml^?1), with an enantiomer ratio of 2.48. Chirality, 2012. © 2012 Wiley Periodicals, Inc.     

Bioleaching of chalcopyrite concentrate using Leptospirillum ferriphilum, Acidithiobacillus ferrooxidans and Acidithiobacillus thiooxidans in a continuous bubble column reactor

To estimate the bioleaching performance of chalcopyrite for various hydraulic residence times (HRTs), laboratory-scale bioleaching of chalcopyrite concentrate was carried out in a continuous bubble column reactor with three different HRTs of 120, 80 and 40 h, respectively. An extraction rate and ratio of 0.578 g Cu l⁻¹ h⁻¹ and 39.7%, respectively, were achieved for an HRT of 80 h at a solids concentration of 10% (w/v). Lower bioleaching performances than this were obtained for a longer HRT of 120 h and a shorter HRT of 40 h. In addition, there was obvious competition between Leptospirillum ferriphilum and Acidithiobacillus ferrooxidans to oxidize ferrous iron, causing large compositional differences between the microbial communitys obtained for the different HRTs. Leptospirillum ferriphilum and Acidithiobacillus thiooxidans were found to be the dominant microbes for the longer HRT (120 h). Acidithiobacillus ferrooxidans became the dominant species when the HRT was decreased. The proportion of Acidithiobacillus thiooxidans was comparatively constant in the microbial community throughout the three process stages.     

Chromatographic characterisation of 11 phytocannabinoids: Quantitative and fit‐to‐purpose performance as a function of extra‐column variance

Introduction

Cannabis sativa L. (cannabis) is utilised as a therapeutic and recreational drug. With the legalisation of cannabis in many countries and the anticipated regulation of potency that will accompany legalisation, analytical testing facilities will require a broadly applicable, quantitative, high throughput method to meet increased demand. Current analytical methods for the biologically active components of cannabis (phytocannabinoids) suffer from low throughput and/or an incomplete complement of relevant phytocannabinoids.

Objective

To develop a rapid, quantitative and broadly applicable liquid chromatography–tandem mass spectrometry analytical method for 11 phytocannabinoids in cannabis with acidic and neutral character.

Methodology

Bulk diffusion coefficients were calculated using the Taylor–Aris open tubular method, with four reference compounds used to validate the experimental set‐up. Three columns were quantitatively evaluated using van Deemter plots and fit‐to‐purpose performance metrics. Low (1.2 μL²) and standard (3.6 μL²) extra‐column variance ultra‐high pressure liquid chromatography (UPLC) configurations were contrasted. Method performance was demonstrated with methanolic cannabis flower extracts.

Results

Bulk diffusion coefficients and van Deemter plots for 11 phytocannabinoids are reported. The developed chromatographic method includes the challenging Δ⁸/Δ⁹‐tetrahydrocannabinol isobars and, at 6.5 min, is faster than existing methods targeting similar panels of biologically active phytocannabinoids.

Conclusions

The bulk diffusion coefficients and van Deemter curves informed the development of a rapid quantitative method and will facilitate potential expansion to include additional compounds, including synthetic cannabinoids. The developed method can be implemented with low or standard extra‐column variance UPLC configurations.     

Methods for measuring the bubble size in bubble column bioreactors II

Summary Electrooptical and light scattering methods for measuring the structure of two phase flow, especially bubble size, are described and compared with photographic and electrical conductivity methods.Symbols L length - T time - M mass - d bubble diameter - mean bubble diameter - d_K inside diameter of the sensor capillary - 1 longitudinal distance between start and stop sensors - 1_B pierced length of bubble - t time - t₁ length of square-wave signal at start sensor - t₂ length of square-wave signal at stop sensor - 1₁₂ time delay between start and stop signals - V_B bubble volume - V_B bubble velocity - w_SG superficial gas velocity     

Semicontinuous penicillin production by two Penicillium chrysogenum strains immobilized in calcium alginate beads

Summary The growth rates of immobilized Penicillium chrysogenum strains are important in their application to semicontinuous penicillin production. Immobilized P. chrysogenum strains produced about 10–15% less biomass but about 1–2 times more penicillin than free suspended mycelia.In a chemically defined medium an industrial P. chrysogenum strain, S₁, produced about 10–12 times more penicillin than strain ATCC 12690. In a complex medium the immobilized P. chrysogenum S₁ produced about 12% penicillin more than in shaken cultures. In bubble column fermentations, penicillin production was 163% higher in the complex medium than in the chemically defined medium.     

Gas holdup and mass transfer characteristics of carboxymethyl cellulose solutions in a bubble column with a radial gas sparger

The gas phase holdup and mass transfer characteristics of carboxymethyl cellulose (CMC) solutions in a bubble column having a radial gas sparger have been determined and a new flow regime map has been proposed. The gas holdup increases with gas velocity in the bubbly flow regime, decreases in the churn-turbulent flow regime, and increases again in the slug flow regime. The volumetric mass transfer coefficient (k _La) significantly decreases with increasing liquid viscosity. The gas holdup and k _La values in the present bubble column of CMC solutions are found to be much higher than those in bubble columns or external-loop airlift columns with a plate-type sparger. The obtained gas phase holdup ( _g) and k _La data have been correlated with pertinent dimensionless groups in both the bubbly and the churn-turbulent flow regimes.List of Symbols a m^–1 specific gas-liquid interfacial area per total volume - A _d m² cross-sectional area of downcomer - A _r m² cross-sectional area of riser - d _b m individual bubble diameter - d _vs m Sauter mean bubble diameter - D _c m column diameter - D _L m²/s oxygen diffusivity in the liquid - Fr Froude number, U _g/(g D_c)^1/2 - g m/s² gravitational acceleration - G _a Galileo number, gD _c ^{3 2}/² _app - H _a m aerated liquid height - H _c m unaerated liquid height - K Pa · sⁿ fluid consistency index - k _L a s^–1 volumetric mass transfer coefficient - n flow behavior index - N _i number of bubbles having diameter d _bi - Sc Schmidt number, _app/( D _L) - Sh Sherwood number, k _L a D _c ² /D_L - U _sg m/s superficial gas velocity - U _gr m/s superficial riser gas velocity - V _a m³ aerated liquid volume - V _c m³ unaerated liquid volume - N/m surface tension of the liquid phase - _g gas holdup - _app Pa · s effective viscosity of non-Newtonian liquid - kg/m³ liquid density - ý s^–1 shear rate - Pa shear stress     

Foam fractionation of globular proteins

Foam fractionation of bovine serum albumin (BSA) was studied as a model system for potato wastewater. The effects of feed concentration, superficial gas velocity, feed flow rate, bubble size, pH, and ionic strength on the enrichment and recovery of BSA were investigated in a single-stage continuous foam fractionation column. Enrichments ranged from 1.5 to 6.0 and recoveries from 5 to 85%. The feed concentrations were varied from 0.01 to 0.2 wt %, and enrichments were found to increase with lower feed concentrations. Enrichments also increased with lower superficial gas velocities and larger bubble sizes. At sufficiently low feed flow rates, enrichment was found to increase with an increase in the flow rate, eventually becoming insensitive to the feed flow rate at higher values. The pH was varied from 3.5 to 7.0 and ionic strength from 0.001M to 0.2M. The effects of pH and ionic strength were found to be coupled with bubble size. A minimum bubble size was found at pH 4.8, the isoelectric point of BSA, resulting in a minimum in the enrichment. Bubble size, and thus enrichment, was found to increase as the ionic strength decreased from 0.2M to 0.01M. Previous models(1,2) for the hydrodynamics of foam column were extended for a singlestage continuous foam fractionation column for the prediction of enrichment and recovery. The model assumed adsorption equilibrium, infinite surface viscosity, and bubbles of the same size. Though coalescence was formally accounted for in the model by considering bubble size as a function of foam height, calculations for the experimental runs were performed only for the case of no coalescence. Quantitative predictions of enrichment and recovery could not be made with a single representative bubble size because of the broad inlet bubble size distribution as well as broadening of the distribution as a result of coalescence. The experimental enrichments were higher and recoveries were lower than the model predictions, the discrepancy being more pronounced at lower feed concentrations because of increased coalescence. The higher enrichments are due to the predominant effect of internal reflux as a result of coalescence whereas the lower recoveries are a result of detrimental effects of broadening bubble size distributions.