Improved oxygen delivery in a continuous-roller-bottle reactor

Variations to the original aeration system in a continuous roller bottle reactor of novel design have been tested and compared for optimal oxygen (O) delivery. Reactor operating parameters that affect O transfer are rotation rate, liquid-volume level, fresh-feed rate, and supplementary-aeration rate. Design modifications to enhance gas-liquid O transfer include the addition of wall baffles and center baffles. The number and location of each of these baffles are compared for their effect on k_La values in the reaction chamber. The liquid feed into the system has been modified to improve the axial liquid mixing and O transfer.     

Growth of aerobic bacteria on alkali- solubilized lignite

Applied Biochemistry and Biotechnology -     

Biogasification of community-derived biomass and solid wastes in a pilot-scale solcon® reactor

Applied Biochemistry and Biotechnology - The Institute of Gas Technology has developed a novel, solids-concentrating (SOLCON®) bioreactor to convert a variety of individual or mixed feedstocks...     

Development of a novel,two-step process for treating municipal biosolids for beneficial reuse

Modern municipal sewage waste treatment plants use conventional mechanical and biological processes to reclaim wastewaters. This process has an overall effect of converting a water pollution problem into a solid waste disposal problem (sludges or biosolids). An estimated 10 million tons of biosolids, which require final disposal, are produced annually in the United States. Although numerous disposal options for biosolids are available, including land application, landfilling, and incineration, disposal costs have risen, partly because of increased federal and local environmental restrictions(1). A novel, thermomechanical biosolids pre-treatment process, which allows for a variety of potential value-added uses, was developed. This two-step process first employs thermal explosive decompression to inactivate or kill the microbial cells and viruses. This primary step also results in the rupture of a small amount of the microbial biomass and increases the intrinsic fluidity of the biosolids. The second step uses shear to effect a near-complete rupturing of the microbial biomass, and shears the nondigested organics, which increases the overall surface area. Pretreated biosolids may be subjected to a secondary anaerobic digestion process to produce additional fuel gas, and to provide for a high-quality, easily dewatered compost product. This novel biosolids pretreatment process was recently allowed a United States patent.     

Production of ethanol from starch by co-immobilizedZymomonas mobilis-glucoamylase in a fluidized-bed reactor

The production of ethanol from starch was studied in a fluidized-bed reactor (FBR) using co-immobilizedZymomonas mobilis and glucoamylase. The FBR was a glass column of 2.54 cm in diameter and 120 cm in length. TheZ. mobilis and glucoamylase were co-immobilized within small uniform beads (1.2-2.5 mm diameter) of κ-carrageenan. The substrate for ethanol production was a soluble starch. Light steep water was used as the complex nutrient source. The experiments were performed at 35κC and pH range of 4.0-5.5. The substrate concentrations ranged from 40 to 185 g/L, and the feed rates from 10 to 37 mL/min. Under relaxed sterility conditions, the FBR was successfully operated for a period of 22 d, during which no contamination or structural failure of the biocatalyst beads was observed. Volumetric productivity as high as 38 g ethanol/(Lh), which was 74% of the maximum expected value, was obtained. Typical ethanol volumetric productivity was in the range of 15-20 g/(Lh). The average yield was 0.49 g ethanol/g substrate consumed, which was 90% of the theoretical yield. Very low levels of glucose were observed in the reactor, indicating that starch hydrolysis was the rate-limiting step.     

Bioconversion of vanillin to vanillyl alcohol in a two-phase reactor

One of the next challenges in the use of biocatalysts (enzyme or microbial cells) is the upgrading of biological reactions of oxidoreduction. The oxidoreductases need cofactors that must be regenerated. Practical experience shows that this is most readily achieved by using living cells of microorganisms. Living cells ofSaccharomyces cerevisiae are able to bioconvert vanillin to vanillyl alcohol (1). By working with a two-phase reactor (dodecanol—feeding medium) it has been possible to use higher vanillin concentrations without inhibiting the bioconversion (2).     

Microbial conversion of 4-oxoisophorone by thermomonospora curvata using an air-bubbling hollow fiber reactor

Microbial conversion of 4-oxoisophorone (OIP) by thermophilic bacteriumThermomonospora curvata was attempted in a continuous process. The correlation between cell growth and microbial conversion was first examined in a batch culture. The results indicated that this microbial conversion was strongly dependent upon cell growth. In a continuous microbial conversion of OIP using a continuous stirred tank reactor, the cell density in the reactor seemed to be the limiting factor in the OIP conversion. Therefore, we developed an air-bubbling hollow fiber reactor to achieve a high density culture. By using this bioreactor, more than 3.3 times higher productivity was achieved. In addition, during the process, only a slight cell contamination to the product was observed. Therefore, this bioreactor is suitable for the continuous microbial conversion, considering further downstream processes and high productivity.     

Simultaneous saccharification and fermentation of lignocellulose

Simultaneous saccharification and fermentation (SSF) processes for producing ethanol from lignocellulose are capable of improved hydrolysis rates, yields, and product concentrations compared to separate hydrolysis and fermentation (SHF) systems, because the continuous removal of the sugars by the yeasts reduces the end-product inhibition of the enzyme complex. Recent experiments using Genencor 150L cellulase and mixed yeast cultures have produced yields and concentrations of ethanol from cellulose of 80% and 4.5%, respectively. The mixed culture was employed because B.clausenii has the ability to ferment cellobiose (further reducing end-product inhibition), while the brewing yeastS. cerevisiae provides a robust ability to ferment the monomeric sugars. These experimental results are combined with a process model to evaluate the economics of the process and to investigate the effect of alternative processes, conditions, and organisms.     

Technical and economic evaluation of different reactors for methanotrophic cultures for propylene oxide production

A two-stage process for the manufacture of propylene oxide is described. The preliminary economics based on use of methanol as a regeneration factor has resulted in a production cost of $12.10/lb of propylene oxide based on propylene oxide production rate of 40 mg/g-cell/h in conventional reactor. Increasing the propylene oxide production from 40 to 500 mg/g-cell/h resulted in a cost reduction from $12.10 to 5.8/lb of propylene oxide. The granular-activated, carbon-fluidized bed reactor (GAC-FBR) absorbs the propylene oxide and when saturated is eluted with ethyl acetate, and the bed is regenerated by steam to drive off the residual solvents. The estimated manufacturing costs are approx 59% lower (from $12.10/lb in conventional reactors to $5.00/lb for GAC-FBRs) for products that are highly inhibitory such as epoxides. In the GAC-FBR reactor, enhancing the propylene oxide production rate from 120 to 1500 mg/g-cell/h has resulted in the cost reduction to $2.00/lb. Enhancing the production capacity from 1 million lb to 10 million lb/yr has further reduced the cost of production to $1.00/lb.     

Anaerobic bioconversion of municipal solid wastes using a novel high-solids reactor design

Novel, laboratory-scale, high-solids reactors operated under mesophilic conditions were used to study the anaerobic fermentation of processed municipal solid waste (MSW) to methane. Product gas rate data were determined for organic loading rates ranging from 2.99–18.46 g of volatile solids (VS) per liter (L) per day (d). The data represent the anaerobic fermentation at high-solids levels within the reactor of 21–32%, while feeding a refuse-derived fuel (RDF)/MSW feedstock supplemented with a vitamin/mineral/nutrient solution. The average biogas yield was 0.59 L biogas/g VS added to the reactor system/d. The average methane composition of the biogas produced was 57.2%. The data indicate a linear relationship of increasing total biogas production with increasing organic loading rate to the process. The maximum organic loading rate obtainable with high-solids anaerobic digestion is in the range of 18–20 g VS/L·d to obtain 80% or greater bioconversion for the RDF/MSW feedstock. This loading rate is approximately four to six times greater than that which can be obtained with comparable low-solids anaerobic bioreactor technology.     

Purification and partial characterization of two lectin isoforms fromCratylia mollis mart. (camaratu bean)

Two additional electrophoretically distinct molecular forms, isoforms (iso) 2 and 3, with lectin properties were isolated fromCratylia mollis Mart, seeds (FABACEAE), by extraction with 0.15M NaCl and ammonium sulfate fractionation, followed by chromatography on Sephadex G-75 and Bio-Gel P-200 (iso 2), as well as CM-Cellulose and Sephadex G-75 (iso 3). Both isoforms were human group nonspecific and showed distinct specificity. Polyacrylamide gel electrophoresis resolved iso 2 and 3 in polypeptides of apparent mol wts 60 and 31 kDa, respectively; a distinct isoelectric focusing pattern was obtained for iso 2 and 3, under denaturing and reducing conditions.     

Development of a novel laboratory scale high solids reactor for anaerobic digestion of processed municipal solid wastes for the production of methane

Economic evaluations of the capital costs for anaerobic digestion systems for gas production show that the reactor is a significant cost component. The successful application of high solids digestion of processed MSW (e.g., greater than 10% solids within the digester) would allow a decrease in reactor volume with maintenance of relatively high gas production rates. However, high solids slurries do not mix well in conventional stirred tank reactors. A horizontal shaft, hydraulically driven reactor was designed and fabricated to test the anaerobic digestion of high solids concentrations. Digester performance was evaluated as a function of experimental parameters such as nutrient requirements, feeding rates, pH control, and agitator design/ rotation speed; horsepower of mixing was also evaluated for the reactor. Several startup protocols were examined to obtain a biologically stable anaerobic fermentation at high solids levels.     

Urease purification from the seeds ofCajanus Cajan and its application in a biosensor construction

Urease has been purified from the seeds of Cajanus Cajan. The purification process involves three solvent extraction steps followed by DEAE-cellulose column chromatography. The specific activity of the purified enzyme is found to be 1920 U/mg with the recovery of 8%. The application of the purified enzyme in a biosensor construction is discussed.     

Mode of action and synergism of cellulases fromPenicillium funiculosum

A 1,4-β-d-glucan cellobiohydrolase (EC 3.2.1.91) and l,4-β-d-glucan glucanohydrolase (EC 3.2.1.4) were purified from the culture filtrates ofPenicillium funiculosum by using preparative isoelectric focusing. Both the enzymes were homogeneous on polyacrylamide gel with and without sodium dodecyl sulphate. The mol wt of the cellobiohydrolase and endoglucanase were 14,400 and 25,000 respectively. The purified enzymes were free of β-glucosidase activity. Acting in isolation, the cellobiohydrolase had little capacity for solubilizing Avicel or Walseth cellulose, but showed increased rates of hydrolysis when combined with endoglucanase. Cellobiose inhibition (50%) was observed in the initial rate of the hydrolysis of Walseth cellulose. It was also observed that cellobiohydrolase initiates the attack on crystalline cellulose. † NCL communication no. 3898.     

Synthesis of Cyclodextrin Glucosyl Transferase byBacillus cereus for the production of cyclodextrins

A potent indigenous bacillus isolate identified asBacillus cereus (RJ-30) was found to produce Cyclodextrin Glucosyl Transferase (CGTase) extracellularly. Process optimization of various fermentation parameters has been established for optimal growth of bacillus and the maximum enzyme synthesis. The organism had the highest specific growth rate (0.7μ) with a generation time of 1 h in glucose containing medium at the conditions of pH 7.0, 37°C at 300 rpm, 1.5 vvm of agitation, and aeration. At these conditions, it exhibited the maximum activity of 54 U/mL at the synthesis rate of 2.7 U/L/h. CGTase was produced from the early exponential growth and peaked during the midsporulating stage of about 16 h thereafter maintained at the same level of 50 U/mL. Saccharides containing media were better inducers than starch, and the influence of carbohydrate substrates has shown that enzyme synthesis is promoted by xylose (65 U/mL) and, more remarkably, by the supplementation of wheat bran extract in glucose medium (106 U/mL). This organism produced CGTase stably in a chemostat culturing over a period of 400 h with a maximum productivity of 5.4 kU/L/h (threefold higher than obtained in batch culturing [1.75 kU/L/h]). Comparatively, CGTase was produced by immobilized cells in a continuous fluidized bed reactor for over approx 360 h, at a relatively high dilution rate of 0.88 h⁻¹ resulting in the productivity of 23.0 kU/L/h.     

Principles for efficient utilization of light for mass production of photoautotrophic microorganisms

Outdoor production of microalgae could be set on a sound industrial basis if solar energy were utilized at a much higher efficiency than presently obtained. Many types of photobioreactors have been developed in the past in an attempt to answer this challenge, but their photosynthetic efficiency has been rather similar to the basically inefficient open raceway commonly used today. Efficient utilization of the oversaturating solar energy flux mandates that reactors should have a narrow lightpath to facilitate ultra-high cell densities, be maximally exposed to sunshine, and have an efficient mixing system to create strong turbulent streaming to affect dark–light cycles of the highest possible frequency.     

Use of novel immobilized β-galactosidase reactor to hydrolyze the lactose constituent of skim milk

A novel chemical reactor, consisting of β-galactosidase fromAspergillus oryzae immobilized on a ribbed membrane made from polyvinyl-chloride and silica, was used to hydrolyze the lactose constituent of skim milk. Multiresponse nonlinear regression methods were employed to determine the kinetic parameters of rate expressions based on a proposed enzymatic mechanism that includes the formation of oligosaccharides. HPLC methods were employed to monitor the concentrations of all species present in the effluent stream. For the experimental conditions used in this research, a rate expression that includes the inhibition effect of α-galactose is sufficient to model the reaction network.     

Growth inhibition in animal cell culture

Eight independent cell lines accumulated ammonia in culture to concentrations between 1.3 and 2.9 mM. The growth inhibition of such concentrations of ammonium chloride when added to culture medium was variable. The cell lines tested could be divided into 3 groups depending on their growth response to 2 mM added NH4Cl. In the first group (293, HDF, Vero, and PQXB1/2) little (less than 14%) or no growth inhibition occurred. In the second group (McCoy and MDCK) a reduction in final cell yield of 50-60% was observed. The third group (HeLa and BHK) was most sensitive to the effects of NH4Cl with growth inhibition (greater than 75%) compared to controls. The growth inhibitory effect of added lactate up to 20 mM was negligible (less than 10%) for 3 cell lines, although one cell line (PQXB1/2) showed greater sensitivity. The interactive effects of ammonia and lactate were determined in a matrix experiment. At lactate (greater than 12 mM) and ammonia (1-4 mM), the growth inhibitory effects of the two components were synergistic. However, at low concentrations of lactate (less than 12 mM) the toxic effect of ammonia was reduced. A proposed mechanism for the sparing effect of lactate on ammonia toxicity is discussed. This may have importance in developing strategies for the optimal growth of ammonia-sensitive cell lines.     

Preparation of Tyr-C-peptide from genetically altered human insulin precursor

C-peptide radioimmunoassay (C-peptide RIA) is widely used in determination of pancreatic B-cell secretion activity.¹²⁵I labeled TyrC-peptide is indispensable in C-peptide RIA kit. Herein we discuss a way of obtaining recombinant Tyr-C-peptide. Arg32Tyr human proinsulin mutant (R32Y-proinsulin) gene was constructed by site-directed mutagenesis and overexpressed inEscherichia coli. Purified R32Y-proinsulin was converted to insulin and Tyr-C-peptide by trypsin and carboxypeptidase B codigestion. Tyr-C-peptide was isolated through reverse-phase HPLC (RP-HPLC) and identified by C-peptide RIA and amino acid analysis.