Continuous hydrolysis of sucrose by invertase adsorbed in a tubular reactor

Studies were made of invertase adsorption on Amberlite ion exchange resins. Up to 4000 units of adsorbed enzymatic activity (aea) were obtainedper g of IRA 93 resin; for an aea of 1600 units, the maximum ratio of aea over units of soluble enzyme used for adsorption was close to 50%. Nodesorption occurred during extensive washing at 30°C with 0.01M sodiumacetate buffer at pH 5. Progressive desorption of aea from the invertase–IRA 93 complex occurred when buffer molarity and temperature were increased. Desorption differed only slightly when the buffer pH was 3 or 5. Theoptimum pH of aea was 3.2 with IRA 93 resin, and varied between 3.2 and 5.1with other resins, depending on their anionic or cationic nature. Batch hydrolysis of sucrose by IRA 93–adsorbed invertase followed 1st order kinetics with respect to the substrate concentration, as in the case of soluble invertase. Continuous sucrose hydrolysis with IRA 93–adsorbed invertase was performed in a tubular reactor, and the percent conversion was experimentally determined as a function of the flow rate. The reaction was experimentally determined 50% (w/v) sucrose solution, at pH4 and 30°C; at the selected flow rate, the ratio of sucrose hydrolysis remained constant and close to 76%. This shows that invertase was not desorbed from the tubular reactor. Some continuous hydrolyses were performed with an industrial sucrose solution: enzymatic activity seemed to be stable for anextended period for time (1 month) at 30°C and pH 3 or 4.     

Elimination of phenylalanine from amino acid mixtures obtained from yeast hydrolysates and autolyzates]

In order to obtain amino acid mixtures devoid of phenylalanine, adsorption of aromatic amino acids was studied during their chromatography on ion-exchange resin IA-Ip. When 40 g of the amino acid mixture were passed through 400 ml of the swollen ion-exchange resin, phenylalanine and tyrosine were eliminated and 67% of dry substance were yielded. Ion-exchange resin adsorbed phenylalanine and tyrosine could be regenerated. The use IA-Ip is advantageous as compared with that of ion-exchange resin Amberlite IR-45 and activated charcoal.     

Recovery of lactic acid from simultaneous saccharification and fermentation media using anion exchange resins

The physicochemical properties (capacity, kinetics and selectivity) of the ion exchange resins Amberlite IRA900, IRA400, IRA96 and IRA67 were determined to evaluate their comparative suitability for lactic acid recovery. Both the kinetics of lactic acid sorption from aqueous solutions and the equilibrium were assessed using mathematical models, which provided a close interpretation of the experimental results. The best resins (Amberlite IRA96 and IRA67) were employed in further fixed-bed operation using aqueous lactic acid solutions as feed. In this set of experiments, parameters such as capacity, regenerant consumption, percentage of lactic acid recovery and product concentration were measured. Amberlite IRA67, a weak base resin, was selected for lactic acid recovery from SSF (simultaneous saccharification and fermentation) broths. Owing to the presence of nutrients and ions other than lactate, a slightly decreased capacity was determined when using SSF media instead aqueous lactic acid solutions, but quantitative lactic acid recoveries at constant capacities were obtained in four sequential load/regeneration cycles.     

Immobilization of α-amylase and amyloglucosidase onto ion-exchange resin beads and hydrolysis of natural starch at high concentration

α-Amylase was immobilized on Dowex MAC-3 with 88 % yield and amyloglucosidase on Amberlite IRA-400 ion-exchange resin beads with 54 % yield by adsorption process. Immobilized enzymes were characterized to measure the kinetic parameters and optimal operational parameters. Optimum substrate concentration and temperature were higher for immobilized enzymes. The thermal stability of the enzymes enhanced after the immobilization. Immobilized enzymes were used in the hydrolysis of the natural starch at high concentration (35 % w/v). The time required for liquefaction of starch to 10 dextrose equivalent (DE) and saccharification of liquefied starch to 96 DE increased. Immobilized enzymes showed the potential for use in starch hydrolysis as done in industry.     

Wood hydrolyzate treatments for improved fermentation of wood sugars to 2,3-butanediol

Acid-hydrolyzed hardwood contains compounds inhibitory to microorganisms that convert wood sugars to fermentation products such as fuels and chemicals. Several methods of treating acid-hydrolyzed hardwood (hydrolyzate) to reduce the levels of potential microbial inhibitors (acetate, furfural, sulfate, and phenolics) were evaluated. The methods evaluated were precipitation with calcium hydroxide, extraction with organic solvents, treatment with ion-exchange resins, adsorption resins, and activated charcoal. Treatment of the hydrolyzate with an anion exchange resin (Amberlite IRA-400) was the most effective method for removing potential inhibitors. Non-treated hydrolyzate adjusted to pH 6 inhibited growth of a 2,3-butanediol-producing culture of Klebsiella pneumoniae. However, hydrolyzate treated with Amberlite IRA-400 was not inhibitory and resulted in yields of 2,3-butanediol that were greater than 90% of theoretical.     

Development of a micelle-fractional precipitation hybrid process for the pre-purification of paclitaxel from plant cell cultures

A micelle-fractional precipitation hybrid process was developed for the effective pre-purification of the anticancer agent paclitaxel extracted from plant cell cultures. First, it was found that the efficiency of such a developed process could be remarkably enhanced by removing waxy substances originating from plant cells using the adsorbent sylopute. Paclitaxel yield was improved and the fractional precipitation time was shortened by increasing the surface area per working volume (S/V) of the reacting solution through the addition of a cation exchange resin (Amberlite IR120 or Amberlite 200), an anion exchange resin (Amberlite IRA400 or Amberlite IRA96), or glass beads. Most of the paclitaxel (>98%) could be obtained after about 12 h of fractional precipitation using Amberlite 200. Purity increased with increasing fractional precipitation time up to 9 h to about 85%, after which it showed little change. On the other hand, no paclitaxel precipitate was formed using either of the nonionic exchange resins because paclitaxel, which is hydrophobic, was strongly adsorbed on the hydrophobic resin surface. Since high-purity paclitaxel can be obtained in high yield and the precipitation time can be reduced by combining micelle formation with fractional precipitation, this hybrid method is expected to significantly enhance the final purification process.     

茶氨酸提取纯化工艺研究

系统研究了从茶多酚工业废液中提取纯化茶氨酸的工艺。采用絮凝、吸附、阳离子树脂交换、重结晶工艺来分离纯化茶氨酸。结果表明,絮凝能有效的去除茶多酚工业废液中的蛋白质等杂质,杂质的去除率为50％;吸附能进一步去除色素、多酚类物质及大分子有机物;阳离子交换树脂能较专-吸附氨基酸。茶多酚工业废液经絮凝→吸附→阳离子树脂交换工艺可得纯度50%的茶氨酸,得率为1．8％;通过重结晶可得到纯度90％的茶氨酸,得率为0．8%。     

Distribution of saccharides and salts on amphoteric ion-exchange resin

An amphoteric ion-exchange resin hardly shrank in 550 and 300 g/L glucose and sodium chloride solutions, respectively; however, the bed packed with a cation-exchange resin shrank considerably. From the distribution coefficients of some saccharides, the swelling pressure of the amphoteric ion-exchange resin was estimated to be 2.0 MPa at 25 °C. The distribution coefficients of glucose, galactose, fructose, and mannose were independent of their concentration and were about 0.621. On the other hand, the apparent distribution coefficients of NaF, NaCl, NaBr, NaI, LiCl, KCl, and CsCl largely depended on concentration. A model for the distribution of salts on the amphoteric resin was proposed, assuming an interaction between the anion of the salt and the positively charged fixed ions with binding constant B. The B values of the chloride salts were nearly the same (1.69–2.94 L/mol), while the values of the sodium salts were largely different depending on the anion.     

Recovery of native protein from potato root water by expanded bed adsorption with amberlite XAD7HP

Potato root water (PRW) contains ～1.5% protein. In this study, expanded bed adsorption (EBA) chromatography with Amberlite XAD7HP resin adsorbent was used to isolate native protein from crude PRW. The optimal pH and ionic strength for potato protein binding onto Amberlite XAD7HP were 5.0 and 20 mmol/L. The EBA-refined proteins were dried by vacuum freeze drying and spray drying at varying outlet temperatures. Results indicated that low temperature spray drying was the most cost effective method with respect to retaining protease inhibitor activities. The dried protein concentrates appeared bright yellow or dark reddish brown, with a total glycoalkaloid content of ～170 μg/g. The protease inhibitor activity was ～400 mg/g and 11 ～ 12 mg/g for trypsin inhibition and chymotrypsin inhibition, respectively. The results presented here suggest that EBA using Amberlite XAD7HP as the adsorbent is a feasible strategy for the direct adsorption of native protein from crude PRW.     

Amberlite IRA 900 versus calcium alginate in immobilization of a novel,engineered β-fructofuranosidase for short-chain fructooligosaccharide synthesis from sucrose

The immobilization of β-fructofuranosidase for short-chain fructooligosaccharide (scFOS) synthesis holds the potential for a more efficient use of the biocatalyst. However, the choice of carrier and immobilization technique is a key to achieving that efficiency. In this study, calcium alginate (CA), Amberlite IRA 900 (AI900) and Dowex Marathon MSA (DMM) were tested as supports for immobilizing a novel engineered β-fructofuranosidase from Aspergillus japonicus for scFOS synthesis. Several immobilization parameters were estimated to ascertain the effectiveness of the carriers in immobilizing the enzyme. The performance of the immobilized biocatalysts are compared in terms of the yield of scFOS produced and reusability. The selection of carriers and reagents was motivated by the need to ensure safety of application in the production of food-grade products. The CA and AI900 both recorded impressive immobilization yields of 82 and 62%, respectively, while the DMM recorded 47%. Enzyme immobilizations on CA, AI900 and DMM showed activity recoveries of 23, 27, and 17%, respectively. The CA, AI900 immobilized and the free enzymes recorded their highest scFOS yields of 59, 53, and 61%, respectively. The AI900 immobilized enzyme produced a consistent scFOS yield and composition for 12 batch cycles but for the CA immobilized enzyme, only 6 batch cycles gave a consistent scFOS yield. In its first record of application in scFOS production, the AI900 anion exchange resin exhibited potential as an adequate carrier for industrial application with possible savings on cost of immobilization and reduced technical difficulty.     

Lactate from cultures of Lactobacillus casei recovered in a fluidized bed column using ion exchange resin

Summary Lactic acid produced by continuous culture of L.casei in an upflow packed bed reactor, was recovered with Amberlite IRA 400 in a fluidized bed column. Bed expansions of 1.25 and 2.25 were applied. Reutilization did not alter the capability of net recovery of 0.048 ± 0.01 g lactic acid/g resin. When 2200 cm/h of ascensional velocity was used, (bed expansion of 2.25), the resin adsorbed 39.3% of the initial lactic acid and 63.5% was eluted. This resin supported the highest exchange capacity of 0.126 g lactic acid/g resin. Applying high flow rates, the process has potential industrial applications due to the short time employed.     

Partial Purification and Properties of the Inhibitors of Na,K-ATPase and Ouabain-Binding in Bovine Central Nervous System

Abstract: Endogenous inhibitors of Na,K-ATPase and ouabain-binding were partially purified from bovine central nervous system, and some of their properties were studied. They were eluted as low-molecular-weight fractions by gel filtration. They could be adsorbed by both Amberlite IR 120 and Amberlite IRA 400 at acidic and basic pH, respectively, indicating that they could act as both anions and cations at different pH. These inhibitors of ouabain-binding appeared to affect specific binding of ouabin, and Scatchard plot analysis showed that the in hibition was competitive, suggesting that they could bind to the same site as ouabain, presumably to Na,K-ATPase itself. The inhibitory activities were heat stable, but charring inactivated them completely.     

In situ separation of lactic acid from fermentation broth using ion exchange resins

Lactic acid fermentation is an end product inhibited reaction. In situ separation of lactic acid from fermentation broth using ion exchange resins was investigated and compared with conventional fermentation system. Amberlite resin (IRA-400, Cl⁻) was used to separate lactic acid from fermentation broth and pH was controlled online with an automatic pH controller. The effect of process variables on lactic acid production by Lactobacillus casei in whey permeate was studied. The maximum productivity was obtained at pH = 6.1, T = 37 °C and impeller speed = 200 rpm. The maximum concentration of lactic acid at optimum condition was found to be 37.4 g/L after 38 h of fermentation using in situ separation system. The productivity of in situ separation system was five times increased in comparison with conventional system.     

Preparative purification of Escherichia coli heat-stable enterotoxin

Heat-stable enterotoxin (STa) isolated from bovine Escherichia coli strains was purified to homogeneity by growing the bacterial strains in a chemically defined medium, desalting, and concentrating the culture filtrate by batch adsorption chromatography on Amberlite XAD-2 resin, batch adsorption chromatography on reversed-phase silica, and preparative reversed-phase high-performance liquid chromatography. This rapid preparative purification scheme gave high recovery yields of pure STa which exhibited biochemical homology to STa purified by more complicated procedures.     

Zn2+ biosorption by Oscillatoria anguistissima

Oscillatoria anguistissima showed a very high capacity for Zn²⁺ biosorption (641 mg g⁻¹ dry biomass at a residual concentration of 129·2 ppm) from solution and was comparable to the commmercial ion-exchange resin IRA-400C. Zn²⁺ biosorption was rapid, pH dependent and temperature independent phenomenon. Zn²⁺ adsorption followed both Langmuir and Freundlich models. The specific uptake (mg g⁻¹ dry biomass) of metal decreased with increase in biomass concentration. Pretreatment of biomass did not significantly affect the biosorption capacity of O. anguistissima. The biosorption of zinc by O. anguistissima was an ion-exchange phenomenon as a large concentration of magnesium ions were released during zinc adsorption. The zinc bound to the biomass could be effectively stripped using EDTA (10 mM) and the biomass was effectively used for multiple sorption–desorption cycles with in-between charging of the biomass with tap water washings. The native biomass could also efficiently remove zinc from effluents obtained from Indian mining industries.     

Potential use of green algae as a biosorbent for hexavalent chromium removal from aqueous solutions

The hexavalent chromium Cr(VI) poses a threat as a hazardous metal and its removal from aquatic environments through biosorption has gained attention as a viable technology of bioremediation. We evaluated the potential use of three green algae (Cladophora glomerata, Enteromorpha intestinalis and Microspora amoena) dry biomass as a biosorbent to remove Cr(VI) from aqueous solutions. The adsorption capacity of the biomass was determined using batch experiments. The adsorption capacity appeared to depend on the pH. The optimum pH with the acid-treated biomass for Cr(VI) biosorption was found to be 2.0 at a constant temperature, 45?°C. Among the three genera studied, C. glomerata recorded a maximum of 66.6% removal from the batch process using 1.0?g dried algal cells/100?ml aqueous solution containing an initial concentration of 20?mg/L chromium at 45?°C and pH 2.0 for 60?min of contact time. Langmuir and Freundlich isotherm equations fitted to the equilibrium data, Freundlich was the better model. Our study showed that C. glomerata dry biomass is a suitable candidate to remove Cr(VI) from aqueous solutions.     

Immobilization of pectinase fromSclerotium rolfsii

Immobilization of pectinase fromS. rolfsii was studied on different matrices of which Amberlite XAD-7 showed maximum adsorption and expression of the enzyme. The most active preparation was obtained when XAD-7 was activated with 2% glutaraldehyde and 1.7 μkat of enzyme per g resin was used for immobilization at pH 5.5 and 28°C. Optimum pH and temperature of theS. rolfsii pectinase remain unaltered, 3.5 and 55°C, respectively, after immobilization. However, the apparentK _M value of the enzyme decreased from 1.75 g/L for soluble enzyme preparation to 1.4 g/L for immobilized enzyme preparation. Both soluble and immobilized enzyme preparations were most stable at pH 4.0. The immobilized enzyme preparation was more stable than the soluble enzyme.     

Recovery of potassium clavulanate from fermentation broth by ion exchange chromatography and desalting electrodialysis

Ion exchange chromatography (IEC) and desalting electrodialysis (DSED) processes were developed for the recovery and purification of potassium clavulanate (KCA) from fermentation broth. A strong anion exchanger, Amberlite IRA 400 resin, a potassium acetate solution as equilibrium buffer, and a potassium chloride (KCl) solution as elution buffer were used for the recovery of KCA in IEC. In order to determine optimal operating conditions, the effects of various operating parameters such as equilibrium buffer pH and concentration, elution buffer concentration, gradient length, and volumetric flow rate on KCA recovery and by-product removal were investigated using a simulated fermentation broth. In the subsequent step of DSED, employing cation (Neocepta CMS, Tokuyama, Japan) and anion (Neocepta ACS, Tokuyama, Japan) exchange membranes were carried out to remove KCl that existed in a large amount in the ion exchanged solution. The effects of operation voltage and feed composition on the performance of DSED were investigated. Based on the operating conditions determined above, IEC and DSED were applied in sequence to an ultrafiltered fermentation broth. Almost complete removal of KCl was possible with no significant loss of KCA, although the KCA recovery was slightly lower than that with the simulated fermentation broth. Based on this observation, it was concluded that IEC and DESD could be an effective process combination for the recovery of KCA from fermentation broth.     

Removal of potassium chloride by nanofiltration from ion-exchanged solution containing potassium clavulanate

In this study, nanofiltration with NF200 membrane was employed to remove KCl from ion-exchanged solutions containing potassium clavulanate. The pore radius of NF200 membrane was estimated to be around 0.39 nm. The effects of operating pressure on separation performance were investigated in a range of 100–400 psig. The influences of cross-flow velocity (0.14–0.70 cm/s), temperature (4–25 °C), and feed composition were also investigated. In all experiments, clavulanate rejection showed high levels from 0.91 to 0.99, while chloride rejection ranged from 0.06 to 0.54. In a case at an operating pressure of 50 psig and 25 °C, as much as 94% of clavulanate was retained while 94% of chloride was removed, indicating that NF200 membrane was a suitable choice for selectively removing KCl. NF200 membrane also showed a stable performance in the operational stability test with an ion-exchanged solution obtained by treating actual fermentation broth.     

The role of lactic acid adsorption by ion exchange chromatography

Background

The polyacrylic resin Amberlite IRA-67 is a promising adsorbent for lactic acid extraction from aqueous solution, but little systematic research has been devoted to the separation efficiency of lactic acid under different operating conditions.

Methodology/Principal Findings

In this paper, we investigated the effects of temperature, resin dose and lactic acid loading concentration on the adsorption of lactic acid by Amberlite IRA-67 in batch kinetic experiments. The obtained kinetic data followed the pseudo-second order model well and both the equilibrium and ultimate adsorption slightly decreased with the increase of the temperature at 293–323K and 42.5 g/liter lactic acid loading concentration. The adsorption was a chemically heterogeneous process with a mean free energy value of 12.18 kJ/mol. According to the Boyd^_plot, the lactic acid uptake process was primarily found to be an intraparticle diffusion at a lower concentration (<50 g/liter) but a film diffusion at a higher concentration (>70 g/liter). The values of effective diffusion coefficient D_i increased with temperature. By using our Equation (21), the negative values of ΔG° and ΔH° revealed that the adsorption process was spontaneous and exothermic. Moreover, the negative value of ΔS° reflected the decrease of solid-liquid interface randomness at the solid-liquid interface when adsorbing lactic acid on IRA-67.

Conclusions/Significance

With the weakly basic resin IRA-67, in situ product removal of lactic acid can be accomplished especially from an open and thermophilic fermentation system without sterilization.