Biodiesel production using anionic ion-exchange resin as heterogeneous catalyst

The transesterification reactions of triolein with ethanol using various ion-exchange resin catalysts were conducted to produce ethyl oleate as a biodiesel. The anion-exchange resins exhibited much higher catalytic activities than the cation-exchange resin. The anion-exchange resin with a lower cross-linking density and a smaller particle size gave a high reaction rate as well as a high conversion. By combining the three-step regeneration method, the resin could be repeatedly used for the batch transesterification without any loss in the catalytic activity. A continuous transesterification reaction was carried out using an expanded bed reactor packed with the most active resin. The reactor system permitted the continuous production of ethyl oleate with a high conversion.     

Extractive lactic acid fermentation using ion-exchange resin

Lactic acid fermentation is an end-product-inhibited reaction. The restriction imposed by lactic acid on its fermentation can be avoided by extractive fermentation techniques. Studies were performed by attaching an ion-exchange resin packed column with a 2-L fermentor for separation of lactic acid. The fermentation, in a conventional batch mode, resulted in a lactic acid yield of 0.828 g . g(-1) and a lactic acid productivity of 0.313 g . L(-1) . h(-1). However, these could be further enhanced to 0.929 g . g(-1) and 1.665 g . L(-1) . h(-1) by extractive fermentation techniques. The effect of temperature on extractive fermentation was remarkable and has been included in this work.     

采用一种新介质纯化神经生长因子

采用一种新闻子交换介质Ｅｘｐｒｅｓｓ－ＩｏｎＣ（简称ＩｏｎＣ）使鼠神经生长因子（ｍＮＧＦ）能得到规模纯化。以鼠颌下腺为原料,匀浆后,经Ｅｘｐｒｅｓｓ－ＩｏｎＣ纯化得单一组分的ｍＮＧＦ,用ＳＤＳ－ＰＡＧＥ测定其相对分子质量约为１３．７×１０＾３,比活约２．３×１０＾５Ｕ／ｍｇ。结果表明ＩｏｎＣ可替代ＣＭ－５２,且更适于工因子的规模化生产。     

Glycosylation reaction via a mild and efficient one-pot reaction using doped natural phosphate with iodine as catalyst

Several alpha/beta-D-ribonucleosides were synthesized in good yields under mild conditions by N-glycosylations of acetyl 2,3,5-tri-O-benzoyl-beta-D-ribofuranose with silylated nucleobases in acetonitrile using NP doped with iodine as catalyst.     

Gram-scale purification of phosphorothioate oligonucleotides using ion-exchange displacement chromatography.

The purification of oligonucleotides by ion-exchange displacement chromatography is demonstrated on the gram-scale. Using a 50 mmD x 100 mmL (203 ml) column operated in the displacement mode, 1.2 g of a 24mer phosphorothioate oligonucleotide was purified. Product yield for this separation was 70% (780 mg) at a purity of 96.4% and the mass balance recovery of all oligonucleotide was 97.5%. The displacement purification of four additional phosphorothioate oligonucleotides ranging in length from 18 to 25 bases is also demonstrated on the semi-preparative (10-50 mg) scale. All of these oligonucleotides were purified using similar displacement conditions and typical results were 60% yield at 96% purity. The displacement portion of these separations required <15 min and total cycle time including equilibration, feed loading and regeneration can be performed in under 30 min. These results seem to indicate that displacement chromatography may be amenable to generalizations in separation protocol that would greatly reduce the effort required to obtain an optimized purification scheme for moderately long oligonucleotides.     

Rapid purification of a recombinant protein using tandem radial flow ion-exchange column chromatography.

Tandem radial flow anion- and cation-exchange columns were used to partially purify and concentrate a dilute recombinant protein that had been refolded in vitro after production as insoluble inclusion bodies in E. coli. The refolded sample was first passed through a Q-Sepharose Fast Flow column in order to remove the majority of E. coli contaminating proteins and endotoxins, then purified on an S-Sepharose Fast Flow column connected to the outlet of the Q-Sepharose column. This tandem arrangement enabled the rapid processing of multiple preparations of refolded material during production method development.     

Further purification of adenosine kinase from rat heart using affinity and ion-exchange chromatography

Adenosine kinase (EC 2.7.1.20) in a cytoplasmic fraction of rat heart was subjected to 5′-AMP-Sepharose 4B chromatography. The enzyme showed affinity for the column in contrast to adenosine deaminase, and was eluted with adenosine plus MgATP. Fractions containing adenosine kinase were put on a column of DEAE-Sephacel and eluted with a gradient. The enzyme was purified up to 3000-fold (yield 10%). The specific activity exceeded 8000 units per gram of protein and sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed only one band. We conclude that the method presented is a simple, quick, and elegant way of purifying myocardial adenosine kinase to virtual homogeneity.     

Large-scale purification of fungal glucoamylases using anion-exchange resin chromatography

Anion-exchange chromatography on polystyrene resin is shown to be more effective than DEAE-cellulose for purification of glucoamylase from crude enzyme extracts of Aspergillus awamori or from commercial preparations. The glucoamylase from A. awamori culture medium was purified to electrophoretic homogeneity with yields approaching 80%.     

The use of polyaniline nanofibre as a support for lipase mediated reaction

Highly stable and recoverable polianiline nanofibres are developed for enzyme immobilisation and recovery. Candida rugosa lipase (LP) was immobilised onto a polyaniline nanofibre with cross-linking for enzyme aggregation. The optimal LP loading was 5 mg LP/1 mg polyaniline. The stability of the immobilised LP was measured and shown to be high under vigorous shaking at room temperature. This polyaniline nanofibre LP was easily separable with low-speed centrifugation and repeatedly usable. LP immobilised on polyaniline nanofibre demonstrated high stereoselectivity in the kinetic resolution of racemic (R,S)-ibuprofen and improved the long-term stability as compared to that by the free enzyme, allowing the supported enzyme to be repeatedly used for a series of chiral resolution reactions. The conversion from racemic ibuprofen to a chirally selective compound, a prophilic ester of ibuprofen, was approximately 30% with free LP and approximately 10% with immobilised LP. The enantiomeric excess using immobilised LP after 96 h reaction was 0.884.     

Acetylation of sugarcane bagasse using NBS as a catalyst under mild reaction conditions for the production of oil sorption-active materials

Sugarcane bagasse was esterified with acetic anhydride using N-bromosuccinimide as a catalyst under mild conditions in a solvent free system. The extent of acetylation was measured by weight percent gain, which varied from 2.1% to 24.7% by changing the reaction temperature (25-130 degrees C) and duration (0.5-6.0 h). N-Bromosuccinimide was found to be a novel and highly effective catalyst for acetylation of hydroxyl groups in bagasse. At a concentration of 1% of the catalyst in acetic anhydride, a weight percent gain of 24.7% was achieved at 120 degrees C for 1 h, compared with 5.1% for the un-catalyst reaction under the same reaction condition. FT-IR and CP-MAS 13C-NMR studies produced evidence for acetylation. The thermal stability of the products decreased slightly upon chemical modification, but no significant decrease in thermal stability was observed for WPG > or = 24.7%. More importantly, the acetylation significantly increased hydrophobic properties of the bagasse. The oil sorption capacity of the acetylated bagasse obtained at 80 degrees C for 6 h, was 1.9 times higher than the commercial synthetic oil sorbents such as polypropylene fibres. Therefore, these oil sorption-active materials can be used to substitute non-biodegradable materials in oil spill cleanup.     

Renaturation and purification of rhGM-CSF with ion-exchange chromatography

The renaturation and purification of recombinant human granulocyte macrophage colony stimulation factor (rhGM-CSF) expressed in Escherichia coli with strong anion-exchange chromatography (SAX) were studied. The effects of pH values, ratios of concentrations of GSH/GSSG, and urea concentrations in the mobile phase on the renaturation and purification of rhGM-CSF with SAX were investigated, respectively. The results show that the above three factors have remarkable influences on the efficiency of renaturation and mass recovery of rhGM-CSF. The addition of GSH/GSSG in the mobile phase can improve the formation of correct disulfide bonds in rhGM-CSF so that its renaturation yield increases. In addition, to enhance the mass recovery of rhGM-CSF with SAX, the low concentration of urea was added in the mobile phase to prevent denatured protein aggregation. Under the optimal conditions, rhGM-CSF was renatured with simultaneous purification on SAX column within 30 min only by one step. After that its specific bioactivity, mass recovery, and purity reached 1.66 x 10(7) IU x mg, 58.8%, and 96.2%, respectively.     

A facile transphosphatidylation reaction using a culture supernatant of actinomycetes directly as a phospholipase D catalyst with a chelating agent

An attempt was made to use the phospholipase D (PLD)- containing culture supernatants of actinomycetes directly as catalysts for the transphosphatidylation reaction of phosphatidylcholine (PC) to phosphatidylethanolamine (PE) in a biphasic system. Of the five actinomycetes (three Streptomyces sp. and two Streptoverticillium sp.) examined, three (St. mediocidicus, Stv. cinnamoneum and Stv. hachijoense) exhibited good PLD production performance, but the selectivity (ratio of transphosphatidylation to hydrolysis) of the PLDs in the culture supernatant of all three actinomycetes were significantly low. However, the addition of EDTA to the reaction mixture as a chelating agent remarkably improved the selectivity of the PLDs, which approached 100% in all the culture supernatants. Commercially available PLDs were also investigated and classified into two types. The PLDs of one type had high selectivity and no metal was required for the enzyme activity, while those of the other type showed low selectivity and a metal was necessary for the enzyme to be activated. From this finding, it was considered that the culture supernatants used in this study contained several PLDs of both types. When the chelating agent was added to the reaction mixture, the hydrolysis due to PLDs with low selectivity was suppressed by removal of the essential metal, resulting in an increased in the overall selectivity of the PLDs in the culture supernatant. Repeated batch transphosphatidylation reactions were performed 20 times, reusing the PLDs in the aqueous phase by centrifugation; the reaction rate gradually decreased to 60% of that of batch 1 by batch 20. This suggests that the transphosphatidylation reaction using a culture supernatant has potential for industrial application. (c) 1994 John Wiley & Sons, Inc.     

Concentration and purification of rubella virus using monolithic chromatographic support

The production of economically acceptable viral vaccines of high quality requires simple and efficient methods for purification and concentration of viral particles. Ion-exchange chromatography (IEC) has become one of commonly used methods for large-scale downstream purification of viruses. Viruses possess different biological and/or biochemical properties and therefore IEC conditions must be established specifically for each virus. Live attenuated rubella virus vaccines have been manufactured and successfully used widely to protect people from rubella and congenital rubella syndrome for almost 40 years. The aim of this study was to search for an efficient method for concentration and purification of rubella virus using IEC. The selected operating conditions using quaternary amine monolithic supports enabled highly efficient binding, purification and concentration of rubella virus from complex biological suspension without additional procedures. Eluted viral particles maintained their infectivity and viral recovery was almost 100%. At the same time, viral preparation was successfully depleted from host cell protein and DNA. This work indicates the possibility of using monoliths to improve the rubella virus yields in productions where high virus titers during cultivation can hardly be achieved.     

Isolation and purification of plastocyanin from spinach stored frozen using hydrophobic interaction and ion-exchange chromatography

A new simple three-day procedure for preparative isolation and purification of plastocyanin from spinach stored in the frozen state is described. This procedure is based on batch adsorption on ion-exchange resin, ammonium sulphate precipitation, and purification on a Phenyl-Sepharose hydrophobic interaction column and a single Q Sepharose High Performance ion-exchange column. Approximately 100 mg of plastocyanin with an absorbance ratio A₂₇₈/A₅₉₇ of 1.10±0.02 in the oxidized state was typically obtained from 12 kg of spinach leaves. The purified spinach plastocyanin is shown to be homogeneous to the resolution of free solution capillary electrophoresis.Abbreviations MES 2(N-morpholino)ethanesulfonic acid - Tris Tris(hydroxymethyl)aminomethane - FSCE free solution capillary electrophoresis     

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.     

Aminopropyl silica gel as a solid support for preparation of glycolipid immunoadsorbent and purification of antibodies.

Aminopropyl silica gel was prepared from porous silica gel and was used as a solid support for immunoadsorbent in the purification of anti-glycolipid antibodies. For neutral glycosphingolipids, a carboxyl function was generated by oxidation of the olefinic double bond of the sphingosine moiety, whereas for gangliosides the carboxyl group of sialic acid was used to couple with aminopropyl silica gel in the presence of a carbodiimide. These compounds were used for purifying anti-glycolipid antibodies from serum of immunized rabbits. The antibodies bound to the su-strate were released by 2 M potassium thiocyanate and their immunological properties were studied. Aminopropyl silica gel may be preferred over conventional organic solid supports for the following reasons: 1) faster flow rate; 2) higher capacity; 3) easier handling; 4) more economical; and 5) lower susceptibility to microbial attack.     

Application of mixed modal resin for purification of a fibrinolytic enzyme

Recent advances in purification technologies for therapeutic molecules have stirred the research consortium. Mixed mode chromatography, having multiple interactions with the solute molecule, has drawn significant attention due to its overall advantage over traditional ion-exchange and reverse-phase chromatography. Capto adhere, a mixed mode chromatography resin with strong anion-exchange and reverse-phase interaction with solutes, was explored for purification of fibrinolytic enzyme from Bacillus sphaericus MTCC 3672. Static and dynamic resin binding study revealed that 30°C temperature, pH 8, and 0.5 mL/min flow rate were optimum for maximum binding of fibrinolytic enzyme. Maximum static dynamic binding and breakthrough capacities for Capto adhere were 249 and 196 U/mL of resin, respectively. Final purification with Sephadex G 100 gel chromatography resulted in 38-fold purity of fibrinolytic enzyme with 39% enzyme recovery. Purified enzyme was further characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) analysis to homogeneity, and molecular mass was found to be around 55–70 kD. Like most of the serine alkaline proteases, purified fibrinolytic enzyme was stable in a temperature range of 25–40°C and pH range of 7–9. Offshoots of our research findings have revealed a broad application area of mixed mode chromatography.     

Peptide synthesis on a phenolic resin support

Protected peptides assembled on a phenolic resin support were cleared by peroxide-catalysed hydrolysis. In genenal peptide phenyl ester resins were more labile to nucleophiles than were corresponding Merrifield resin derivatives; transesterification with dimethylaminoethanol providing on alternative cleavage method for peroxide-sensitive peptides. Losses of radiolabelled peptide from both Merrifield and phenolic resins were determined during acid deprotection, base wash and coupling steps in the synthesis of a tetrapeptide. Using 40% (v/v) trifluoroacetic acid in dichloromethane for Boc-deprotection the phenolic resin gave improved results compared to the Merrifield resin. The merits of the procedure for the preparation of protected peptide acids suitable for subsequent condensation reactions were exemplified by the synthesis of an octapeptide sequence of a modified lysozyme.     

A continuous process for biodiesel production in a fixed bed reactor packed with cation-exchange resin as heterogeneous catalyst

Continuous esterification of free fatty acids (FFA) from acidified oil with methanol was carried out with NKC-9 cation-exchange resin in a fixed bed reactor with an internal diameter of 25 mm and a height of 450 mm to produce biodiesel. The results showed that the FFA conversion increased with increases in methanol/oil mass ratio, reaction temperature and catalyst bed height, whereas decreased with increases in initial water content in feedstock and feed flow rate. The FFA conversion kept over 98.0% during 500 h of continuous esterification processes under 2.8:1 methanol to oleic acid mass ratio, 44.0 cm catalyst bed height, 0.62 ml/min feed flow rate and 65°C reaction temperature, showing a much high conversion and operational stability. Furthermore, the loss of sulfonic acid groups from NKC-9 resin into the production was not found during continuous esterification. In sum, NKC-9 resin shows the potential commercial applications to esterification of FFA.