Bioseparation最新文献

Clavulanic acid (CA) is a beta-lactamase inhibitor produced by strains of Streptomyces clavuligerus. Nowadays, the combination of CA with amoxycillin is the most successful example of the use of a beta-lactam antibiotic sensitive to beta-lactamases together with an inhibitor of these enzymes. Clavulanic acid is purified from fermentation broth by a series of steps consisting mainly of two-phase separation processes such as liquid-liquid extraction, adsorption or ion-exchange chromatography, among others. Amberlite IRA 400, a strong anion-exchange resin, has a very high adsorption capacity for CA (Mayer et al. 1997). This resin can be pre-treated with NaCl (chloride cycle), to remove selectively only those anions, which are able to displace chloride from the resin or with NaOH (hydroxyl cycle), to remove all species of anions. In order to decide the best operating conditions for CA recovery by ion-exchange resins and then to construct a model of this separation process, batch experiments were conducted using Amberlite IRA 400 in the chloride cycle. These runs were carried out in a 200 ml stirred tank, at two different initial solution pH, 6.2 and 4.0; the temperature was maintained at 10 degrees C and 20 degrees C during adsorption and 30 degrees C during the desorption step. It was possible, on the basis of these batch results, to model the separation process, including the adsorption kinetics, equilibrium data and mass transfer limitations.

The adsorption and desorption pattern of alkaline protease was studied using different aliphatic and aromatic hydrophobic ligands. Overall, higher adsorption was obtained on ligands coupled to 6% cross-linked gel than the 4% gel. The highest adsorption was obtained on butyl (94%) and phenyl (98.4%) of 6% cross-linked gel. The adsorption was dependent on concentration and nature of the ligand. In a single-step operation, almost 20-fold purification with 40% yield of the enzyme was obtained using all the optimized experimental parameters.

The selectivity of carbohydrate-imprinted polymers for several disaccharides, namely cellobiose, maltose, lactose and gentiobiose, is investigated. An ternary ligand-Cu(II)-carbohydrate complex was formed in alkaline solution and captured afterwards in the polymer. The accessibility of the polymer matrix for disaccharides was investigated by HPLC analysis, refractometry and 1H NMR spectroscopy applying excess of the original template during rebinding experiments under saturation conditions in unbuffered, aqueous solution at neutral pH and 20 degrees C. The selective discrimination of the alpha- and beta-glycosidic linkage of cellobiose and maltose is demonstrated. It is further shown, that the disaccharide-imprinted polymers slightly distinguish between the 1,4-beta- and the 1,6-beta-glycosidic linkage of cellobiose and gentiobiose, while cellobiose and lactose are not selectively recognized. Due to the weak apparent binding constant of the functional Cu(II) monomers with the targeted disaccharides at physiological pH, the recognition process is dominated by the shape of the created imprinted cavity under the applied conditions.

HPLC-based separations of amino acids and peptides, nucleotide bases, drugs, sugars and steroids using molecularly imprinted polymers (MIPs) have been reviewed in this article. The molecular recognition mechanisms of the template molecules on the MIPs in organic and aqueous eluents were discussed. Furthermore, new polymerization methods suitable for preparations of HPLC columns and packing materials using molecular imprinting techniques, and their applications to HPLC-based separations are also dealt with.

Molecularly imprinted polymers (MIPs) for quercetin have been successfully prepared by a thermal polymerization method using 4-vinylpyridine (4-VP) and ethylene glycol dimethacrylate (EDMA) as functional monomer and cross-linker, respectively. The obtained molecularly imprinted polymers were evaluated by HPLC using organic eluents, with respect to their selective recognition properties for quercetin and related compounds of the flavonoid class. Two equivalent control polymers, a blank polymer and a polymer imprinted with a structural analogous template, were synthesized, in order to confirm the obtained results. Furthermore, preliminary experiments confirm the applicability of the prepared MIPs for solid phase extraction (SPE), as rapid and facile clean-up of wine samples for HPLC analysis is an envisaged field of application. The successful preparation of molecularly imprinted polymers for flavones provides an innovative opportunity for the development of advanced separation materials, with applications in the field of wine and fermentation analysis.

Acoustic field assisted demixing was employed to decrease the demixing time in polymer-polymer (polyethylene glycol-maltodextrin) two-phase system. Application of acoustic field has decreased the demixing time in these systems up to 2-fold. Ultrasonication has induced mild circulation currents in the phase dispersion, which has enhanced the rate of droplet coalescence, eventually resulting in decreased demixing time. In polymer-polymer systems, phase demixing was found to depend greatly on which of the phases is continuous and viscosity of the continuous phase was observed to have a strong influence on the movement of the droplets and hence the phase demixing. Addition of NaCl increased the demixing time and presence of E. coli cells did not seem to have any influence on phase demixing.

The isolation and preliminary characterization of a trypsin-like protease with high activity at 20 degrees C is described. This protease was isolated from Antarctic krill (Euphasia superba) by a two-step chromatography process and the use of zymogram analyses. The protease has a molecular weight of 30 kDa and a pI of 4.1. Its specific activity at 20 degrees C on BAPNA is 0.5 U/mg.