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Chemical Extraction (chemical + extraction)
Selected AbstractsIntensified Process for the Purification of an Enzyme from Inclusion Bodies Using Integrated Expanded Bed Adsorption and RefoldingBIOTECHNOLOGY PROGRESS, Issue 4 2006Matthew H. Hutchinson This work describes the integration of expanded bed adsorption (EBA) and adsorptive protein refolding operations in an intensified process used to recover purified and biologically active proteins from inclusion bodies expressed in E. coli. ,5 -3-Ketosteroid isomerase with a C-terminal hexahistidine tag was expressed as inclusion bodies in the cytoplasm of E. coli. Chemical extraction was used to disrupt the host cells and simultaneously solubilize the inclusion bodies, after which EBA utilizing immobilized metal affinity interactions was used to purify the polyhistidine-tagged protein. Adsorptive refolding was then initiated in the column by changing the denaturant concentration in the feed stream from 8 to 0 M urea. Three strategies were tested for performing the refolding step in the EBA column: (i) the denaturant was removed using a step change in feed-buffer composition, (ii) the denaturant was gradually removed using a gradient change in feed-buffer composition, and (iii) the liquid flow direction through the column was reversed and adsorptive refolding performed in the packed bed. Buoyancy-induced mixing disrupted the operation of the expanded bed when adsorptive refolding was performed using either a step change or a rapid gradient change in feed-buffer composition. A shallow gradient reduction in denaturant concentration of the feed stream over 30 min maintained the stability of the expanded bed during adsorptive refolding. In a separate experiment, buoyancy-induced mixing was completely avoided by performing refolding in a settled bed, which achieved comparable yields to refolding in an expanded bed but required a slightly more complex process. A total of 10% of the available KSI,(His6) was recovered as biologically active and purified protein using the described purification and refolding process, and the yield was further increased to 19% by performing a second iteration of the on-column refolding operation. This process should be applicable for other polyhistidine tagged proteins and is likely to have the greatest benefit for proteins that tend to aggregate when refolded by dilution. [source] Fractionated extraction of polycyclic aromatic hydrocarbons (PAHs) from polluted soils: estimation of the PAH fraction degradable through bioremediationEUROPEAN JOURNAL OF SOIL SCIENCE, Issue 3 2004S. Thiele-Bruhn Summary Chemical extractions were tested to estimate the fraction of 15 polycyclic aromatic hydrocarbons (PAHs) in polluted soils degradable through bioremediation. Of 30 soil samples, differing in PAH concentrations, from 10 long-term polluted sites, four were used in screening tests conducted with 14 different extractants. PAH extractability increased in the order salt solutions < surfactant solutions < organic solvents and organic solvent,water mixtures. Closest correlations (r = 0.73,0.91) and ratios of c. 1:1 of the extracted to the degradable PAH concentrations were found with aqueous solutions of two surfactants (Genapol UDD 88, Synperonic LF/RA 30). The PAH extractability by surfactant solutions increased with surfactant concentration (0,50 mg ml,1) and with decreasing degree of PAH condensation (6,2 rings). Kinetic behaviour of PAH solubilization by surfactants resembled the kinetics of PAH degradation (r = 0.84,0.99). With repeated extractions, a finite fraction of PAHs was extracted, which is thought to be the mobile and mobilizable PAH fraction, the latter being controlled by back-diffusion from intra- and inter-particle sites. Experiments with all 30 soil samples and a priori testing with an additional 10 soil samples confirmed that single extractions with surfactant solutions (5 mg ml,1; soil:solution ratio 1:20) were suitable for estimating the degradable fraction of PAHs in soil. With separate linear regression equations for PAHs grouped according to their degree of condensation, the extracted concentrations described the degradable concentrations with correlation coefficients (r) ranging from 0.62 to 0.80 (P < 0.001). PAH extractions with the surfactant solutions can be used to assess the prospects of bioremediation of PAH-polluted sites. [source] Interaction of copper and zinc with allophane and organic matter in the B horizon of an AndosolEUROPEAN JOURNAL OF SOIL SCIENCE, Issue 2 2003C. Latrille Summary Andosols developed on basaltic material are naturally rich in metals. Organic matter and allophane, the key colloids of these soils, have a strong affinity for trace metals, but are intimately mixed so that speciation of trace metal is difficult to determine. We used three complementary approaches, namely physical fractionation, chemical extraction and potentiometric measurement, to distinguish them. Physical particle-size separations and chemical selective dissolution of allophanes and organic matter were combined to demonstrate relations between the occurrence of colloids and contents of Cu and Zn in an andic B horizon. About 22% of total soil Cu and 7% of total soil Zn were present in the < 5-,m fraction, associated with organic or amorphous mineral constituents. To support this association further, the affinity of soil colloids for Cu and Zn in a mimicked system was demonstrated. An Al-rich allophane was synthesized, and a portion of the organic matter was extracted from the clay fraction, and their reactivities towards Cu2+ or Zn2+ were studied by potentiometry. The two metallic cations displayed specific affinity towards allophane or soluble organic matter. Furthermore, the behaviour of copper and zinc in the ternary system, allophane + soluble organic matter + trace element, revealed a synergy in the surface complexation. The use of these three speciation approaches highlighted the linkage between metals and constituents, and showed how important the colloidal constituents are in the behaviour of Cu and Zn in Andosols. [source] Effects of genotype, location and baking on the phenolic content and some antioxidant properties of cereal speciesINTERNATIONAL JOURNAL OF FOOD SCIENCE & TECHNOLOGY, Issue 1 2010Valeria Menga Summary In order to assess the effect of genotype, location and their interaction on total phenolic content (TPC) of chemical extracts, the whole grains of durum and soft wheat, oat, barley and triticale were evaluated. Data showed differences in phenolic content of chemical extracts among cereal species and the analysis of variance confirmed the key role of location. Besides TPC and trolox equivalent antioxidant capacity (TEAC) values assessed by chemical extraction were compared with those obtained with an in vitro digestive enzymatic extraction. Differences were found between methanolic and enzymatic extracts, and data confirmed that enzymatic technique enhanced extraction of antioxidants but pointed out lesser differences among cereal types. The breads obtained by flours enriched with different levels of bran were also evaluated. Chemical extracts highlighted the increasing levels of antioxidants according to bran enrichments, without pointing out changes caused by baking. The enzymatic extraction instead did not show differences regarding to bran enrichments, but documented a loss in antioxidant properties of breads in respect to corresponding flours. On the other hand the scarce differences between flours and corresponding breads did not allow asserting that baking modified the TPC and TEAC, independently of the extraction methods used. Indeed, during baking process, also the observed phenolic acids profile variations did not vary the antioxidant properties of breads. [source] Development of a simulated earthworm gut for determining bioaccessible arsenic, copper, and zinc from soil,ENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 7 2009Wai K. Ma Abstract Soil physicochemical characteristics and contamination levels alter the bioavailability of metals to terrestrialinvertebrates. Current laboratory-derived benchmark concentrations used to estimate risk do not take into account site-specific conditions, such as contaminant sequestration, and site-specific risk assessment requires a battery of time-consuming and costly toxicity tests. The development of an in vitro simulator for earthworm bioaccessibility would significantly shorten analytical time and enable site managers to focus on areas of greatest concern. The simulated earthworm gut (SEG) was developed to measure the bioaccessibility of metals in soil to earthworms by mimicking the gastrointestinal fluid composition of earthworms. Three formulations of the SEG (enzymes, microbial culture, enzymes and microbial culture) were developed and used to digest field soils from a former industrial site with varying physicochemical characteristics and contamination levels. Formulations containing enzymes released between two to 10 times more arsenic, copper, and zinc from contaminated soils compared with control and 0.01 M CaCl2 extractions. Metal concentrations in extracts from SEG formulation with microbial culture alone were not different from values for chemical extractions. The mechanism for greater bioaccessible metal concentrations from enzyme-treated soils is uncertain, but it is postulated that enzymatic digestion of soil organic matter might release sequestered metal. The relevance of these SEG results will need validation through further comparison and correlation with bioaccumulation tests, alternative chemical extraction tests, and a battery of chronic toxicity tests with invertebrates and plants. [source] | ||||||||||