| 51. |
- Sparr Eskilsson, Cecilia, et al.
(author)
-
Harmful azo colorants in leather. Determination based on their cleavage and extraction of corresponding carcinogenic aromatic amines using modern extraction techniques.
- 2002
-
In: Journal of Chromatography A. - 0021-9673. ; 955:2, s. 215-227
-
Journal article (peer-reviewed)abstract
- This study concerns the possibilities of using microwave-assisted extraction (MAE) or supercritical fluid extraction (SFE) for detection of harmful azo colorants in leather. After degreasing of the leather sample with SFE there follows a reductive cleavage of the azo colorants to their corresponding aromatic amines in the MAE or SFE equipment. The aromatic amines are subsequently extracted using either MAE or SFE and then finally determined by liquid chromatography with diode-array detection. The results have been compared with recoveries obtained using the German DIN method 53316. This standard method, based on conventional solvent extraction, is used in several European countries. Overall much better recoveries were obtained using MAE or SFE. With both MAE and SFE the amine recoveries of spiked leather samples were generally above 50%. The average recoveries were 62% for MAE and 60% for SFE (solvent collection) compared to 24% with the DIN method. For genuine leather samples the recoveries decreased, especially for benzidine. In this case the average values for MAE, SFE and DIN were 54, 38 and 19%, respectively. The quantification limits in leather samples using MAE or SFE were below 1 mg/kg for all amines investigated. The within-laboratory precision was generally better than 10%, varying somewhat with the analyte considered. With the proposed methodology, the amount of hazardous organic solvents used could be decreased and the sample throughput increased with at least a factor of two with less manual handling compared to the DIN method.
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| 52. |
- Svensson, Mårten, et al.
(author)
-
Aqueous two-phase systems containing self-associating block copolymers - Partitioning of hydrophilic and hydrophobic biomolecules
- 1999
-
In: Journal of Chromatography A. - 0021-9673. ; 839:1-2, s. 71-83
-
Journal article (peer-reviewed)abstract
- A series of proteins and one membrane-bound peptide have been partitioned in aqueous two-phase systems consisting of micelle-forming block copolymers from the family of Pluronic block copolymers as one polymer component and dextran T500 as the other component. The Pluronic molecule is a triblock copolymer of the type PEO-PPO-PEO, where PEO and PPO are poly(ethylene oxide) and poly(propylene oxide), respectively. Two different Pluronic copolymers were used, P105 and F68, and the phase diagrams were determined at 30oC for these polymer systems. Since the temperature is an important parameter in Pluronic systems (the block copolymers form micellar-like aggregates at higher temperatures) the partitioning experiments were performed at 5 and 30oC, to explore the effect of temperature-triggered micellization on the partitioning behaviour. The temperatures correspond to the unimeric (single Pluronic chain) and the micellar states of the P105 polymer at the concentrations used. The degree of micellization in the F68 system was lower than that in the P105 system, as revealed by the phase behaviour. A membrane-bound peptide, gramicidin D, and five different proteins were partitioned in the above systems. The proteins were lysozyme, bovine serum albumin, cytochrome c, bacteriorhodopsin and the engineered B domain of staphylococcal protein A, named Z. The Z domain was modified with tryptophan-rich peptide chains in the C-terminal end. It was found that effects of salt dominated over the temperature effect for the water-soluble proteins lysozyme, bovine serum albumin and cytochrome c. A strong temperature effect was observed in the partitioning of the integral membrane protein bacteriorhodopsin, where partitioning towards the more hydrophobic Pluronic phase was higher at 30oC than at 5oC. The membrane-bound peptide gramicidin D partitioned exclusively to the Pluronic phase at both temperatures. The following trends were observed in the partitioning of the Z protein. (i) At the higher temperature, insertion of tryptophan-rich peptides increased the partitioning to the Pluronic phase. (ii) At the lower temperature, lower values of K were observed for ZT2 than for ZT1.
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| 53. |
- Svensson, Mårten, et al.
(author)
-
Partitioning of hydrophobic amino acids and oligopeptides in aqueous two-phase system containing self-aggregating block copolymer - Effects of temperature, salts and surfactants
- 1997
-
In: Journal of Chromatography A. - 0021-9673. ; 761:1-2, s. 91-101
-
Journal article (peer-reviewed)abstract
- The partitioning of hydrophobic amino acids and oligopeptides in the Pluronic P105-dextran-water system has been studied. Pluronic P105 is a member of a family of triblock copolymers with the structure PEO-PPO-PEO, where PEO is poly(ethylene oxide) and PPO poly(propylene oxide). The partitioning was studied for tryptophan, phenylalanine and di- and tri-peptides composed of these amino acids at 5 and 40oC. These temperatures correspond to a unimeric (5oC) and a micellar (40oC) state of the P105 molecule. Partitioning depended strongly on the temperature which is attributed to the increased hydrophobicity of Pluronic P105 with increasing temperature. However, it appears that the presence of the micelles plays no major direct role. The effect of different pH, salts and surfactants (both cationic and anionic) on partitioning has also been investigated.
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| 54. |
- Torto, Nelson, et al.
(author)
-
On-line quantitation of enzymatic mannan hydrolysates in small-volume bioreactors by microdialysis sampling and column liquid chromatography-integrated pulsed electrochemical detection
- 1996
-
In: Journal of Chromatography A. - : Elsevier BV. - 0021-9673. ; 725:1, s. 165-175
-
Journal article (peer-reviewed)abstract
- Quantitative aspects of on-line microdialysis sampling are investigated with respect to sampling in small-volume bioreactors. Three modes of microdialysis sampling; continuous-flow microdialysis sampling (CFMS), stopped-flow microdialysis sampling with continuous stirring, and stopped-flow microdialysis sampling with stopped stirring were investigated as a possible means for studying bioprocesses. The hydrolytic properties of a well characterised endomannanase from Aspergillus niger were studied using 0.01% ivory nut mannan as substrate in a 5-ml reaction vessel. On-line sampling was achieved using a microdialysis probe fitted with an SPS 6005 membrane. Stopped-flow microdialysis sampling was found to give the least analyte depletion and thus used for quantitation of the enzymatic hydrolysates. However, CFMS can be used when analyte depletion is not significant (large-volume reactor). Hydrolysis of ivory nut mannan gave mainly mannobiose and mannotriose in almost equal amounts, which is consistent with an endo-wise hydrolysis. The concentrations of mannose and mannopentaose did not change significantly over the monitoring period, however, that of mannotetraose increased gradually up to 11 h where it starts to decrease.
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| 55. |
- Turner, Charlotta, et al.
(author)
-
Collection in analytical-scale supercritical fluid extraction.
- 2002
-
In: Journal of Chromatography A. - 0021-9673. ; 947:1, s. 1-22
-
Research review (peer-reviewed)abstract
- This review is a comprehensive summary of available collection techniques in supercritical fluid extraction (SFE), with emphasis on which parameters are especially important for a successful analyte collection. Environmental, biological and agricultural applications, including several types of sample matrices and analyte groups, are discussed with respect to choice of collection mode and optimization of collection conditions. This review also includes discussions about collection when a modifier is used or when the sample contains large amounts of fat or water, as well as possibilities to achieve enhanced selectivity.
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| 56. |
- Turner, C, et al.
(author)
-
Supercritical fluid extraction and chromatography for fat-soluble vitamin analysis.
- 2001
-
In: Journal of Chromatography A. - 0021-9673. ; 936:1, s. 37-215
-
Journal article (peer-reviewed)abstract
- Extraction and chromatographic separation of fat-soluble vitamins is a challenging task, due to the sensitivity of these compounds towards light, oxygen, heat and pH. In light of this, supercritical fluid extraction (SFE) and supercritical fluid chromatography (SFC) are attractive techniques as they function at considerably milder conditions than conventional solvent-based analytical techniques. Moreover, supercritical techniques consume much less amounts of organic solvents than conventional ones. This review gives a brief description of suitable supercritical media as well as basic theory on SFE and SFC processes. Furthermore, guidelines are provided for optimizing the important extraction and separation parameters to facilitate a successful method development. Finally, applications employing SFE and/or SFC for fat-soluble vitamin enrichment and final determination are reviewed.
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| 57. |
- Welinder, Charlotte, et al.
(author)
-
Restricted access chromatographic sample preparation of low mass proteins expressed in human fibroblast cells for proteomics analysis
- 2001
-
In: Journal of Chromatography A. - 0021-9673. ; 909:2, s. 279-288
-
Journal article (peer-reviewed)abstract
- Two-dimensional electrophoresis and modern image analysis systems have made it possible to study protein expression and regulation of proteins in biological systems. Proteins in the molecular mass region of 20-120 kDa are well investigated and described. However, proteins with masses below 20 kDa are the least investigated as they are rarely seen on 2D-PAGE due to fast migrations in the electric field and lack of staining efficiency. This paper describes a technique that enriches proteins in the lower mass region using solid-phase extraction. The purification step is carried out using C18 functionalised "restricted access" affinity chromatography whereby simultaneous trace enrichment and sample clean up is achieved. In this study expression patterns of TGF-beta stimulated and non-stimulated fibroblasts were compared after the solid-phase fractionation procedure. An increased expression pattern was obtained whereby 400 protein spots could be detected by image analysis in the <20-kDa region. Out of these, specific regulations of 14 spots were found by quantitative image analysis and spots of interest were identified with MALDI TOF-MS. The regulated and identified proteins were triosephosphate isomerase, cofilin and heat shock 27-kDa protein.
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| 58. |
- Wikström, Per, et al.
(author)
-
Affinity fibre — a new support for rapid enzyme purification by high-performance liquid affinity chromatography
- 1987
-
In: Journal of chromatography. A. - 0021-9673. ; 388, s. 123-134
-
Journal article (peer-reviewed)abstract
- A new type of support for chromatographic use has been developed. Non-porous quartz fibre, with a mean diameter of 0.5 μm, was silylated with mercaptopropyltrimethoxysilane. Tresyl chloride-activated dextran was covalently coupled to the SH groups on the fibre. Remaining active tresyl groups on the dextran were then coupled with an NAD derivative.The affinity fibre contained 0.3 μmol NAD derivative per g and awas able to bind 15 mg of lactate dehydrogenase per g of fibre.The affinity fibre was used for large-scale purification of ox heart lactate dehydrogenase and its performance was compared with other commonly used chromatographic matrices. The operational capacity was found to be 1.0 g of pure lactate dehydrogenase per hour per 100 g opf fibre material (adsorption followed by salt elution). The affinity fibre was found to be particularly suitable for very rapid processing of large volumes of dilute enzyme solutions.
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| 59. |
- Wikström, Per, et al.
(author)
-
Gas phase silylation, a rapid method for preparation of high-performance liquid chromatography supports
- 1988
-
In: Journal of chromatography. A. - 0021-9673. ; 455, s. 105-117
-
Journal article (peer-reviewed)abstract
- A new method for preparing chromatographic supports is described. Porous silica was treated with gaseous silanes and gaseous triethylamine at high temperature and reduced pressure. The silylation procedure was rapid and gave supports with a covalently bound monolayer of aminopropyl-or epoxysilane. Diol silica prepared by gas phase silylation was compared with diol silica prepared in the aqueous or organic phase (toluene). Seven proteins were chromatographed and the protein recovery, plate number and peak asymmetry were calculated. Silica prepared by the gas phase silylation method showed improved performance. The separation of carbonhydrates on aminopropylsilica and the isolation of lactate dehydrogenase with nicotinamide-adenine dinucleotide-silica, prepared from gas phase silylated diol silica, are described.
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| 60. |
- Önnerfjord, Patrik, et al.
(author)
-
High sample throughput flow immunoassay utilising restricted access columns for the separation of bound and free label
- 1998
-
In: Journal of Chromatography A. - 0021-9673. ; 800:2, s. 219-230
-
Journal article (peer-reviewed)abstract
- A flow immunodetection system with high sample throughput capacity is described for the screening of various analytes. The immunochemical detection principle is based on the chromatographic separation of the formed immunocomplex (AbAg or AbAg*) and the free antigen (Ag) by a restricted access (RA) column, utilising size-exclusion and reversed-phase mechanism. A fluorescein labelled analyte (Ag*) was used in the competitive assay format with fluorescence detection. The speed and simplicity of the assay were the greatest advantages, allowing measurement of the analyte to be carried out in less than 1 min. The biocompatibility and capacity of the restricted access material allowed multiple injections of up to 5000, without any breakthrough of the fluorescent tracer molecule and thus need for regeneration. The flow immunoassay was developed using the well-known atrazine herbicide and some transformation products as model compounds, due to their human toxicity and widespread use. The sample throughput was 80 samples per hour and the detection limits were 1.4 nM (300 pg/ml) for atrazine (Ab I) and 2.3 nM (500 pg/ml) for the sum of triazines (Ab II-III). Different sample matrices, PBS buffer, creek water, and urine were successfully applied in the flow system without the need for any sample handling step. For plasma samples an additional clean-up step using solid-phase extraction had to be included. The resulting detection limits for atrazine in plasma and water samples using this clean-up and trace enrichment procedure were found to be 2 ng/ml and 20 pg/ml, respectively. The analysis could be performed at a sample throughput rate of 400 per 6-h working shift.
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