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  • Lim, H, et al. (författare)
  • Benzo[a]pyrene-specific online high-performance liquid chromatography fractionation of air particulate extracts : a tool for evaluating biological interactions.
  • 2014
  • Ingår i: Journal of chromatography. A. - Stockholm : Karolinska Institutet, Institute of Environmental Medicine. - 0021-9673.
  • Tidskriftsartikel (refereegranskat)abstract
    • Benzo[a]pyrene (B[a]P) is a known human carcinogen and is commonly used as a surrogate for assessing the carcinogenic risk posed by complex mixtures of polycyclic aromatic hydrocarbons (PAHs) present in air particulate matter (PM). However, studies have shown that using B[a]P as a surrogate may underestimate the carcinogenic potential of PAH mixtures, as the risk assessment approach does not consider interaction effects. Thus, toxicological studies using B[a]P to assess its carcinogenic potential in environmentally derived complex mixtures, as opposed to single compound experiments, could improve risk assessment. The intention of the present study was to develop an online HPLC fractionation system for the selective removal of B[a]P from air PM extracts. Two serial pyrenylethyl (PYE) columns enabled selective separation of B[a]P from its isomers and other PAHs as well as a short fractionation cycle of 30min. One run consisted of three collection steps: the first fraction contained PAHs eluting earlier than B[a]P, the second contained B[a]P and the last contained later-eluting PAHs. The selectivity and recovery of the system was investigated using extracts of Stockholm air PM samples. The overall recovery for all PAHs was approximately 80%, and the system proved to be selective, as it removed 94% of B[a]P and less than 3% of benzo[b]fluoranthene from the complex PAH mixture. Exposing human cells to blanks generated by the fractionation system did not induce cytotoxicity or DNA damage signalling. In conclusion, the online HPLC system was selective for B[a]P fractionation whilst minimising run-to-run variation and allowing repeated fractionations for larger samples due to its relatively short run time.
  • Nilsson, GA, et al. (författare)
  • New halogen-specific detector applied to the analysis of chlorinated fatty acids
  • 2001
  • Ingår i: Journal of Chromatography A. - : Elsevier. - 0021-9673. ; 912:1, s. 99-106
  • Tidskriftsartikel (refereegranskat)abstract
    • A new halogen-specific detection method (XSD) was tested for determination of chlorinated fatty acids in marine biota. In XSD, an increased emission of ions and electrons is caused by the high-temperature combustion of halogen-containing compounds. The detection limit of methyl dichlorooctadecanoate and the selectivity at a reactor temperature of 900 degreesC match those of electrolytic conductivity detection (ELCD). The relative standard deviation is less than 11% for greater than or equal to0.2 ng methyl dichlorooctadecanoate. An XSD chromatogram of a complex sample, chlorinated fatty acid methyl esters liberated from fish lipids, agreed with a previously obtained ELCD chromatogram. (C) 2001 Elsevier Science B.V. All rights reserved.
  • Arfvidsson, Cecilia, et al. (författare)
  • Mass overloading in the flow field-flow fractionation channel studied by the behaviour of the ultra-large wheat protein glutein.
  • 2003
  • Ingår i: Journal of Chromatography A. - : Elsevier. - 0021-9673. ; 1011:1-2, s. 99-109
  • Tidskriftsartikel (refereegranskat)abstract
    • Flow field-flow fractionation (FFF) has previously been used in successful fractionation and characterisation of the ultra-large wheat protein glutenin. The many parameters, which may influence the retention behaviour, especially when analysing extremely high-molecular-mass samples such as glutenin, are here reported. Size determination from the sample retention time, using FFF theory, will as a result have a very low accuracy. The need for direct molecular mass determination, such as by light scattering, in combination with FFF, in order to do accurate size measurements of glutenin is pointed out as well as the importance to minimise the overloading.
  • Arvidsson, Pär, et al. (författare)
  • Chromatography of microbial cells using continuous supermacroporous affinity and ion-exchange columns
  • 2002
  • Ingår i: Journal of Chromatography A. - : Elsevier. - 0021-9673. ; 977:1, s. 27-38
  • Tidskriftsartikel (refereegranskat)abstract
    • Continuous supermacroporous chromatographic columns with anion-exchange ligands [2-(dimethylanlino)ethyl group] and immobilized metal affinity (IMA) ligands (Cu2+-loaded iminodiacetic acid) have been developed allowing binding of Escherichia coli cells and the elution of bound cells with high recoveries. These poly(acrylamide)-based continuous supermacroporous columns have been produced by radical co-polymerization of monomers in aqueous solution frozen inside a column (cryo-polymerization). After thawing, the column contains a continuous matrix (so-called cryogel) with interconnected pores of 10-100 mum in size. The large pore size of the matrix makes it possible for E. coli cells to pass unhindered through a plain column containing no ligands. E. coli cells bound to an ion-exchange column at low ionic strength were eluted with 70-80% recovery at NaCl concentrations of 0.35-0.40 M, while cells bound to an IMA-column were eluted with around 80% recovery using either 10 mM imidazole or 20 mM EDTA solutions, respectively. The cells maintain their viability after the binding/elution procedure. These preliminary results indicate that microbial cells can be handled in a chromatographic mode using supermacroporous continuous columns. These columns are easy to manufacture from cheap and readily available starting materials, which make the columns suitable for single-time use. (C) 2002 Published by Elsevier Science B.V.
  • Arvidsson, Pär, et al. (författare)
  • Direct chromatographic capture of enzyme from crude homogenate using immobilized metal affinity chromatography on a continuous supermacroporous adsorbent
  • 2003
  • Ingår i: Journal of Chromatography A. - : Elsevier. - 0021-9673. ; 986:2, s. 275-290
  • Tidskriftsartikel (refereegranskat)abstract
    • A continuous supermacroporous matrix has been developed allowing direct capture of enzyme from non-clarified crude cell homogenate at high flow-rates. The continuous supermacroporous matrix has been produced by radical co-polymerization of acrylamide, allyl glycidyl ether and N,N′-methylene-bis(acrylamide) which proceeds in aqueous solution of monomers frozen inside a column (cryo-polymerization). After thawing, the column contains a continuous matrix having interconnected pores of 10–100 m size. Iminodiacetic acid covalently coupled to the cryogel is a rendering possibility for immobilized metal affinity chromatographic purification of recombinant His-tagged lactate dehydrogenase, (His)6-LDH, originating from thermophilic bacterium Bacillus stearothermophilus, but expressed in Escherichia coli. The large pore size of the adsorbent makes it possible to process particulate-containing material without blocking the column. No preliminary filtration or centrifugation is needed before application of crude extract on the supermacroporous column. A total of 210 ml crude homogenate, 75 ml of it non-clarified, was processed on a single 5.0 ml supermacroporous column at flow speeds up to 12.5 ml/min without noticeable impairment of the column properties. Mechanically the cryogel adsorbent is very stable. The continuous matrix could easily be removed from the column, dried at 70 °C and kept in a dry state. After rehydration and reinsertion of the matrix into an empty column, (His)6-LDH was purified as efficiently as on the newly prepared column. The procedure of manufacturing the supermacroporous continuous cryogel is technically simple. Starting materials and initiators are cheap and available and are simply mixed and frozen under specified conditions. Altogether these qualities reveal that the supermacroporous continuous cryogels is a very interesting alternative to existing methods of protein purification from particulate-containing crude extracts
  • Barinaga-Rementeria Ramírez, Irene, et al. (författare)
  • Affinity partitioning for membrane purification exploiting the biotin-NeutrAvidin interaction - Model study of mixed liposomes and membranes
  • 2002
  • Ingår i: Journal of Chromatography A. - : Elsevier. - 0021-9673. ; 971:1-2, s. 117-127
  • Tidskriftsartikel (refereegranskat)abstract
    • Biotinylated negatively charged liposomes as well as membranes were affinity partitioned in an aqueous poly(ethylene glycol)-dextran two-phase system using NeutrAvidin conjugated to dextran as affinity ligand. Both liposomes and membranes redistributed from top to bottom phase upon addition of NeutrAvidin-dextran. The presence of 35-60 mM Li2SO4 was necessary both to force the components into the top phase without ligand and for ligand-dependent redistribution into the bottom phase. Attaching biotin via a hexanamidohexanoyl spacer and an increased density of biotin or NeutrAvidin enhanced the affinity separation. The separation conditions in these model experiments provide a basis for affinity partitioning of membranes using other affinity ligands.
  • Berggren, Kristina, et al. (författare)
  • Effects of salts and the surface hydrophobicity of proteins on partitioning in aqueous two-phase systems containing thermoseparating ethylene oxide-propylene oxide copolymers
  • 1995
  • Ingår i: Journal of Chromatography A. - : Elsevier. - 0021-9673. ; 718:1, s. 67-79
  • Tidskriftsartikel (refereegranskat)abstract
    • The partitioning of five well-characterised model proteins, bovine serum albumin (BSA), lysozyme, [beta ]-lactoglobulin A, myoglobin and cytochrome c, in aqueous two-phase systems has been studied. As top phase polymers PEG (polyethylene glycol, 100% EO) and the thermoseparating ethylene oxide (EO)-propylene oxide (PO) random copolymers, Ucon 50-HB-5100 (50% EO, 50% PO) and EO30PO70 (30% EO, 70% PO), respectively, were used. The top phase polymers are increasing in hydrophobicity with increasing content of PO. Reppal PES 200 (hydroxypropyl starch) was used as the bottom phase polymer. Phase diagrams for Reppal PES 200-PEG and Reppal PES 200-EO30PO70 two-phase systems were determined. The partitioning of four salts with different hydrophobicity, and also the effect of the salts on protein partitioning in these systems, was studied. It was found that the partitioning of the salts followed the Hofmeister series. The partitioning of proteins with low surface hydrophobicity, myoglobin and cytochrome c, was little affected by hydrophobic polymers and salts. However, the partitioning of a protein with higher surface hydrophobicity, lysozyme, was strongly affected when polymer hydrophobicity was increased and a hydrophobic counterion was used. A protein with a relatively hydrophobic surface can be partitioned to a phase containing a thermoseparating EO-PO copolymer by using a hydrophobic counterion. The partitioning of lysozyme and cytochrome c in the polymer-water system formed after temperature-induced phase separation was also examined. Both proteins partitioned exclusively to the water phase. A separation of the protein and polymer was obtained by temperature-induced phase separation on the isolated phase containing the EO-PO copolymer. The partitioning data also indicated that the hydroxypropyl starch polymer had a weak negative charge.
  • Blomquist, G., et al. (författare)
  • Reproducibility of pyrolysis-gas chromatographic analyses of the mold Penicillium brevi-compactum
  • 1979
  • Ingår i: Journal of Chromatography A. - : Elsevier. - 0021-9673. ; 173:1, s. 7-17
  • Tidskriftsartikel (refereegranskat)abstract
    • The reproducibility of multivariate measurements is discussed. Reproducibility of a second kind is defined, in which part of the variability between samples is described by a principal components model. The use of this generalized reproducibiity is shown to give an improved precision in the pyrolysis-gas chromatography of a Penicillium species.
  • Brüggemann, Oliver, et al. (författare)
  • New configurations and applications of molecularly imprinted polymers
  • 2000
  • Ingår i: Journal of Chromatography A. - : Elsevier. - 0021-9673. ; 889:1-2, s. 15-24
  • Tidskriftsartikel (refereegranskat)abstract
    • Molecularly imprinted polymers (MIPs) are applicable in a variety of different configurations. For example, bulk polymers imprinted with β-lactam antibiotics are presented to be used as stationary phases for the chromatographic separation of β-lactam antibiotics with both aqueous and organic mobile phases. However, in some analytical applications, monosized spherical beads are preferred over the currently used ground bulk polymers. A precipitation polymerization technique allows preparation of monosized spherical imprinted beads with diameters down to 200 nm having excellent recognition properties for different target molecules. Nevertheless, with current imprinting protocols a substantial amount of template has to be used to prepare the polymer. This can be problematic if the template is poorly soluble, expensive or difficult to obtain. It is shown that for analytical applications, the functional monomer:template ratio can be drastically increased without jeopardizing the polymer's recognition properties. Furthermore, a substantial reduction of the degree of crosslinking is demonstrated, resulting in much more flexible polymers that are useful for example the preparation of thin imprinted films and membranes for sensors. Apart from analysis, MIPs also are applicable in chemical or enzymatic synthesis. For example, MIPs using the product of an enzyme reaction as template are utilized for assisting the synthetic reaction by continuously removing the product from the bulk solution by complexation. This results in an equilibrium shift towards product formation. Copyright (C) 2000 Elsevier Science B.V.
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