ABSTRACT
BACKGROUND: Increased autocorrelation (AR) of system-specific measures has been suggested as a predictor for critical transitions in complex systems. Increased AR of mood scores has been reported to anticipate depressive episodes in major depressive disorder, while other studies found AR increases to be associated with depressive episodes themselves. Data on AR in patients with bipolar disorders (BD) is limited and inconclusive. METHODS: Patients with BD reported their current mood via daily e-diaries for 12 months. Current affective status (euthymic, prodromal, depressed, (hypo)manic) was assessed in 26 bi-weekly expert interviews. Exploratory analyses tested whether self-reported current mood and AR of the same item could differentiate between prodromal phases or affective episodes and euthymia. RESULTS: A total of 29 depressive and 20 (hypo)manic episodes were observed in 29 participants with BD. Self-reported current mood was significantly decreased during the two weeks prior to a depressive episode (early prodromal, late prodromal), but not changed prior to manic episodes. The AR was neither a significant predictor for the early or late prodromal phase of depression nor for the early prodromal phase of (hypo)mania. Decreased AR was found in the late prodromal phase of (hypo)mania. Increased AR was mainly found during depressive episodes. CONCLUSIONS: AR changes might not be better at predicting depressive episodes than simple self-report measures on current mood in patients with BD. Increased AR was mostly found during depressive episodes. Potentially, changes in AR might anticipate (hypo)manic episodes.
Subject(s)
Bipolar Disorder , Prodromal Symptoms , Self Report , Humans , Bipolar Disorder/physiopathology , Bipolar Disorder/psychology , Female , Male , Adult , Middle Aged , Affect/physiology , Mania , Young AdultABSTRACT
This study has examined the batch binding behaviour of different thermo-responsive co-polymer grafted chromatographic materials under different temperature and protein loading conditions. The effect of molecular composition of poly(N-isopropylacrylamide) (PNIPAAm)-based co-polymers on the phase transition properties has been documented. Sixteen co-polymers of different compositions were synthesized by free radical polymerization methods. Most underwent relatively sharp phase transitions upon application of increasing temperature. However, the value of the lower critical solution temperature (LCST) varied due to differences in co-polymer compositions. In general, it was found that the LCST increased for co-polymers containing more hydrophilic moieties, but decreased for co-polymers with more hydrophobic moieties. Moreover, the LCST increased, together with increased width of the transition temperature, when highly branched monomeric moieties (i.e. Ntertoctyl groups) were present. When bulky side chains (octadecyl or triphenylmethyl groups) were located in the polymer structures the LCST transition was absent. Based on these findings, 6 thermo-responsive co-polymers of different compositions were individually immobilised onto cross-linked Sepharose 6 Fast Flow by a "grafting-from" method. Bovine holo-lactoferrin and bovine holo-transferrin at different concentrations in the range 1-100 mg/mL were then employed as target proteins to evaluate the adsorption behaviour under batch binding conditions with these different polymer grafted Sepharose 6 Fast Flow sorbents at two different temperatures. In general, all sorbents exhibited greater affinity and adsorption capacity for bovine holo-lactoferrin at 50 °C compared to 20 °C. In addition, the affinity and adsorption capacity of bovine holo-lactoferrin with positively charged copolymer grafted Sepharose 6 Fast Flow chromatographic sorbents at 20 °C and 50 °C were much lower than that found for negatively charged copolymer grafted Sepharose 6 Fast Flow sorbents, whilst the opposite trend was found with bovine holo-transferrin due to differences in the surface charge properties of these two proteins, indicative of different separation selectivity. Furthermore, the structure of the side chains present in the grafted copolymer structure was found to affect the adsorption performance of both proteins at 20 °C and 50 °C.
Subject(s)
Polymers/chemistry , Proteins/chemistry , Rheology , Sepharose/chemistry , Temperature , Acrylic Resins/chemical synthesis , Acrylic Resins/chemistry , Adsorption , Animals , Cattle , Hydrophobic and Hydrophilic Interactions , Lactoferrin/chemistry , Polymerization , Polymers/chemical synthesis , Transferrin/chemistryABSTRACT
This investigation describes an integrated workflow for the parallel extraction and recovery of polyphenols and phytosterols from Pinot noir grape seeds. Using (E)-resveratrol and stigmasterol as exemplars, the approach employs two different molecular imprinted polymers in tandem for the extraction of these compounds and their subsequent analysis by capillary high-performance liquid chromatography (capHPLC) interfaced with electrospray ionisation tandem mass spectrometry (ESI MS/MS). Information on the selectivity of the solid-phase extraction processes was obtained through analysis of the binding behaviour of (E)-resveratrol- and stigmasterol-imprinted polymers using structurally similar polyphenols or phytosterols with the extent of binding determined from the capHPLC-ion trap ESI MS/MS data. This study documents with Pinot noir grape seed extracts and optimised solid-phase extraction protocols that the (E)-resveratrol-templated MIP enabled a very high recovery (99%) of the health-beneficial polyphenol (E)-resveratrol with co-purification of procyanidin and catechin/epicatechin. Further, the stigmasteryl-3-O-methacrylate-templated polymer resulted in high recovery (96%) of the phytosterol stigmasterol with co-purification of campesteryl glycoside. The results also demonstrate that rapid and high-resolution capHPLC-ESI MS/MS methods can be used as part of the work flow for selectivity optimisation and monitoring of the performance of MIPs intended for use in the solid-phase extraction of bioactive molecules with nutraceutical properties from agricultural waste streams.
Subject(s)
Molecular Imprinting , Phytosterols/chemistry , Phytosterols/isolation & purification , Polymers/chemical synthesis , Polyphenols/chemistry , Polyphenols/isolation & purification , Vitis/chemistry , Chromatography, High Pressure Liquid , Hexanes/chemistry , Seeds/chemistry , Solvents/chemistry , Spectrometry, Mass, Electrospray Ionization , Tandem Mass SpectrometryABSTRACT
In this investigation, application of linear solvent energy relationships (LSERs) and principal component analysis (PCA) methods have been employed to investigate the structure-retention dependencies of E-resveratrol analogues separated under different stationary and mobile phase conditions. To this end, the retention of 22 analogues have been determined with phenyl, diol, bidentate anchored C18 (BDC18) and Diamond Hydride™ C18 (DHC18) substituted silica hydride stationary phases under isocratic chromatographic conditions using mobile phases containing 0.1 (% v/v) formic acid and different acetonitrile or methanol contents from 10 to 90% (v/v) in 10% increments. In general, these compounds showed decreasing retention with increased acetonitrile or methanol content in the mobile phase with all the stationary phases. The retention order generally followed their log P values, although some unique selectivity variations were apparent depending on the nature of the selected stationary and mobile phases. These 22 compounds contained different backbone functionalities linking the phenyl ring A to phenyl ring B and different numbers of hydroxyl groups in the phenyl ring A/phenyl ring B. Structure-retention descriptors, derived according to LSER concepts, were analysed by PCA methods to provide group classification of these resveratrol analogues from the associated PC1 versus PC2 score plots. These results revealed that the selectivity of these compounds was dominated by hydrophobic and steric interactions. Based on the number and position of hydroxyl groups in a specific resveratrol analogue, a reliable curve fitting approach (indicated by R2â¯>â¯0.99 for the correlation between experimental and predicted log k values) was derived for prediction of the retention of these analytes under different mobile phase isocratic separation conditions. The application of similar methods are anticipated to find general utility for the analysis of diverse classes of other low molecular mass compounds in the different modes of liquid chromatography, permitting enhanced levels of prediction and evaluation of the retention attributes of polar and non-polar compounds.
ABSTRACT
Determination of the extent of host cell protein (HCP) contamination is an essential pre-requisite to validate the chromatographic purification of recombinant proteins. This study explores how different experimental conditions affect the HCP profiles generated during the immobilised metal ion affinity chromatographic (IMAC) purification with a Ni2+-1,4,7-triaza-cyclononane (tacn) Sepharose FF™ sorbent of the Bacillus halodurans N- and C-terminal His6-tagged xylanase A, expressed by Escherichia coli BL21(DE3) cells, and captured directly from cell lysates. Comparative studies were also carried out under identical loading, wash and elution conditions using nitrilotriacetic acid (NTA), also immobilised onto an agarose support and complexed with Ni2+ ions. High-resolution tandem mass spectrometry of the tryptic peptides derived from the proteins present in the IMAC flow-through, wash and elution fractions confirmed that the E. coli BL21(DE3) HCP profiles were dependent on the choice of adsorbent. With feedstocks containing the N- or C-terminal His6-tagged xylanase A, in several instances the same E. coli BL21(DE3) HCPs were found to co-elute with the tagged protein from either adsorbent, indicating a preferential ability of some HCPs to bind to both the IMAC resin and to the recombinant protein. This promiscuous behaviour has been found to be due to factors other than just the presence of histidine-rich motifs within the amino acid sequences of these HCPs. This case study demonstrates that the choice of protein expression and separation conditions impact on the levels of HCP contamination when different IMAC systems are employed.
Subject(s)
Bacterial Proteins/isolation & purification , Chromatography, Affinity/methods , Endo-1,4-beta Xylanases/isolation & purification , Escherichia coli Proteins/chemistry , Nickel/chemistry , Adsorption , Bacillus/enzymology , Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Chromatography, Affinity/instrumentation , Endo-1,4-beta Xylanases/chemistry , Endo-1,4-beta Xylanases/genetics , Endo-1,4-beta Xylanases/metabolism , Escherichia coli/genetics , Escherichia coli/metabolism , Escherichia coli Proteins/genetics , Escherichia coli Proteins/isolation & purification , Escherichia coli Proteins/metabolism , Histidine/genetics , Histidine/metabolism , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/isolation & purification , Recombinant Fusion Proteins/metabolismABSTRACT
This review documents recent advances in the design, synthesis, characterization, and application of molecularly imprinted polymers in the form of monoliths and particles/beads for the use in the separation and analysis of proteins with solid-phase extraction or liquid chromatography. The merits of three-dimensional molecular imprinting, whereby the molecular template is randomly embedded in the polymer, and two-dimensional imprinting, in which the template is confined to the surface, are described. Target protein binding can be achieved by either using the entire protein as a template or by using a protein substructure as template, that is, a peptide, as in the "epitope" approach. The intended approach and strategy then determine the choice of polymerization method. A synopsis has been provided on methods used for the physical, chemical, and functional characterizations and associated performance evaluations of molecularly imprinted and nonimprinted control polymers, involving a diverse range of analytical techniques commonly used for low and high molecular mass analytes. Examples of recent applications demonstrate that, due to the versatility of imprinting methods, molecularly imprinted monoliths or particles/beads can be adapted to protein extraction/depletion and separation procedures relevant to, for example, protein biomarker detection and quantification in biomedical diagnostics and targeted proteomics.
Subject(s)
Molecular Imprinting , Polymers/chemistry , Proteins/analysis , Biomarkers/analysis , Chromatography, Liquid , Humans , Polymers/chemical synthesis , Solid Phase ExtractionABSTRACT
The interaction thermodynamics of the thrombin receptor agonistic peptide (TRAP-1), H-Ser-Phe-Leu-Leu-Arg-Asn-Pro-OH, and a set of alanine scan substitution peptides, have been investigated with an n-octadecylacrylic polymer-bonded porous silica (Sil-ODA18) and water-acetonitrile mobile phases at temperatures ranging from 5 to 80⯰C in 5⯰C increments. The retention of these peptides on the Sil-ODA18 stationary phase decreased as the water content in the mobile phase was lowered from 80% (v/v) to ca. 45% (v/v) and reached a minimum value for each peptide at a specific water-acetonitrile composition. Further decreases in the water content of the mobile phase led to increased retention. The magnitude of the changes in enthalpy of interaction, Δ H a s s o c 0 , changes in entropy of interaction, Δ S a s s o c 0 , and changes in heat capacity, Δ C p 0 , were found to be dependent on the molecular properties of the mobile phase, the temperature, the structure/mobility of the stationary phase, and the conformation and solvation state of the peptides. With water-rich mobile phases, the retention behaviour of the TRAP analogues was dominated by enthalpic processes, consistent with the participation of strong hydrogen bonding effects, but became dominated by entropic effects with acetonitrile-rich mobile phases as the temperature was increased. These changes in the retention behaviour of these TRAP peptides are consistent with the generation of water or acetonitrile clusters in the mobile phase depending on the volume fractions of the organic solvent as the Sil-ODA18 stationary phase transitions from its crystalline to its isotropic state.
ABSTRACT
Perfluorinated C8-(PerfluoroC8) and bidentate anchored C8-(BDC8)-modified silica hydride stationary phases have been employed for the isocratic separation of homologous phenylalkanols and phenylalkylamines differing in their n-alkyl chain length, using aqueous-acetonitrile (ACN) mobile phases of different ACN contents from 10 to 90% (v/v) in 10% increments. These analytes showed reversed-phase (RP) retention behaviour with mobile phases of <40% (v/v) ACN content with both stationary phases but with the BDC8 stationary phase providing longer retention. The PerfluoroC8, but not the BDC8, stationary phase also exhibited significant retention of these analytes under conditions typical of an aqueous normal phase (ANP) mode (i.e. with mobile phases of >80% (v/v) ACN content), with the analytes exhibiting overall U-shape retention dependencies on the ACN content of the mobile phase. Further, these stationary phases showed differences in their selectivity behaviour with regard to the n-alkyl chain lengths of the different analytes. These observations could not be explained in terms of pK a , log P, molecular mass or linear solvation energy concepts. However, density functional theory (DFT) simulations provided a possible explanation for the observed selectivity trends, namely differences in the molecular geometries and structural organisation of the immobilised ligands of these two stationary phases under different solvational conditions. For mobile phase conditions favouring the RP mode, these DFT simulations revealed that interactions between adjacent BDC8 ligands occur, leading to a stationary phase with a more hydrophobic surface. Moreover, under mobile phase conditions favouring retention of the analytes in an ANP mode, these interactions of the bidentate-anchored C8 ligands resulted in hindered analyte access to potential ANP binding sites on the BDC8 stationary phase surface. With the PerfluoroC8 stationary phase, the DFT simulations revealed strong repulsion of individual perfluoroC8 ligand chains, with the perfluoroC8 ligands of this stationary phase existing in a more open brush-like state (and with a less hydrophobic surface) compared to the BDC8 ligands. These DFT simulation results anticipated the chromatographic findings that the phenylalkanols and phenylalkylamines had reduced retention in the RP mode with the PerfluoroC8 stationary phase. Moreover, the more open ligand structure of the PerfluoroC8 stationary phase enabled greater accessibility of the analytes to water solvated binding sites on the stationary phase surface under mobile phase conditions favouring an ANP retention mode, leading to retention of the analytes, particularly the smaller phenylalkylamines, via hydrogen bonding and electrostatic effects.
ABSTRACT
BACKGROUND: Alcohol relapse is often occurring to regulate negative affect during withdrawal. On the neurobiological level, alcoholism is associated with gray matter (GM) abnormalities in regions that regulate emotional experience such as the orbitofrontal cortex (OFC). However, no study to our knowledge has investigated the neurobiological unpinning of affect in alcoholism at early withdrawal and the associations of OFC volume with long-term relapse risk. METHODS: One hundred and eighty-two participants were included, 95 recently detoxified alcohol dependent patients (ADP) and 87 healthy controls (HC). We measured affective states using the positive and negative affect schedule (PANAS). We collected T1-weighted brain structural images and performed Voxel-based morphometry (VBM). RESULTS: Findings revealed GM volume decrease in alcoholics in the prefrontal cortex (including medial OFC), anterior cingulate gyrus, and insula. GM volume in the medial OFC was positively associated with NA in the ADP group. Cox regression analysis predicted that risk to heavy relapse at 6 months increases with decreased GM volume in the medial OFC. CONCLUSIONS: Negative affect during alcohol withdrawal was positively associated with OFC volume. What is more, increased GM volume in the OFC also moderated risk to heavy relapse at 6 months. Reduced GM in the OFC poses as risk to recovery from alcohol dependence and provides valuable insights into transient negative affect states during withdrawal that can trigger relapse. Implications exist for therapeutic interventions signifying the OFC as a neurobiological marker to relapse and could explain the inability of ADP to regulate internal negative affective states.
Subject(s)
Affect , Alcoholism/pathology , Alcoholism/therapy , Prefrontal Cortex/pathology , Biomarkers , Female , Gray Matter/pathology , Gyrus Cinguli/pathology , Humans , Magnetic Resonance Imaging , Male , Middle Aged , Recurrence , Substance Withdrawal Syndrome/pathology , Time FactorsABSTRACT
A new class of efficient stationary phase has been investigated for use in the liquid chromatographic separation of low molecular weight analytes and high molecular weight biomolecules, based on the application of immobilised stimuli-responsive polymers (SRPs). To this end, two polymeric units, namely poly(2-dimethylaminoethyl methacrylate) (PDMAEMA) and poly(acrylic acid) (PAA) were tethered to a triazine core. The derived poly(2-dimethyl-aminoethyl methacrylate)-block-poly(acrylic acid) (PDMAEMA-b-PAA), as a diblock co-polymer, was then immobilised onto the surface of porous silica particles. The performance of this microparticulate adsorbent was evaluated under various temperature, ionic strength and/or pH conditions in packed columns in a high-performance liquid chromatography (HPLC) format. Baseline separations of a variety of low molecular weight analytes were achieved at different temperatures with this SRP-based adsorbent using 10 mM sodium phosphate buffer, pH 6.0, as the mobile phase. Moreover, when the ionic strength of the mobile phase was increased to 40 mM sodium phosphate buffer, pH 6.0, similar temperature changes resulted in further increases in resolution for the hydrophobic analytes. In addition, changes in the pH of the mobile phase from pH 6.0 to pH 8.0 led to significant changes in selectivity of the analytes, including reversal in their elution orders. Upon increasing the temperature, the retention times of all analytes decreased but without loss of resolution. These findings can be attributed to the consequence of the immobilised copolymer undergoing a phase transition at its lower critical solution temperature (LCST), which leads to changes in its solvated structure, including how the electrostatic, hydrophilic and hydrophobic regions/domains of the copolymer are exposed to the bulk mobile phase. Thermodynamic data were indicative of a temperature-related re-organisation of the structure of the immobilised PDMAEMA-b-PAA stationary phase with exothermic binding of the analytes occurring at temperatures below the lower critical solution temperature (LCST). In this manner; changes in the system temperature could directly be used to manipulate the adsorption and desorption behaviour of these analytes with this stimuli-responsive, polymer-modified porous silica stationary phase. Additional studies with several proteins further documented the versatility of these stimuli-responsive separation materials. The results indicated that these separations could be tuned by variation of the temperature with fully aqueous mobile phases at specific ionic strength and pH values, without the need to use an organic solvent as a component in the mobile phase.
Subject(s)
Acrylic Resins/chemistry , Acrylic Resins/chemical synthesis , Chromatography, High Pressure Liquid/methods , Drug Design , Ethylamines/chemistry , Methacrylates/chemistry , Amitriptyline/isolation & purification , Chemistry Techniques, Synthetic , Hydrogen-Ion Concentration , Hydrophobic and Hydrophilic Interactions , Ketoprofen/isolation & purification , Osmolar Concentration , Proteins/isolation & purification , Silicon Dioxide/chemistry , TemperatureABSTRACT
This investigation demonstrates the application of a new peak sharpening technique to improve the separation of difficult-to-resolve racemic mixtures in capillary electro-chromatography. Molecularly imprinted porous layer open tubular (MIP-PLOT) capillaries, prepared by a layer-on-layer polymerization approach with Z-l-Asp-OH as the template, were selected to validate the approach. SEM revealed that the polymer film thickness can be varied by changes in both the polymer composition and the layer-on-layer regime. Capillaries made with methacrylic acid as the functional monomer could not separate the Z-Asp-OH racemate, due to weak interactions between the MIP-PLOT material and the target analytes. In contrast, MIP-PLOT capillaries prepared with 4-vinylpyridine as the functional monomer resulted in increased ionic interactions with the target analytes. Separation of the enantiomers could be enhanced when a peak zone sharpening effect was exploited through the use of specific BGE compositions and by taking advantage of eigenpeak phenomena. In this manner, the position of a sharpening zone and the peak shape of the sample analytes could be fine-tuned, so that when the sharpening zone and the target analyte co-migrated the separation of the Z-l-Asp-OH enantiomer from its d-enantiomer in a racemic mixture could be achieved under overloading conditions.
Subject(s)
Capillary Electrochromatography/instrumentation , Capillary Electrochromatography/methods , Molecular Imprinting , Polymers , Asparagine/analogs & derivatives , Asparagine/isolation & purification , Capillary Electrochromatography/standards , Equipment Design , Pyridines , StereoisomerismABSTRACT
This review describes recent advances associated with the development of surface imprinting methods for the synthesis of polymeric membranes and thin films, which possess the capability to selectively and specifically recognize biomacromolecules, such as proteins and single- and double-stranded DNA, employing "epitope" or "whole molecule" approaches. Synthetic procedures to create different molecularly imprinted polymer membranes or thin films are discussed, including grafting/in situ polymerization, drop-, dip-, or spin-coating procedures, electropolymerization as well as micro-contact or stamp lithography imprinting methods. Highly sensitive techniques for surface characterization and analyte detection are described, encompassing luminescence and fluorescence spectroscopy, X-ray photoelectron spectroscopy, FTIR spectroscopy, surface-enhanced Raman spectroscopy, atomic force microscopy, quartz crystal microbalance analysis, cyclic voltammetry, and surface plasmon resonance. These developments are providing new avenues to produce bioelectronic sensors and new ways to explore through advanced separation science procedures complex phenomena associated with the origins of biorecognition in nature.
Subject(s)
Biosensing Techniques , Chemistry Techniques, Analytical/instrumentation , Macromolecular Substances/analysis , Macromolecular Substances/isolation & purification , Polymers/chemical synthesis , Membranes, ArtificialABSTRACT
This investigation describes a general procedure for the selectivity mapping of molecularly imprinted polymers, using (E)-resveratrol-imprinted polymers as the exemplar, and polyphenolic compounds present in Pinot noir grape skin extracts as the test compounds. The procedure is based on the analysis of samples generated before and after solid-phase extraction of (E)-resveratrol and other polyphenols contained within the Pinot noir grape skins using (E)-resveratrol-imprinted polymers. Capillary reversed-phase high-performance liquid chromatography (RP-HPLC) and electrospray ionisation tandem mass spectrometry (ESI MS/MS) was then employed for compound analysis and identification. Under optimised solid-phase extraction conditions, the (E)-resveratrol-imprinted polymer showed high binding affinity and selectivity towards (E)-resveratrol, whilst no resveratrol was bound by the corresponding non-imprinted polymer. In addition, quercetin-3-O-glucuronide and a dimer of catechin-methyl-5-furfuraldehyde, which share some structural features with (E)-resveratrol, were also bound by the (E)-resveratrol-imprinted polymer. Polyphenols that were non-specifically retained by both the imprinted and non-imprinted polymer were (+)-catechin, a B-type procyanidin and (-)-epicatechin. The compounds that did not bind to the (E)-resveratrol molecularly imprinted polymer had at least one of the following molecular characteristics in comparison to the (E)-resveratrol template: (i) different spatial arrangements of their phenolic hydroxyl groups, (ii) less than three or more than four phenolic hydroxyl groups, or (iii) contained a bulky substituent moiety. The results show that capillary RP-HPLC in conjunction with ESI MS/MS represent very useful techniques for mapping the selectivity of the binding sites of imprinted polymer. Moreover, this procedure permits performance monitoring of the characteristics of molecularly imprinted polymers intended for solid-phase extraction of bioactive and nutraceutical molecules from diverse agricultural waste sources.
Subject(s)
Fruit/chemistry , Plant Extracts/chemistry , Polymers/chemistry , Polyphenols/analysis , Stilbenes/chemistry , Vitis , Binding Sites , Chromatography, High Pressure Liquid , Molecular Imprinting , Porosity , Resveratrol , Spectrometry, Mass, Electrospray Ionization , Surface Properties , Tandem Mass SpectrometryABSTRACT
A semi-covalent imprinting strategy has been developed for the synthesis of molecularly-imprinted polymers specific for the fungal sterol, ergosterol, a biological precursor of vitamin D2. This imprinting approach involved a novel post-synthesis cleavable monomer-template composite, namely ergosteryl methacrylate, and resulted in the formation of an imprinted polymer that selectively and efficiently recognized ergosterol through non-covalent interactions. The derived molecularly-imprinted polymer and the corresponding non-imprinted polymer were systematically evaluated for their selectivity towards ergosterol via static and dynamic binding studies using various ergosteryl esters (e.g. ergosteryl-cinnamate, -ferulate, -coumarate, -ferulate acetate and -acetate, respectively) as competitors. Moreover, the binding capacity of the molecularly imprinted polymer for ergosterol was enhanced when the sample loading conditions involved the use of partially aqueous solvent mixtures, such as acetonitrile/water (9:1 (v/v) or 8:2 (v/v)). These attributes were exploited in a solid-phase extraction format, whereby ergosterol was obtained with excellent recoveries from an extract of the fruiting body powder of the medicinal fungus Ganoderma tsugae var. Janniae.
Subject(s)
Chemistry Techniques, Analytical/methods , Ergosterol/isolation & purification , Ganoderma/chemistry , Polymers/chemistry , Solid Phase Extraction , Acetonitriles/chemistry , Methacrylates/chemistry , Molecular Imprinting , Solvents/chemistry , Water/chemistryABSTRACT
This study documents the use of an integrated approach, involving on-line hydrophobic interaction chromatography interfaced with Small Angle X-ray Scattering (HIC-SAXS) measurements, to monitor the conformational status of proteins immediately upon elution from a chromatographic column operated at different temperatures. Moreover, this approach provides an additional avenue to interrogate the changes in protein shape that may occur across the eluted chromatographic peak. To this end, radii of gyration were extrapolated from the Guinier approximation with the HIC-SAXS data, whilst pair distribution functions and bead model simulations were generated by using the indirect transform program GNOM and ab initio reconstruction with GASBOR to provide further insight into protein conformational changes that occur during hydrophobic interaction chromatography.
Subject(s)
Egg White/chemistry , Muramidase/chemistry , Scattering, Small Angle , X-Ray Diffraction , Animals , Chickens , Chromatography , Hydrophobic and Hydrophilic Interactions , Protein ConformationABSTRACT
Micro-patterned surfaces with alternate hydrophilic and hydrophobic rectangular areas effectively confine water droplets down to attolitre volumes. The contact angle, volume, and geometry of the confined droplets as a function of the geometry and physico-chemical properties of the confining surfaces have been determined by phenomenological simulations, validated by atomic force microscopy measurements. The combination between experiments and simulations can be used for the purposeful design of arrays with surface-addressable hydrophobicity employed in digital microfluidics and high-throughput screening nanoarrays.
ABSTRACT
A new type of stimuli-responsive polymeric (SRP) coating has been prepared for use in open tubular capillary electrochromatography (OT-CEC), by grafting poly(2-dimethylaminoethylmethacrylate)-block-poly(acrylic acid) (PDMAEMA-b-PAA) as a Y-shaped block copolymer with two dissimilar chain compositions onto the inner walls of aminopropyl-modified silica capillaries. The grafting process introduced weakly charged functional groups from the PAA and PDMAEMA, enabling the generation of electroendosmotic flow with magnitude and direction adjustable by changing the pH of the running buffer electrolyte. This stimuli-responsive PDMAEMA-b-PAA block copolymer was found to provide excellent resolution of various acidic and basic compounds, leading to efficient analyte separation. When operated in the OT-CEC mode, separation selectivities could be readily manipulated via differential contributions from chromatographic and electrophoretic mechanisms, simply by changing the pH or the ionic strength of the running buffer electrolyte.
ABSTRACT
In this study, the surface charge properties of perfluorinated C8 (PerfluoroC8) and undecanoic acid (UDA) modified silica hydride stationary phases have been investigated. The zeta potential values of these stationary phases were measured in aqueous/acetonitrile mobile phases of different pH, buffer concentrations and acetonitrile contents. The retention behaviour of several basic, acidic and neutral compounds were then examined with these two stationary phases, with U-shaped retention dependencies evident with regard to the organic solvent content of the mobile phase. Plots of the logarithmic retention factor versus buffer concentration revealed slopes ≥ -0.41 for both stationary phases, indicating the involvement of mixed mode retention mechanisms with contributions from both ionic and non-ionic interactions. Using a linear solvation energy relationship approach, the origins of these interactions under different mobile phase conditions were differentiated and quantified. The PerfluoroC8 stationary phase exhibited stronger retention for basic compounds under high acetonitrile content mobile phase conditions, whilst stronger retention was observed for all compounds with the UDA stationary phase under high aqueous content mobile phase conditions. The more negative zeta potentials of the UDA stationary phase correlated with higher total charge density, surface charge density and charge density at the beta plane (the outer plane of the double layer) compared to the PerfluoroC8 stationary phase. With mobile phases of low buffer concentrations, more negative zeta potential values were unexpectedly observed for the PerfluoroC8 stationary phase with slight increases in the C descriptor value, reflecting also the greater accessibility of the analytes to the stationary phase surface. Comparison of the retention behaviours on these phases with other types of silica hydride stationary phases has revealed different patterns of selectivity.
Subject(s)
Fatty Acids/isolation & purification , Fluorocarbons/isolation & purification , Silicon Dioxide/chemistry , Fatty Acids/chemistry , Fluorocarbons/chemistryABSTRACT
Molecularly imprinted polymers (MIPs) templated with either the phytoalexin, (E)-resveratrol, or its structural analog, 3,5-dihydroxy-N-(4-hydroxyphenyl)benzamide, have been used in tandem for the sequential extraction of (E)-resveratrol from aqueous peanut meal extracts in high purity and in near quantitative yields. Re-processing of the (E)-resveratrol-depleted peanut meal extract with the 3,5-dihydroxy-N-(4-hydroxyphenyl)benzamide imprinted MIP yielded additional polyphenolic components, identified as A-type procyanidins. Tandem liquid chromatography-electrospray ionization mass spectrometry confirmed the identity and purity of the isolated products. This study documents the advantages of tandem approaches with MIPs for the solid phase extraction and analysis of multiple bioactive compounds present in complex biomass waste streams.
Subject(s)
Chemistry Techniques, Analytical/methods , Polyphenols/analysis , Polyphenols/isolation & purification , Solid Phase Extraction , Stilbenes/analysis , Stilbenes/isolation & purification , Arachis/chemistry , Chromatography, High Pressure Liquid , Industrial Waste/analysis , Molecular Imprinting , Resveratrol , Spectrometry, Mass, Electrospray Ionization , Water/chemistryABSTRACT
This investigation has examined the origin of the molecular recognition associated with the interaction of monoclonal IgG2's with terpyridine-based ligands immobilized onto agarose-derived chromatographic adsorbents. Isothermal titration calorimetric (ITC) methods have been employed to acquire thermodynamic data associated with the IgG2-ligand binding. These ITC investigations have documented that different enthalpic and entropic processes are involved depending on the nature of the chemical substituents in the core structure of the terpyridinyl moiety. In addition, molecular docking studies have been carried out with IgG2 structures with the objective to identify possible ligand binding sites and key interacting amino acid residues. These molecular docking experiments with the different terpyridine-based ligands have shown that all of the examined ligands can potentially undergo favorable interactions with a site located within the Fab region of the IgG2. However, another favorable binding site was also identified from the docking poses to exist within the Fc region of the IgG2 for some, but not all, of the ligands studied. These investigations have provided a basis to elucidate the unique binding properties and chromatographic behaviors shown by several substituted terpyridine ligands in their interaction with IgGs of different isotype. Copyright © 2016 John Wiley & Sons, Ltd.