Anion Exchange Interaction Research Articles

Identification and Characterization of Pectate Lyase as a Novel Allergen in Artemisia sieversiana Pollen

Introduction: Artemisia species are widely spread in north hemisphere. Artemisia sieversiana pollen is one of the common pollen allergens in the north of China. At present, seven allergens were identified and had been listed officially from A. sieversiana pollen, but the remaining allergens are still insufficiently studied, which need to be found. Methods: Pectate lyase was purified from the extracts of A. sieversiana pollen by anion exchange, size exclusion, and HPLC-hydrophobic interaction chromatography. The gene of A. sieversiana pectate lyase (Art si pectate lyase) was cloned and expressed in Escherichia coli. The enzyme activity and circular dichroism (CD) spectrum of natural and recombinant proteins were analyzed. The allergenicity of Art si pectate lyase was characterized by enzyme-linked immunosorbent assay (ELISA), Western blot, inhibition ELISA, and basophil activation test. The allergen’s physicochemical properties, three-dimensional structure, sequence profiles with homologous allergens and phylogenetic tree were analyzed by in silico methods. Results: Natural Art si pectate lyase (nArt si pectate lyase) was purified from A. sieversiana pollen extracts by three chromatographic strategies. The cDNA sequence of Art si pectate lyase had a 1191-bp open reading frame encoding 396 amino acids. Both natural and recombinant pectate lyase (rArt si pectate lyase) exhibited similar CD spectrum, and nArt si pectate lyase had higher enzymatic activity. Moreover, the specific immunoglobulin E (IgE) binding rate against nArt si pectate lyase and rArt si pectate lyase was determined as 40% (6/15) in patients’ serum with Artemisia species pollen allergy by ELISA. The nArt si pectate lyase and rArt si pectate lyase could inhibit 76.11% and 47.26% of IgE binding activities to the pollen extracts, respectively. Art si pectate lyase was also confirmed to activate patients’ basophils. Its structure contains a predominant motif of classic parallel helical core, consisting of three parallel β-sheets, and two highly conserved features (vWiDH, RxPxxR) which may contribute to pectate lyase activity. Moreover, Art si pectate lyase shared the highest sequence identity of 73.0% with Art v 6 among currently recognized pectate lyase allergen, both were clustered into the same branch in the phylogenetic tree. Conclusion: In this study, pectate lyase was identified and comprehensively characterized as a novel allergen in A. sieversiana pollen. The findings enriched the allergen information for this pollen and promoted the development of component-resolved diagnosis and molecular therapy of A. sieversiana pollen allergy.

An improved subtraction model applied in supercritical fluid chromatography to characterise polar stationary phases

Herein, an improved subtraction model was proposed to characterise the polar stationary phases in supercritical fluid chromatography (SFC). Fifteen stationary phases were selected, including two types of aromatic columns, Waters Torus and Viridis series columns, as well as silica and amino columns. Ethylbenzene and Torus 1-AA were defined as the reference solute and column, respectively. Identifying the interaction with the maximum contribution to retention in SFC separation and using it as the initial term is a key step in modelling. The dipole, or induced dipole interaction (θ′P), replaced the hydrophobic interaction (η′H) as the starting term. The improved model was expressed as logα=η′H+β′A+α′B+κ′C+θ′P+ε′E+σ′S, where the term ε′E indicated that anion exchange interaction was intentionally supplemented. A 7-step modelling process, including bidirectional fitting and residual analysis, was proposed. The obtained column parameters had reasonable physical significance, with the adjusted determination coefficient (R2adj) greater than 0.999 and the standard error (SE) less than 0.029. Methodological validation was further performed using the other four columns and 12 solutes that were not involved in the modelling. The result revealed good predictions of solutes’ retention, as demonstrated by R2adj from 0.9923 to 0.9979 and SE from 0.0636 to 0.1088. This study indicated the feasibility of using the improved subtraction model to characterise polar stationary phases in SFC, with the most crucial being the determination of an initial term, followed by the addition of a new descriptor and the selection of an appropriate reference column. The study expanded the application scope of the subtraction model in SFC, which will help gain an in-depth understanding of the SFC separation mechanism.

A new approach towards highly sensitive detection of endogenous N-acetylaspartic acid, N-acetylglutamic acid, and N-acetylaspartylglutamic acid in brain tissues based on strong anion exchange monolith microextraction coupled with UHPLC-MS/MS.

Anovel in-tube solid-phase microextraction coupled withan ultra-high performance liquid chromatography-mass spectrometry method has been established for simultaneous quantification of three crucial brain biomarkers N-acetylaspartic acid (NAA), N-acetylglutamic acid (NAG), and N-acetylaspartylglutamic acid (NAAG). A polymer monolith with quaternary ammonium as the functional group was designed and exhibited efficient enrichment of target analytes through strong anion exchange interaction. Under the optimized conditions, the proposed method displayed wide linear ranges (0.1-80 nM for NAA and NAG, 0.2-160 nM for NAAG) with good precision (RSDs were lower than 15%) and low limits of detection (0.019-0.052 nM), which is by far the most sensitive approach for NAA, NAG, and NAAG determination. Furthermore, this approach has been applied to measure the target analytes in mouse brain samples, and endogenous NAA, NAG, and NAAG were successfully detected and quantified from only around 5mg of cerebral cortex, cerebellum, and hippocampus. Compared with existing methods, the newly developed method in the current study provides highest sensitivity and lowest sample consumption for NAA, NAG, and NAAG measurements, which would potentially be utilized in determining and tracking these meaningful brain biomarkers in diseases or treatment processes, benefiting the investigations of pathophysiology and treatment of brain disorders.

Fabrication of polyethyleneimine grafted maleopimaric acid glycidyl methacrylate ester stationary phase for mixed-mode liquid chromatography

Due to their advantages of low green toxicity, diverse sources, renewable nature, and excellent biocompatibility, natural products have been extensively utilized in the exploration and development of novel stationary phase materials for liquid chromatography. In this study, we fabricated a rosin-based stationary phase material for mixed-mode chromatography by utilizing maleopimaric acid glycidyl methacrylate ester (MPAG) and polyethyleneimine (PEI) as ligands. Initially, silica was functionalized with MPAG through thiol-ene click reaction, followed by the grafting of PEI via an imidization reaction to produce Sil-MPAG-PEI material. This material exhibited both anion exchange and hydrophilic interaction properties due to the presence of PEI, allowing for the switching of its retention mode from reversed-phase mode to a mixed-mode incorporating reversed-phase, hydrophilic interaction and ion-exchange chromatography (RPLC/HILIC/IEC) under different mobile phase conditions. Consequently, Sil-MPAG-PEI demonstrated the ability to provide various interactions, including hydrophobic, π-π, hydrogen bond, hydrophilic, ion exchange, and electrostatic interactions. These interactions enabled selective separation among compounds with different polarities, such as nucleosides/bases, benzoic acids, flavonoids, alkylbenzenes, phenols, anilines, and polycyclic aromatic hydrocarbons (PAHs). Furthermore, Sil-MPAG-PEI demonstrated excellent chromatographic stability, as evidenced by the low relative standard deviations (RSDs) of the retention time for interday precision and interday precision were 0.042 %-0.275 % (n = 10) and 0.159 %-0.932 % (n = 7). The column was also successfully employed for the quantitative determination of evodiamine and rutaecarpine in an extract solution of Evodiae fructus, achieving concentrations of 9.102 and 3.256 mg/g, respectively. These results demonstrate the potential of Sil-MPAG-PEI in the analysis of complexsamples. Briefly, this study investigated and confirmed the potential application of rosin derivatives in mixed-mode liquid chromatography separations has been investigated and verified. The exceptional separation ability of Sil-MPAG-PEI indicates that it is a promising separation material for chromatographic separation.

Validation of a Liquid Chromatography Coupled to Mass Spectrometry Method for Glyphosate and Aminomethylphosphonic Acid in Urine for Human Biomonitoring Using Combined Hybrid Anion-Exchange and Hydrophilic Interaction Liquid Chromatography

Glyphosate-based herbicides are the most widely used pesticides in the world; however, the toxicity of glyphosate (GlyP) toward humans, especially its carcinogenicity, is controversial. The aim of this work was to validate a rapid assay for measuring GlyP and its metabolite aminomethylphosphonic acid (AMPA) in urine for human biomonitoring. The analytes were purified via solid-phase extraction in the presence of isotopically labeled internal standards. An LC-MS/MS assay was developed using a column with a novel hybrid stationary phase combined with anion exchange and hydrophilic interaction liquid chromatography. Detection and quantification were performed using negative electrospray ionization in a hybrid triple quadrupole/linear ion trap mass spectrometer. The retention times for AMPA and GlyP were 1.44 and 7.24 min, respectively. Calibration curves showed a linear dynamic range of up to 40 µg/L, inter- and intra-run precisions <7.5%, and accuracies within 10% of the theoretical concentrations. The limits of quantification were 0.1 µg/L and 0.5 µg/L for GlyP and AMPA, respectively. The matrix effect bias was controlled using internal standards. Successful participation in external quality assurance exercises strengthens the validity of the method. The assay was applied to the measurement of GlyP and AMPA in the urine of 9 urban residents, 26 rural residents, and 12 agricultural workers; while AMPA was mostly not quantifiable, the median GlyP values were 0.1 and 0.34 µg/L in rural residents and workers, respectively. The assay is useful to assess GlyP and AMPA in human urine following different exposure scenarios.

Dendrimer functionalized spinel ferrite for robust sorption of precious metals from strongly acidic medium

The separation of noble metal ions from strongly acidic solutions is considered to be a challenging task due to the significantly low pH of these solutions. Spinel ferrites possess unique physicochemical properties, such as excellent magnetic properties, high specific surface area, high chemical stability, active surface sites, and easy modification or functionalization, making them suitable for various applications. In this study, we have introduced a facile method for synthesizing polyamidoamine (PAMAM) dendrimer-functionalized spinel ferrite (DFSF) with a particle size of approximately 10 nm. The prepared DFSF materials have investigated its ability to adsorb Pt(IV) and Pd(II) anions (PtCl62− and PdCl42−) from strongly acidic solutions. The results indicate that the adsorption kinetics and isotherm studies of synthesized materials have a rapid and high adsorption capacity for Pt(IV) and Pd(II) with simple solid-liquid separation. The data analysis suggests an anion exchange interaction between the adsorbate and the adsorbent, specifically the electrostatic interaction between the protonated amine group (–NH3+) and the noble metal anions (PtCl62–and PdCl42−). The Pseudo second-order rate model fitting suggests that the adsorption of noble metals by DFSF is a function of valence. The excellent superparamagnetic properties (0.346 emu g−1) of DFSF make it a promising candidate for large-scale applications, as it exhibited good reusability with a short recovery time (25 s) and high recovery efficiency through the adsorption-reduction-precipitation route. This study suggests that DFSF can be a potential sorbent for noble metal ions in various applications.

Maximal performance of intact N-glycopeptide enrichment using sequential HILIC and MAX columns.

Low abundance and heterogeneity of N-glycosylation at the peptide level poses a great challenge to the structural and functional analysis of glycosylation in the field of glycobiology. Solving this conundrum requires a sufficient and specific method for intact N-glycopeptide enrichment. Using the C18 or HLB desalting column followed by the mixed-mode strong anion exchange (MAX) or hydrophilic interaction chromatography (HILIC) glycopeptide enrichment column are commonly applied approaches for sample preparation of intact N-glycopeptides from complex samples. Herein, we compared the effects of different combinations of two desalting columns and two enrichment columns using equal amounts of mouse brain tissues from the same source. The results revealed the C18 column was a bit superior to the HLB column, and the MAX and HILIC columns were complementary on intact N-glycopeptides enrichment. Additionally, the results also demonstrated that enriching glycopeptides using a HILIC column followed by a MAX column from the flow-through solution got a better enrichment performance than the reversed order. Based on these results, the sequential enrichment of glycopeptides using HILIC and then MAX columns could maximize the enrichment performance of intact N-glycopeptides, and therefore is an option for in-depth analysis of site-specific glycoproteome.

In situ self-sacrificial synthesis of polypyrrole/biochar composites for efficiently removing short- and long-chain perfluoroalkyl acid from contaminated water

Efficient removal of perfluoroalkyl acids (PFAAs), especially short-chain ones, from contaminated water is of great challenge and is urgently called for so as to safeguard the ecosystem and human health. Herein, polypyrrole (PPy) functionalized biochar (BC) composites were innovatively synthesized by an in situ self-sacrificial approach to allow efficient capture of PFAAs with different chain lengths. Compared with conventional PPy-based composites synthesized by direct polymerization using FeCl3 as an oxidizing agent, PPy/BC composites were fabricated utilizing freshly generated Fe3+ as an oxidizing agent from self-sacrificial Fe3O4 for pyrrole monomers in situ polymerizing on BC. As a result, with the support of BC and gradual release of Fe3+, PPy overcame its tendency to aggregate and became uniformly dispersed on BC, and meanwhile, PPy could well tailor the surface chemistry of BC to endow its positively charged surface. Consequently, the composites exhibited strong sorption capacities of 3.89 and 1.53 mmol/g for short-chain perfluorobutanoic acid (PFBA) and perfluorobutane sulfonic acid (PFBS), 2.55 and 1.22 mmol/g for long-chain perfluorooctanoic acid (PFOA) and perfluorooctane sulfonic acid (PFOS), respectively, which were superior to those of pristine BC, commercial activated carbon, and anion exchange resins reported. Additionally, they could effectively remove 17 different classes of per- and polyfluoroalkyl substances (PFAS) (removal >95%) from actual PFAS-contaminated water, and the spent sorbent could be well regenerated and reused at least 5 times. An integrated analysis indicated that such an outstanding PFAA sorption performance on PPy/BC composites could be mainly attributed to surface adsorption enhanced by electrostatic attractions (anion exchange interaction) with the traditional hydrophobic interaction and pore filling of less contribution, particularly for short-chain analogues. These results are expected to inform the design of BC with greater ability to remove PFAS from water and the new sorbent could help water facilities comply with PFAS regulations.

Evaluating orthogonality between ion-pair reversed phase, anion exchange, and hydrophilic interaction liquid chromatography for the separation of synthetic oligonucleotides

The orthogonality of separation between ion-pair reversed phase (IP-RP), anion exchange (AEX), and hydrophilic interaction liquid chromatography (HILIC) was evaluated for oligonucleotides. A polythymidine standard ladder was first used to evaluate the three methods and showed zero orthogonality, where retention and selectivity were based on oligonucleotide charge/size under all three conditions. Next, a model 23-mer synthetic oligonucleotide containing 4 phosphorothioate bonds with 2′ fluoro and 2′-O-methyl ribose modifications typical of small interfering RNA was used for evaluating orthogonality. The resolution and orthogonality were evaluated between the three modes of chromatography in terms of selectivity differences for nine common impurities, including truncations (n-1, n-2), addition (n + 1), oxidation, and de-fluorination. We first evaluated different ion-pairing reagents that provided the best separation of the key impurities while suppressing diastereomer separation due to phosphorothioate linkages. Although different ion-pairing reagents affected resolution, very little orthogonality was observed. We then compared the retention times between IP-RP, HILIC, and AEX for each impurity of the model oligonucleotide and observed various selectivity changes. The results suggest that coupling HILIC with either AEX or IP-RP provide the highest degree of orthogonality due to the differences in retention for hydrophilic nucleobases and modifications under HILIC conditions. IP-RP provided the highest overall resolution for the impurity mixture, whereas more co-elution was observed with HILIC and AEX. The unique selectivity patterns offered by HILIC provides an interesting alternative to IP-RP or AEX, in addition to the potential for coupling with multidimensional separations. Future work should explore orthogonality for oligonucleotides with subtle sequence differences such as nucleobase modifications and base flip isomers, longer strands such as guide RNA and messenger RNA, and other biotherapeutic modalities such as peptides, antibodies, and antibody-drug-conjugates.

Speciation of inorganic arsenic with mixed mode HPLC-ESI-MS and Arsenite Oxidation

It has been challenging to analyze inorganic arsenic (iAs) with anion exchange HPLC-Electrospray Ionization-Mass spectrometry (HPLC-ESI-MS), because arsenite (As(III)) is difficult to retain on column and the salts in mobile phase causes ionization suppression of iAs. To address these issues, a method has been developed involving the determination of arsenate (As(V)) with mixed mode HPLC-ESI-MS and the conversion of As(III) to As(V) for total iAs. As(V) was separated from other chemicals on Newcrom B, a bi-modal HPLC column involving anion exchange and reverse phase interaction. The elution employed a two-dimensional gradient, including a formic acid gradient to elute As(V) and a concurrent alcohol gradient to elute organic anions used in sample preparations. As(V) was detected by Selected Ion Recording (SIR) in negative mode at m/z = 141 with a QDa (single quad) detector. As(III) was quantitatively converted to As(V) by mCPBA oxidation and measured for total iAs. By replacing salt with formic acid in elution, the ionization efficiency for As(V) was greatly enhanced in ESI interface. The limit of detection (LOD) for As(V) and As(III) were 0.0263 μM (1.97 ppb) and 0.0398 μM (2.99 ppb), respectively. The linear range was 0.05–1 μM.The method has been used to characterize iAs speciation change in the solution and precipitation in a simulated iron-rich groundwater caused by air exposure.

Enhance selective separation of vanadium and iron from acid-leaching solution with sulfamic acid modified resin

To address the problems of low saturated adsorption capacity of the chelate resin and enhance selective separation of vanadium and iron when vanadium shale acid leaching solution was treated by ion exchange resin, DS418 resin, an innovative resin was prepared by surface modification of D418 resin with sulfamic acid. FTIR, XPS and the frontier molecular orbital theory show that the amino and the sulfamic acid are under a sulfonamide reaction, removing a molecule of H2O and connecting S and N. The adsorption rate of V on DS418 resin was 94.17 % when the concentration of modifier was 1.35 mol/L, the modified liquid–solid ratio was 6.5 g/mL and the modification temperature was 41℃. When pH = 1.8, the adsorption rate of V was increased by 12 %, the adsorption rate of Fe was reduced by 34 % compared with the D418 resin, and the saturated adsorption capacity of DS418 resin reached 112.36 mg/g, was twice as much as the D418 resin. After 10 cycles of adsorption–desorption experiments, the adsorption rate of DS418 resin for V was 92.19 %, and the desorption rate was 98.37 %. The adsorption mechanism of V was changed from single anion exchange to synergistic adsorption of anion exchange and electrostatic interaction. DS418 resin had a higher affinity for V than Fe. This research is expected to achieve efficient recovery of V and guide the selection and design of special resin for V extraction.

Sequentially activating macrophages M1 and M2 phenotypes by lipopolysaccharide-containing Mg-Fe layered double hydroxides coating on the Ti substrate

As cells of innate immunity, macrophages are a class of innate immune cells existing in almost all tissues and play a crucial role in bone repair. However, it remains a challenge to modulate the sequential activation of the deferent phenotypes in macrophage when designing the titanium (Ti) implants. In this study, the Mg-Fe layered double hydroxides (LDHs) was coated on Ti substrate through hydrothermal treatment. Further on lipopolysaccharide (LPS) was introduced onto the LDHs through adsorption and ions exchange. The adsorption efficiency of the coating on LPS reached 72.8% in 24 h due to the anion exchange and electrostatic interactions between the LPS and the LDH layers in deionized water. The LDHs-LPS coating released a large amount of LPS in the early stage, which induced macrophages into M1 phenotype via activating TLR-4 → MyD88 and TLR-4 → Ticam-1/2 signal pathways. Subsequently, the M1 macrophages were transformed into M2 phenotype by regulating the integrin α5β1 of cells by the nanostructures, wetting angle and Mg2+ of the coating. The LDHs-LPS coating endows Ti with the ability of stage immunomodulation, indicating the positive osteoimmunomodulatory property.

Selective and efficient removal of ReO4- from aqueous solution by imidazolium-based porous organic polymers

In this paper, imidazolium-functionalized porous organic polymers (IM-Br-2OH) are prepared successfully via a quaternization reaction of tetrakis[4-(1-imidazolyl)phenyl]methane (TIPM) and 1,4-dibromo-2,3-butanediol (2,3-BDO). The as-synthesized IM-Br-2OH are characterized by various tools such as Fourier transform infrared spectroscopy (FTIR), Scanning electron microscope and energy dispersive spectrum (SEM-EDS mapping), Thermogravimetric analysis (TGA), N2 adsorption-desorption isotherms, X-ray photoelectron spectroscopy (XPS) and solid-state 13C CP/MAS NMR spectroscopy. The IM-Br-2OH exhibits the maximum adsorption capacity of perrhenate (ReO4-) was 658.4 mg/g with removal efficiency of 98.0 ± 0.2% at 35 ºC. Experimental results showed that the IM-Br-2OH presents a higher adsorption capacity for ReO4- than the previously reported adsorbents. In addition, excellent adsorption selectivity of IM-Br-2OH for ReO4- was observed in the presence of competing anions such as CO32-, Cl-, SO42-, NO3-, and PO43-. Adsorption process of ReO4- from solutions onto IM-Br-2OH follows a pseudo-second-order mode well and the adsorption data are in accordance well with the Langmuir isotherm equations. Adsorption mechanism is confirmed with the help of Zeta potentials, FTIR and XPS, indicating that anion-exchange interactions between N+ and ReO4- contribute to the ReO4- adsorption from aqueous solution. The conclusions imply that IM-Br-2OH has potential application in the removal of radioactive pollutants from effluents.

Anion Exchange Affinity-Based Controllable Surface Imprinting Synthesis of Ultrathin Imprinted Films for Protein Recognition.

Anion exchange affinity-based controllable surface imprinting is an effective approach to overcome low imprinting efficiency and high non-specific binding capacity. The template proteins were first immobilized on the anchored tetraalkylammonium groups of the nanoparticles via anion exchange affinity-based interactions, enabling monolayer sorption using a low template concentration. The combined use of surface-initiated photoiniferter-mediated polymerization to precisely control the imprinted film thickness, allowing the formation of homogeneous binding cavities, and the construction of effective binding sites resulted in a low non-specific binding capacity and high imprinting efficiency. The obtained imprinted films benefited from the anion exchange mechanism, exhibiting a higher imprinting factor and faster binding rate than the reference material. Binding tests revealed that the binding strength and selective recognition properties could be tuned to a certain extent by adjusting the NaCl concentration. Additionally, in contrast to the harsh template elution conditions of the covalent immobilization approach, over 80% of the template molecules were readily removed from the imprinted films using supersonic elution with an aqueous mixture of NaCl and HAc. Introducing template immobilization by anion exchange interactions to the synthesis of imprinted materials may provide a new approach for effective biomacromolecular imprinting.

Enhanced removal of Eriochrome Black T from water using biochar/layered double hydroxide/chitosan hybrid composite: Performance evaluation and optimization using BBD-RSM approach

In this research work, a novel hybrid composite consisting of biochar (B), layered double hydroxide (CuFe) and chitosan (CS) (B-CuFe-CS) was produced using an ultrasonication-assisted co-precipitation method. The resultant composite was employed for adsorptive removal of Eriochrome black T (EBT) from water. Physicochemical characterization indicated that the B-CuFe-CS containing 10 wt % CS exhibited a heterogeneous structure with better crystallographic and textural characteristics. The B-CuFe-CS with abundant surface functionalities (-CO, -C-O, –OH, –NO3, and MMO), facilitates faster and enhanced removal of the EBT. The kinetic results showed better fitting to the pseudo-second order model, and equilibrium was achieved within 30 min. Equilibrium data was well explained by Langmuir and Redlich Peterson isotherm models (R2 > 0.98), indicating the EBT removal onto B-CuFe-CS followed monolayer adsorption. The maximum adsorption capacity was 806.4 mg/g, which was higher than pristine B–CuFe (476.19 mg/g) and many other adsorbents. The spectroscopic analysis (FTIR and XPS) and experimental results suggested that EBT adsorption is mainly governed by electrostatic, chemical and anion-exchange interactions. It is evident from these results that coupling B–CuFe composite with bio-filler (chitosan) resulted in an efficient bio-adsorbent to effectively purify dye-contaminated water streams.

Comparing the relative robustness of anion exchange and the corresponding mixed-mode chromatography on removing a weakly-bound byproduct: A case study of bispecific antibody purification

A certain impurity may bind weaker or tighter to a particular type of chromatography column (e.g., ion exchange, hydrophobic interaction or mixed-mode) than the target protein, and this forms the basis for separation. For impurities that bind weaker, they can be removed by an appropriate pre-elution wash. However, we previously showed that wash-enabled impurity clearance is usually sensitive to loading density, causing poor robustness of the process. In this work, with a bispecific antibody case study, we compared the relative robustness of anion exchange and the corresponding mixed-mode chromatography, which mediates both anion exchange and hydrophobic interactions, on removing a weakly-bound byproduct by wash. It was learned that under a fixed appropriate wash condition, the latter achieves consistent byproduct clearance and good yield over a much wider range of loading density than the former. As wide loading density range is highly desirable for large-scale manufacturing, the above finding suggests that for a chromatography step that employs stepwise elution and relies on pre-elution wash for removing weakly-bound impurities, mixed-mode chromatography could be a better choice than the corresponding monomode ion exchange chromatography.

Field sample preparation of trace inorganic anions in environmental waters with in-tip microextraction device based on anion-exchange monolithic adsorbent followed by ion chromatography quantification

In this study, a new field preparation method for the quantification of inorganic anions (including F−, BrO3−, Cl−, NO2− and Br−) at trace levels in environmental waters was proposed and developed. Glycidyl methacrylate was copolymerized with ethylene dimethacrylate in a pipette tip to obtain epoxy group-rich monolith, and then trimethylamine was employed to perform ring-opening reaction and got anion-exchange monolithic adsorbent (AMA). The resultant adsorbent was used as the extraction medium of three-channel in-tip microextraction device (TCMD) which was employed to conduct field sample preparation of inorganic anions in different water samples followed by analysis with ion chromatography with conductivity detector (IC/CD). The AMA exhibited satisfactory capture capability to studied anions through anion-exchange interaction. After optimization of extraction parameters, a method with low limits of detection (LODs, 0.019–0.33 μg/L), satisfactory coefficients of correlation (0.9925–0.9985) and good precision (RSDs ≤ 10%) was developed. The introduced approach was successful applied to detect studied anions in environmental waters, and the fortified recoveries varied from 77.3 to 113%. Additionally, the accuracy of suggested method was inspected by confirmatory testing. Compared with reported methods, the current approach displays some attractive features such as semi-automation, high throughput and low LODs. The suggested AMA/TCMD for on-site preparation of inorganic anions not only circumvents the storage and transportation of a great quantity of waters, but also ensures the quantification accuracy.

Biochemical Characterization of Two Polygalacturonases Purified from the Digestive Juice of the Snail Limicolaria flammea

Polygalacturonases constitute the major part of pectinase preparations for many bioprocess purposes. Investigation on the digestive juice of snail Limicolaria flammea led to purification of two polygalacturonases named PG1 and PG2. Properties of these enzymes were examined to explore their potential in biotechnology applications. A three steps procedure including size exclusion, anion and cation exchange and hydrophobic interaction chromatography were used for purification. The enzymes PG1 and PG2 had native molecular weights of approximately 46 and 86 kDa, respectively and functioned both as monomeric structures. The purified polygalacturonases PG1 and PG2 showed optimum hydrolysis activities at 50°C in sodium acetate buffer pH 5.6. The common inhibitor of the two purified polygalacturonases activity were Mn2+, Ca2+, Zn2+, EDTA, SDS and L-cystein. NH3+ stimulate the polygalacturonase PG1 while Ba2+ was an activator for polygalacturonase PG2. Substrate specificity indicated that these enzymes hydrolyse a broad range of pectin from different sources. The highest activity of PG1 was observed with apple pectin and lemon pectin while PG2 showed its highest activity with orange pectin. The catalytic efficiency of PG1 was highest for lemon pectin (0.125 µmol/min/mL) and orange pectin (0.124 µmol/min/mL). PG2 displayed highest catalytic efficiency (0.325 µmol/min/mL) towards orange pectin. These results suggest that orange and lemon pectin would be the potential physiological substrates of the two purified enzymes.

Identification of allergens for food-dependent exercise-induced anaphylaxis to shrimp

Shrimp is a causative food that elicits food-dependent exercise-induced anaphylaxis (FDEIA). In this study, we sought to identify IgE-binding allergens in patients with shrimp-FDEIA. Sera were obtained from eight patients with shrimp-FDEIA and two healthy control subjects. Proteins were extracted from four shrimp species by homogenization in Tris buffer. Immunoblot analysis revealed that IgE from patient sera bound strongly to a 70-kDa and a 43-kDa protein in a preparation of Tris-soluble extracts from Litopenaeus vannamei. Mass spectrometry identified the 70-kDa and 43-kDa proteins as a P75 homologue and fructose 1,6-bisphosphate aldolase (FBPA), respectively. To confirm that the putative shrimp allergens were specifically recognized by serum IgE from shrimp-FDEIA patients, the two proteins were purified by ammonium sulfate precipitation followed by reversed-phase HPLC and/or anion-exchange hydrophobic interaction chromatography and then subjected to immunoblot analysis. Purified P75 homologue and FBPA were positively bound by serum IgE from one and three, respectively, of the eight patients with shrimp-FDEIA, but not by sera from control subjects. Thus, P75 homologue and FBPA are identified as IgE-binding allergens for shrimp-FDEIA. These findings could be useful for the development of diagnostic tools and desensitization therapy for shrimp-FDEIA patients.

New deep eutectic solvent based superparamagnetic nanofluid for determination of perfluoroalkyl substances in edible oils

Perfluoroalkyl substances (PFASs) have been identified as global pollutants and raise considerable food safety concerns. However, the development of an analytical method with satisfied pretreatment performance for PFASs with varying alkyl chain length in the fatty samples remains a challenge. We describe herein the preparation of superparamagnetic nanofluid, based on a new choline chloride/1-(o-tolyl)biguanide deep eutectic solvent (DES) system, for direct extraction of perfluoroalkyl carboxylic acids from edible oils. Target PFASs, especially the short-chain one, all possessed high recoveries (90–109% with intra-day and inter-day precision below 10%). This was achieved by adjusting the constituent ratio of DES for balancing the anion-exchange (electrostatic) interactions and hydrogen-bond interactions. Employing the prepared nanofluid in magnetic microextraction, followed by high-resolution mass spectrum analysis, resulted in a rapid (15 min for pretreatment), simple, sensitive (detection limit: 0.3–1.6 pg g−1), and efficient method for the enrichment and determination of trace PFASs. Furthermore, the introducing of N–H⋯F weak force increased the pretreatment selectivity to effectively reduce the matrix effect. At the end of the study, the proposed methodology was successfully applied to the analysis of target analytes in real samples.