Deep eutectic solvent assisted recovery of high molecular weight levan from an isolated Neobacillus pocheonensis BPSCM4.

The present study demonstrates the usage of deep eutectic solvent to recover microbial levan from the clarified fermented broth. The classic ethanol precipitation method for levan recovery is expensive because ethanol can be utilized as a biofuel. Production of ethanol consumes more energy and is not easily recycled. As a result, the current work concentrates on using environmentally friendly solvents for levan recovery. Deep Eutectic Solvents (DES) are greener and can replace ethanol from the microbial polysaccharides precipitation. Thus the proposed approach is environment friendly, technically feasible, reliable and economically viable. The levan was produced from a microbial isolate of aged sugarcane molasses, recovered using traditional ethanol and proposed DES (Choline Chloride and Ethylene Glycol) assisted precipitation. The levan-producing strain was characterized and identified as Neobacillus pocheonensis BPSCM4. The DES-precipitated levan has a high molecular weight of levan, 1.54 × 106 KDa, compared with the ethanol-precipitated levan, 4.246 KDa. The high molecular weight of DES-precipitated levan is due to the low viscosity and hydrogen interaction of ChCl:EG with the levan present in the fermented broth. Further, the optimization enhanced the levan yield to 32.56 g/L when the sucrose concentration was 250 g/L.

C-reactive protein interacts with amphotericin B liposomes and its potential clinical consequences.

OBJECTIVES: Amphotericin B (AmB) is the gold standard for treating invasive fungal infections. New liposomal-containing AmB formulations have been developed to improve efficacy and tolerability. Serum/plasma C-reactive protein (CRP) values are widely used for monitoring infections and inflammation. CRP shows a high affinity to phosphocholine and it aggregates structures bearing this ligand, e.g. phosphocholine-containing liposomes. Therefore, we studied the interaction between CRP and phosphocholine-containing liposomal AmB preparations in vivo and in vitro. METHODS: CRP was prepared by affinity chromatography. Liposomal AmB (L-AmB, AmBisome®) was spiked (final concentrations of L-AmB: 150 mg/L) to CRP-containing serum (final CRP concentration: 300 mg/L). Following the addition of L-AmB, complex formation was monitored turbidimetrically. The size of CRP-L-AmB complexes was assessed using gel filtration. CRP was monitored in patients receiving either L-Amb or AmB lipid complex (ABLC). RESULTS: Following addition of L-AmB to CRP-containing plasma, turbidimetry showed an increase in absorbance. These results were confirmed by gel permeation chromatography. Similarly, in vivo effects were observed following intravenous administration of AmBisome®: a decline in CRP values was observed. In patients receiving L-Amb, decline of CRP concentration was faster than in patients receiving ABLC. CONCLUSIONS: In vitro experiments are suggestive of a complexation between CRP and liposomes in plasma. Interpretation of CRP values following administration of AmBisome® might be impaired due to this complexation. In vivo formation of complexes between liposomes and CRP might contribute, or even lead, to intravascular microembolisation. Similar effects have been described following the administration of Intralipid® and other phosphocholine-containing liposomes.

High-Molecular-Weight Exopolysaccharides Production from Tuber brochii Cultivated by Submerged Fermentation.

Truffles are known worldwide for their peculiar taste, aroma, and nutritious properties, which increase their economic value. However, due to the challenges associated with the natural cultivation of truffles, including cost and time, submerged fermentation has turned out to be a potential alternative. Therefore, in the current study, the cultivation of Tuber borchii in submerged fermentation was executed to enhance the production of mycelial biomass, exopolysaccharides (EPSs), and intracellular polysaccharides (IPSs). The mycelial growth and EPS and IPS production was greatly impacted by the choice and concentration of the screened carbon and nitrogen sources. The results showed that sucrose (80 g/L) and yeast extract (20 g/L) yielded maximum mycelial biomass (5.38 ± 0.01 g/L), EPS (0.70 ± 0.02 g/L), and IPS (1.76 ± 0.01 g/L). The time course analysis of truffle growth revealed that the highest growth and EPS and IPS production was observed on the 28th day of the submerged fermentation. Molecular weight analysis performed by the gel permeation chromatography method revealed a high proportion of high-molecular-weight EPS when 20 g/L yeast extract was used as media and the NaOH extraction step was carried out. Moreover, structural analysis of the EPS using Fourier-transform infrared spectroscopy (FTIR) confirmed that the EPS was ß-(1-3)-glucan, which is known for its biomedical properties, including anti-cancer and anti-microbial activities. To the best of our knowledge, this study represents the first FTIR analysis for the structural characterization of ß-(1-3)-glucan (EPS) produced from Tuber borchii grown in submerged fermentation.

Molecular Characteristics and Antioxidant Activity of Spruce (Picea abies) Hemicelluloses Isolated by Catalytic Oxidative Delignification.

Spruce (Piceaabies) wood hemicelluloses have been obtained by the noncatalytic and catalytic oxidative delignification in the acetic acid-water-hydrogen peroxide medium in a processing time of 3-4 h and temperatures of 90-100 °C. In the catalytic process, the H2SO4, MnSO4, TiO2, and (NH4)6Mo7O24 catalysts have been used. A polysaccharide yield of up to 11.7 wt% has been found. The hemicellulose composition and structure have been studied by a complex of physicochemical methods, including gas and gel permeation chromatography, Fourier-transform infrared spectroscopy, and thermogravimetric analysis. The galactose:mannose:glucose:arabinose:xylose monomeric units in a ratio of 5:3:2:1:1 have been identified in the hemicelluloses by gas chromatography. Using gel permeation chromatography, the weight average molar mass Mw of hemicelluloses has been found to attain 47,654 g/mol in noncatalytic delignification and up to 42,793 g/mol in catalytic delignification. Based on the same technique, a method for determining the α and k parameters of the Mark-Kuhn-Houwink equation for hemicelluloses has been developed; it has been established that these parameters change between 0.33-1.01 and 1.57-472.17, respectively, depending on the catalyst concentration and process temperature and time. Moreover, the FTIR spectra of the hemicellulose samples contain all the bands characteristic of heteropolysaccharides, specifically, 1069 cm-1 (C-O-C and C-O-H), 1738 cm-1 (ester C=O), 1375 cm-1 (-C-CH3), 1243 cm-1 (-C-O-), etc. It has been determined by the thermogravimetric analysis that the hemicelluloses isolated from spruce wood are resistant to heating to temperatures of up to ~100 °C and, upon further heating, start destructing at an increasing rate. The antioxidant activity of the hemicelluloses has been examined using the compounds simulating the 2,2-diphenyl-2-picrylhydrazyl free radicals.

An Efficient Method of Birch Ethanol Lignin Sulfation with a Sulfaic Acid-Urea Mixture.

For the first time, the process of birch ethanol lignin sulfation with a sulfamic acid-urea mixture in a 1,4-dioxane medium was optimized experimentally and numerically. The high yield of the sulfated ethanol lignin (more than 96%) and containing 7.1 and 7.9 wt % of sulfur was produced at process temperatures of 80 and 90 °C for 3 h. The sample with the highest sulfur content (8.1 wt %) was obtained at a temperature of 100 °C for 2 h. The structure and molecular weight distribution of the sulfated birch ethanol lignin was established by FTIR, 2D 1H and 13C NMR spectroscopy, and gel permeation chromatography. The introduction of sulfate groups into the lignin structure was confirmed by FTIR by the appearance of absorption bands characteristic of the vibrations of sulfate group bonds. According to 2D NMR spectroscopy data, both the alcohol and phenolic hydroxyl groups of the ethanol lignin were subjected to sulfation. The sulfated birch ethanol lignin with a weight average molecular weight of 7.6 kDa and a polydispersity index of 1.81 was obtained under the optimum process conditions. Differences in the structure of the phenylpropane units of birch ethanol lignin (syringyl-type predominates) and abies ethanol lignin (guaiacyl-type predominates) was manifested in the fact that the sulfation of the former proceeds more completely at moderate temperatures than the latter. In contrast to sulfated abies ethanol lignin, the sulfated birch ethanol lignin had a bimodal and wider molecular weight distribution, as well as less thermal stability. The introduction of sulfate groups into ethanol lignin reduced its thermal stability.

Development of a gel permeation chromatography method for analysing cellulose nitrate in museums.

This article describes the development of a suitable Gel Permeation Chromatography method for cellulose nitrate plasticised with camphor (celluloid) found in cultural heritage. Current sample preparation and dissolution methods, apart from focusing on native, nonderivatised cellulose, require long preparation times, and often employ solvents that induce degradation. This study aims to develop a systematic method for sample preparation of cellulose nitrate that uses the least sample amount possible, is nondegrading, and can be applied on differently aged samples. This is investigated through identification of a suitable solvent system and a statistically designed experiment testing the critical variables affecting the analysis, namely sample condition, sample, and salt concentration (lithium chloride) in N,N-dimethylacetamide. The use of 0.1% sample was inadequate for analysis because it did not fully dissolve in any salt concentration, while the 0.3% negatively impacted the analysis with its high molecular weight distributions. The 0.2% cellulose nitrate in a solution of 0.5% lithium chloride in N,N-dimethylacetamide offered the most consistent and repeatable molecular weight data. This method miniaturised the sample as much as possible and is suitable for museum objects in various ageing conditions.

Modification of Arabinogalactan Isolated from Larix sibirica Ledeb. into Sulfated Derivatives with the Controlled Molecular Weights.

The process of sulfation of arabinogalactan-a natural polysaccharide from Larix sibirica Ledeb.-with sulfamic acid in 1,4-dioxane using different activators has been studied for the first time. The dynamics of the molecular weight of sulfated arabinogalactan upon variation in the temperature and time of sulfation of arabinogalactan with sulfamic acid in 1,4-dioxane has been investigated. It has been found that, as the sulfation time increases from 10 to 90 min, the molecular weights of the reaction products grow due to the introduction of sulfate groups without significant destruction of the initial polymer and sulfation products. Sulfation at 95 °C for 20 min yields the products with a higher molecular weight than in the case of sulfation at 85 °C, which is related to an increase in the sulfation rate; however, during the further process occurring under these conditions, sulfation is accompanied by the destruction and the molecular weight of the sulfated polymer decreases. The numerical optimization of arabinogalactan sulfation process has been performed. It has been shown that the optimal parameters for obtaining a product with a high sulfur content are a sulfamic acid amount of 20 mmol per 1 g of arabinogalactan, a process temperature of 85 °C, and a process time of 2.5 h.

Poly(Pyridinium Salt)s Containing 2,7-Diamino-9,9'-Dioctylfluorene Moieties with Various Organic Counterions Exhibiting Both Lyotropic Liquid-Crystalline and Light-Emitting Properties.

A series of poly(pyridinium salt)s-fluorene main-chain ionic polymers with various organic counterions were synthesized by using ring-transmutation polymerization and metathesis reactions. Their chemical structures were characterized by Fourier Transform Infrared (FTIR), proton (1H), and fluorine 19 (19F) nuclear magnetic resonance (NMR) spectrometers. These polymers showed a number-average molecular weight (Mns) between 96.5 and 107.8 kg/mol and polydispersity index (PDI) in the range of 1.12-1.88. They exhibited fully-grown lyotropic phases in polar protic and aprotic solvents at different critical concentrations. Small-angle X-ray scattering for one polymer example indicates lyotropic structure formation for 60-80% solvent fraction. A lyotropic smectic phase contains 10 nm polymer platelets connected by tie molecules. The structure also incorporates a square packing motif within platelets. Thermal properties of polymers were affected by the size of counterions as determined by differential scanning calorimetry and thermogravimetric analysis measurements. Their ultraviolet-visible (UV-Vis) absorption spectra in different organic solvents were essentially identical, indicating that the closely spaced π-π* transitions occurred in their conjugated polymer structures. In contrast, the emission spectra of polymers exhibited a positive solvatochromism on changing the polarity of solvents. They emitted green lights in both polar and nonpolar organic solvents and showed blue light in the film-states, but their λem peaks were dependent on the size of the counterions. They formed aggregates in polar aprotic and protic solvents with the addition of water (v/v, 0-90%), and their λem peaks were blue shifted.

Mechanistic Study of the Solvent-Dependent Formation of Extended and Stacked Supramolecular Polymers Composed of Bis(imidazolylporphyrinatozinc) Molecules.

Bis(imidazolylporphyrinatozinc) molecules linked through a 1,3-butadiynylene moiety respond to the solvents they are dissolved in to afford exclusively extended (E) or stacked (S) supramolecular polymers. This system is expected to be a solvation/desolvation indicator. However, the principles underlying the solvent-dependent formation of the two types of polymers and the mechanism of the transformation between them are unclear. The formation of the polymers is considered to depend on the two types of complementary coordination bonds that can be formed and the π-π interactions between the porphyrins. In this study, the contributions and solvent dependence of both the coordination bonds and the π-π interactions have been investigated. The results clearly indicate that the coordination bonds are weakly or little solvent-dependent, and that the π-π interactions function effectively only in the inner porphyrins of the S-polymer and are strongly solvent-dependent. Thermodynamic analysis revealed that the formation of the E- or S-polymer in solution is determined by the total energies and the type of solvent used. The transformation of the E- to S-polymer was investigated by gel permeation chromatography. The kinetics of the transformation were also determined. The role of the terminal imidazolylporphyrinatozinc moieties was also investigated: The results indicate that the transformation from the E- to S-polymer occurs by an exchange mechanism between the polymers, induced by attack of terminal free imidazolyl groups on a polymer to zinc porphyrins on other polymers.

Salinomycin-loaded PLA nanoparticles: drug quantification by GPC and wave voltammetry and biological studies on osteosarcoma cancer stem cells.

A new straightforward gel permeation chromatography (GPC) method was developed to calculate the drug encapsulation efficiency and loading content of Poly(lactic acid) nanoparticles (PLA NPs) loaded with Salinomycin (Sal), exploiting the capability of this technique to separate a macromolecular/molecular mixture on the basis of the molecular weight of each component. The proposed GPC method allowed Sal detection until 1% of Sal content in PLA NPs, avoiding sample pre-treatments. The method was validated by wave voltammetry (SW) technique, using a slightly modified literature procedure, useful to detect Sal in the concentration range 0.4 ≤ C/µmol/L ≤ 12 (linear concentration range). PLA-based NPs were prepared by nanoprecipitation with either native and functionalized PLA. Specifically, folate-decorated PLA NPs (PLA-FA NPs) were obtained by CuAAC click functionalization of alkyne-grafted PLA with azide-folate. Sal-loaded NPs were characterized physicochemically and morphologically. They exhibited adequate physicochemical properties, good drug encapsulation efficiency (98 ± 0.5% and 99 ± 0.5%), and loading content (8.8 ± 0.1% and 8.9 ± 0.1% for PLA/Sal and PLA-FA/Sal NPs, respectively). The size of empty PLA NPs resulted smaller (90 ± 3.2 nm and 680 ± 15.3 nm, for PLA NPs and PLA-FA NPs respectively) than the correspondent drug-loaded NPs (110 ± 3.8 nm and 875 ± 20.5 nm, respectively). Their biological activity was assessed on osteosarcoma bulk cells MG63, healthy osteoblast cell line (hFOB1.19), and enriched osteosarcoma cancer stem cells (CSCs), showing cell-depending effect. Entrapped Sal maintained its cytotoxic effect on CSCs and MG63 cells, with a potency comparable to the free drug and no evident benefit was detected for folate-decorated PLA NPs respect to native PLA NPs. Graphical abstract.

Quality Evaluation and Characterization of Fractions with Biological Activity from Ephedra Herb Extract and Ephedrine Alkaloids-Free Ephedra Herb Extract.

Previously, we reported that the c-Met inhibitory effect of Ephedra Herb extract (EHE) is derived from ingredients besides ephedrine alkaloids. Moreover, analgesic and anti-influenza activities of EHE and ephedrine alkaloids-free Ephedra Herb extract (EFE) have been reported recently. In this study, we examined the fractions containing c-Met kinase inhibitory activity from EHE and the fractions with analgesic and anti-influenza activities from EFE, and elucidated the structural characteristics of the active fractions. Significant c-Met kinase activity was observed in 30, 40, and 50% methanol (MeOH) eluate fractions obtained from water extract of EHE using Diaion HP-20 column chromatography. Similarly, 20 and 40% MeOH, and MeOH eluate fractions obtained from water extract of EFE were found to display analgesic and anti-influenza activities. Reversed phase-HPLC analysis of the active fractions commonly showed broad peaks characteristic of high-molecular mass condensed tannin. The active fractions were analyzed using 13C-NMR and decomposition reactions; the deduced structures of active components were high-molecular mass condensed tannins, which were mainly procyanidin B-type and partly procyanidin A-type, including pyrogallol- and catechol-type flavan 3-ols as extension and terminal units. HPLC and gel permeation chromatography (GPC) analyses estimated that the ratio of pyrogallol- and catechol-type was approximately 9 : 2, and the weight-average molecular weight based on the polystyrene standard was >45000. Furthermore, GPC-based analysis was proposed as the quality evaluation method for high-molecular mass condensed tannin in EHE and EFE.

A Candidate for Multitopic Probes for Ligand Discovery in Dynamic Combinatorial Chemistry.

Multifunctionalized materials are expected to be versatile probes to find specific interactions between a ligand and a target biomaterial. Thus, efficient methods to prepare possible combinations of the functionalities is desired. The concept of dynamic combinatorial chemistry (DCC) is ideal for the generation of any possible combination, as well as screening for target biomaterials. Here, we propose a new molecular design of multitopic probes for ligand discovery in DCC. We synthesized a new Gable Porphyrin, GP1, having prop-2-yne groups as a scaffold to introduce various functional groups. GP1 is a bis(imidazolylporphyrinatozinc) compound connected through a 1,3-phenylene moiety, and it gives macrocycles spontaneously and quantitatively by strong imidazole-to-zinc complementary coordination. Some different types of functional groups were introduced into GP1 in high yields. Formation of heterogeneous macrocycles composed of GP1 derivatives having different types of substituents was accomplished under equilibrium conditions. These results promise that enormous numbers of macrocycles having various functional groups can be provided when the kinds of GP components increase. These features are desirable for DCC, and the present system using GP1 is a potential candidate to provide a dynamic combinatorial library of multitopic probes to discover specific interactions between a ligand and a biomaterial.

[Determination of 3-nitropropionic acid in moldy sugarcane by gel filtration-high performance liquid chromatography].

OBJECTIVE: To establish a method for determination of 3-nitropropionic acid(3-NPA) in moldy sugarcane by gel permeation chromatography-high performance liquid chromatography(GPC-HPLC) method. METHODS: Samples were extracted by acetonitrile and cleaned up by GPC-HPLC. The sample extract was separated on an Agilent Eclipse plus C_(18) column(4.6 mm×150 mm, 5 µm), keeping the column temperature at 40℃ with an injection volume of 10 µL. The condition of mobile phase was KH_2PO_4 aqueous solution(0.02 mol/L, pH 3.0 with phosphoric acid) as mobile phase A and acetonitrile as mobile phase B for isocratic elution. RESULTS: There was a good linearity of the calibration curve for 3-NPA in the range of 0.5-20.0 mg/kg with the correlation coefficient of 0.99989. The detection limit of the method was 0.05 mg/kg, and the limit of quantification was 0.15 mg/kg. The average recovery at three spiked levels was in the range of 91.7%-99.4%, and the relative standard deviation was 7.23%-8.62%(n=6). CONCLUSION: This method is stable and accurate, and is suitable for the detection of 3-nitropropionic acid in sugarcane.

Medium Resolution 1 H-NMR at 62 MHz as a New Chemically Sensitive Online Detector for Size-Exclusion Chromatography (SEC-NMR).

A state-of-the-art, medium-resolution 1 H-NMR spectrometer (62 MHz) is used as a chemically sensitive online detector for size-exclusion chromatography of polymers such as polymethylmethacrylate (PMMA) and polystyrene (PS). The method uses protonated eluents and works at typical chromatographic conditions with trace amounts of analytes (<0.5 g L-1 after separation). Strong solvent suppression, e.g., by a factor of 500, is achieved by means of T1 -filtering and mathematical subtraction methods. Substantial improvements are made with respect to previous work in terms of the sensitivity (signal-to-noise ratio up to 130:1, PMMA OCH3 ) and selectivity (peak width, full width half maximum (FWHM) 4 Hz on-flow). Typical homopolymers and a blend are investigated to deformulate their composition along the dimensions of molecular weight and NMR chemical shift. These results validate this new hyphenated chromatography method, which can greatly facilitate analysis and is much more effective than previously published results.

Parameterizing amylose chain-length distributions for biosynthesis-structure-property relations.

Amylose, one of the components of starch, is a glucose polymer consisting largely of long, linear chains with a few long-chain branch points. The chain-length (molecular weight) distribution (CLD) of the component chains of amylose can provide information on amylose biosynthesis-structure-property relations, as has been done previously by fitting amylopectin CLDs to a model with physically meaningful parameters. Due to the presence of long chains, the CLD of amylose can currently best be obtained by size-exclusion chromatography, a technique that suffers from band-broadening effects which alter the observed distribution. The features of the multiple regions present in amylose chain-length distributions are also difficult to resolve, an issue that combines with band broadening to compound the difficulty of analysis and subsequent parameterization of the structural characteristics of amylose. A new method is presented to fit these distributions with biologically meaningful parameters in a way that accounts for band broadening. This is achieved by assuming that band broadening takes the form of a simple Gaussian over a relatively small region and that chain stoppage is a random process independent of the length of the substrate chain over the same region; these assumptions are relatively weak and expected to be frequently applicable. The method provides inbuilt consistency tests for its applicability to a given data set and, in cases where it is applicable, allows for the first nonempirical parameterization of amylose biosynthesis-structure-property relations from CLDs by using parameters directly linked to the activities of the enzymes responsible for chain growth and chain stoppage. Graphical abstract Model calculation illustrating the method described and showing the division between the three characteristic regions of a typical amylose chain-length distribution.

Synthesis of sharply thermo and PH responsive PMA-b-PNIPAM-b-PEG-b-PNIPAM-b-PMA by RAFT radical polymerization and its schizophrenic micellization in aqueous solutions.

Sharply thermo- and pH-responsive pentablock terpolymer with a core-shell-corona structure was prepared by RAFT polymerization of N-isopropylacrylamide and methacrylic acid monomers using PEG-based benzoate-type of RAFT agent. The PEG-based RAFT agent could be easily synthesized by dihydroxyl-capped PEG with 4-cyano-4-(thiobenzoyl) sulfanylpentanoic acids, using esterification reaction. This pentablock terpolymer was characterized by 1H NMR, FT-IR, and GPC. The PDI was obtained by GPC, indicating that the molecular weight distribution was narrow and the polymerization was well controlled. The thermo- and pH-responsive micellization of the pentablock terpolymer in aqueous solution was investigated using fluorescence spectroscopy technique, UV-vis transmittance, and TEM. The LCST of pentablock terpolymer increased (over 50 °C) compared to the NIPAM homopolymer (~32 °C), due to the incorporation of the hydrophilic PEG and PMA blocks in pentablock terpolymer (PNIPAM block as the core, PEG the block and the hydrophilic PMA block as the shell and the corona). Also, pH-dependent phase transition behavior shows at a pH value of about ~5.8, according to pKa of MAA. Thus, in acidic solution at room temperature, the pentablock terpolymer self-assembled to form core-shell-corona micelles, with the hydrophobic PMA block as the core, the PNIPAM block and the hydrophilic PEG block as the shell and the corona, respectively.

Extraction of cocoa proanthocyanidins and their fractionation by sequential centrifugal partition chromatography and gel permeation chromatography.

Cocoa beans contain secondary metabolites ranging from simple alkaloids to complex polyphenols with most of them believed to possess significant health benefits. The increasing interest in these health effects has prompted the need to develop techniques for their extraction, fractionation, separation, and analysis. This work provides an update on analytical procedures with a focus on establishing a gentle extraction technique. Cocoa beans were finely ground to an average particle size of <100 µm, defatted at 20 °C using n-hexane, and extracted three times with 50 % aqueous acetone at 50 °C. Determination of the total phenolic content was done using the Folin-Ciocalteu assay, the concentration of individual polyphenols was analyzed by electrospray ionization high performance liquid chromatography-mass spectrometry (ESI-HPLC/MS). Fractions of bioactive compounds were separated by combining sequential centrifugal partition chromatography (SCPC) and gel permeation column chromatography using Sephadex LH-20. For SCPC, a two-phase solvent system consisting of ethyl acetate/n-butanol/water (4:1:5, v/v/v) was successfully applied for the separation of theobromine, caffeine, and representatives of the two main phenolic compound classes flavan-3-ols and flavonols. Gel permeation chromatography on Sephadex LH-20 using a stepwise elution sequence with aqueous acetone has been shown for effectively separating individual flavan-3-ols. Separation was obtained for (-)-epicatechin, proanthocyanidin dimer B2, trimer C1, and tetramer cinnamtannin A2. The purity of alkaloids and phenolic compounds was determined by HPLC analysis and their chemical identity was confirmed by mass spectrometry.

Simultaneous determination of seven anticoagulant rodenticides in agricultural products by gel permeation chromatography and liquid chromatography-tandem mass spectrometry.

A sensitive and reliable method for the simultaneous determination of hydroxycoumarin-type (brodifacoum, bromadiolone, coumatetralyl, and warfarin) and indandione-type (chlorophacinone, diphacinone, and pindone) rodenticides in agricultural products by gel permeation chromatography (GPC) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed. The procedure involved extraction of the rodenticides from samples with acetone, followed by liquid-liquid partitioning with hexane/ethyl acetate (1:1, v/v) and 10% sodium chloride aqueous solution, then cleanup using GPC, and finally, analysis using LC-MS/MS. High recoveries from the GPC column were obtained for all rodenticides tested using a mobile phase of acetone/cyclohexane/triethylamine (400:1600:1, v/v/v). An ODS column, which contains low levels of metal impurities, gave satisfactory peak shapes for both hydroxycoumarin- and indandione-type rodenticides in the LC-MS/MS separation. The average recoveries of rodenticides from eight agricultural foods (apple, eggplant, cabbage, orange, potato, tomato, brown rice, and soybean) fortified at 0.0005-0.001 mg/kg ranged from 76 to 116%, except for bromadiolone in orange (53%) and diphacinone in soybean (54%), and the relative standard deviations ranged from 1 to 16%. The proposed method effectively removed interfering components, such as pigments and lipids, and showed high selectivity. In addition, the matrix effects were negligible for most of the rodenticide/food combinations. The results suggest that the proposed method is reliable and suitable for determining hydroxycoumarin- and indandione-type rodenticides in agricultural products.

Simultaneous enantioselective determination of triadimefon and its metabolite triadimenol in edible vegetable oil by gel permeation chromatography and ultraperformance convergence chromatography/tandem mass spectrometry.

A novel, sensitive, and efficient enantioselective method for the determination of triadimefon and its metabolite triadimenol in edible vegetable oil, was developed by gel permeation chromatography and ultraperformance convergence chromatography/tandem triple quadrupole mass spectrometry. After the vegetable oil samples were prepared using gel permeation chromatography, the eluent was collected, evaporated, and dried with nitrogen gas. The residue was redissolved by adding methanol up to a final volume of 1 mL. The analytes of six enantiomers were analyzed on Chiralpak IA-3 column (150 × 4.6 mm) using compressed liquid CO2-mixed 14 % co-solvents, comprising methanol/acetonitrile/isopropanol = 20/20/60 (v/v/v) in the mobile phase at 30 °C, and the total separation time was less than 4 min at a flow rate of 2 mL/min. Quantification was achieved using matrix-matched standard calibration curves. The overall mean recoveries for six enantiomers from vegetable oil were 90.1-97.3 %, with relative standard deviations of 0.8-5.4 % intra-day and 2.3-5.0 % inter-day at 0.5, 5, and 50 µg/kg levels. The limits of quantification were 0.5 µg/kg for all enantiomers based on five replicate extractions at the lowest fortified level in vegetable oil. Moreover, the absolute configuration of six enantiomers had been determined based on comparisons of the vibrational circular dichroism experimental spectra with the theoretical curve obtained by density functional theory calculations. Application of the proposed method to the 40 authentic vegetable oil samples from local markets suggests its potential use in enantioselective determination of triadimefon and triadimenol enantiomers. Graphical Abstract Chemical structures and UPC(2)-MS/MS separation chromatograms of triadimefon and triadimenol.

Comparison of different cleanup procedures for oil crops based on the development of a trace analytical method for the determination of pyraclostrobin and epoxiconazole.

The effects of different cleanup procedures in removing high-molecular-mass lipids and natural colorants from oil-crop extracts, including dispersive solid-phase extraction, low-temperature precipitation and gel permeation chromatography, were studied. The pigment removal, lipid quantity, and matrix effects of the three cleanup methods were evaluated. Results indicated that the gel permeation chromatography method is the most effective way to compare the dispersive solid-phase extraction and low-temperature precipitation. Pyraclostrobin and epoxiconazole applied extensively in oil-crop production were selected as typical pesticides to study and a trace analytical method was developed by gel permeation chromatography and ultra high performance liquid chromatography with tandem mass spectrometry. Average recoveries of the target pesticides at three levels (10, 50, and 100 µg/kg) were in the range of 74.7-96.8% with relative standard deviation values below 9.2%. The limits of detection did not exceed 0.46 µg/kg, whereas the limits of quantification were below 1.54 µg/kg and much lower than maximum residue limit in all matrices. This study may provide the essential data for optimizing the analytical method of pesticides in oil-crop samples.