Electrospray Ionization Source Research Articles

Development and validation of two bioanalysis methods for the determination of etimicin in human serum and urine by liquid chromatography-tandem mass spectrometry: Applications to a human pharmacokinetic and breakpoint study.

Etimicin is a fourth-generation aminoglycoside antibiotic. It has potent activity and low toxicity when employed for the treatment of Gram-negative and Gram-positive bacterial infections. The pharmacokinetics of etimicin in humans have not been elucidated completely. Two liquid chromatography-tandem mass spectrometry (LC-MS/MS) bioanalytical methods, without the use of any ion-pairing reagents, were developed and validated for the quantification of etimicin in human samples of serum and urine. Using a deuterated reagent as the internal standard, analytes in serum and urine samples were extracted by protein precipitation and dilution before LC-MS/MS analysis, respectively. For the two methods, chromatographic separations were undertaken under isocratic elution of water-ammonia solution-acetic acid (96:3.6:0.2, v/v/v) and methanol at 50%:50% and a flow rate of 0.35ml/min within 5min. A Waters XTerra MS C18 column (2.1 × 150mm, 3.5μm) and a column temperature of 40°C were chosen. A Sciex Qtrap 5500 mass spectrometer equipped with an electrospray ion source was used in both methods under multiple-reaction monitoring in positive-ion mode. The two methods showed good linearity, accuracy, and precision with high recovery and a minimal matrix effect in the range of 50.0-20000ng/ml for serum samples and 50.0-10000ng/ml for urine samples, respectively. Carry-over effects were not observed. Etimicin remained stable in human samples of serum or urine under the storage, preparation, and analytical conditions of the two methods. These two simple and reliable methods were applied successfully to a dose-escalation, phase I clinical trial of etimicin in Chinese healthy volunteers after intravenous administration of single and multiple doses. Based on these two methods we ascertained, for the first time, the comprehensive pharmacokinetics of etimicin in humans, which will be used for the exploration of the breakpoint research further.

Phytochemical study of Stachys sylvatica (Lamiaceae) aerial parts

In this article, the first complete phytochemical study of Stachys sylvatica (Lamiaceae) aerial parts, collected in Tuscan-Emilian Apennine, is reported. Twenty-three known compounds were isolated from the chloroform, chloroform-methanol 9:1 and methanol extracts; they were subsequently characterized by means of one- and two-dimensional nuclear magnetic resonance spectroscopy (1D- and 2D-NMR) and mass spectrometry as six simple phenolic derivatives (chlorogenic acid (1), caffeic acid (2), p-coumaric acid methyl ester (3), niduloic acid (4), hydroxytyrosol (5), tyrosol (6)), one glycosylated aliphatic alcohol (ebracteatoside B (7)), six glycosylated phenylpropanoids (decaffeoyl verbascoside (8), lavandulifolioside (9), betanyoside F (10), verbascoside (11), isoacteoside (12), cystanoside D (13)), four flavonoids (stachyspinoside (14), chrysoeriol 7-O-β-D-glucopyranoside (15), naringenin 7-O-(6’’-E + Z-p-coumaroyl)-β-D-glucopyranoside (16), apigenin 7-O-(6’’-E + Z-p-coumaroyl)-β-D-glucopyranoside (17)), three lignans (syringaresinol 4’-O-β-D-glucopyranoside (18), pinoresinol 4-O-β-D-glucopyranoside (19), 2-episesaminol 2-O-β-D-glucopyranoside (20)) and three ionones (loliolide (21), vomifoliol (22), (3S,5R,6R,7E,9R)-3,5,6,9-tetrahydroxy-7-megastigmene (23)). In addition, a further phenolic derivative (ethyl caffeate (b)) and four carboxylic acids (azelaic acid (a), trihydroxyoctadecadienoic acid (c), trihydroxyoctadecenoic acid (d) and dihydroxydodecadienoic acid (e)) have been tentatively identified by ultra-high performance liquid chromatography coupled to an electrospray ionization source high-resolution Orbitrap mass spectrometer (UHPLC-ESI-HR-Orbitrap/MS). Chemosystematic relationships between S. sylvatica and other related species were highlighted.

Development of a 3D-Printed Ionization Source for Single-Cell Analysis.

Understanding the physiologies and pathologies of diseases requires a thorough understanding of metabolic heterogeneity in cells. This technical note presents a 3D printing technology for manufacturing an ionization source that is specially adapted for mass spectrometry-based single-cell analysis. This all-in-one 3D-printed electrospray ionization source integrates the sample introduction, metabolite extraction, and ionization into one device, simplifying the process of single-cell analysis and improving the reproducibility of the measurement. We successfully used it for high-throughput analysis of three types of cancer cells (around 17 cells/min) and used the t-distributed stochastic neighbor embedding algorithm to distinguish different cell types based on detected metabolites. By simply adjusting the printing parameters of the 3D-printed ionization source, it can be applied to cells with different sizes. The proposed 3D-printed ionization source promises to open new possibilities for single-cell analysis.

Systematic analysis of in-source modifications of primary metabolites during flow-injection time-of-flight mass spectrometry

Flow-injection mass spectrometry (FI-MS) enables metabolomics studies with a very high sample-throughput. However, FI-MS is prone to in-source modifications of analytes because samples are directly injected into the electrospray ionization source of a mass spectrometer without prior chromatographic separation. Here, we spiked authentic standards of 160 primary metabolites individually into an Escherichia coli metabolite extract and measured the thus derived 160 spike-in samples by FI-MS. Our results demonstrate that FI-MS can capture a wide range of chemically diverse analytes within 30 s measurement time. However, the data also revealed extensive in-source modifications. Across all 160 spike-in samples, we identified significant increases of 11,013 ion peaks in positive and negative mode combined. To explain these unknown m/z features, we connected them to the m/z feature of the (de-)protonated metabolite using information about mass differences and MS2 spectra. This resulted in networks that explained on average 49 % of all significant features. The networks showed that a single metabolite undergoes compound specific and often sequential in-source modifications like adductions, chemical reactions, and fragmentations. Our results show that FI-MS generates complex MS1 spectra, which leads to an overestimation of significant features, but neutral losses and MS2 spectra explain many of these features.

Vending-Machine-Style Skin Excretion Sensing.

Skin metabolites show huge potential for use in clinical diagnostics. However, skin sampling and analysis workflows are tedious and time-consuming. Here, we demonstrate a vending-machine-style skin excretion sensing platform based on hydrogel-assisted sampling of skin metabolites. In this sensing platform, a sampling probe with hydrogel is held by a robotic arm. The robotic arm manoeuvres the probe to press it onto the forearm of a human subject. Due to the highly hydrophilic nature of the hydrogel, water-soluble metabolites─released by skin─are collected into the hydrogel, leaving behind the nonpolar metabolites. The probe is then inserted into a custom-made open port sampling interface coupled to an electrospray ion source of a high-resolution quadrupole-time-of-flight mass spectrometer. Metabolites in the hydrogel are immediately extracted by a solvent liquid junction in the interface and analyzed using the mass spectrometer. The ion current of the target analyte is displayed on a customized graphical user interface, which can also be used to control the key components of the analytical platform. The automated sampling and analysis workflow starts after the user inserts coins or presents an insurance card, presses a button, and extends an arm on the sampling area. The platform relies on low-cost mechanical and electronic modules (a robotic arm, a single-board computer, and two microcontroller boards). The limits of detection for standard analytes─arginine, citrulline, and histidine─embedded in agarose gel beds were 148, 205, and 199 nM, respectively. Various low-molecular-weight metabolites from human skin have been identified with the high-resolution mass spectrometer.

Simultaneous Determination of nicotinamide, kojic acid, Tranexamic acid, raspberry glycoside, azelaic acid, magnesium ascorbate phosphate and β-Arbutin in whitening cosmetics by UPLC-MS/MS

A UPLC-MS/MS method was developed for simultaneous determination of 7 whitening ingredients: nicotinamide, kojic acid, Tranexamic acid, raspberry glycoside, azelaic acid, magnesium ascorbate phosphate and β-Arbutin in cosmetics. The whitening active components were extracted from cosmetics by supersonic extraction with sodium chloride and dichloromethane to disperse the sample, and supersonic extraction with 0.015 mol/L potassium dihydrogen phosphate solution purified by HLB solid phase extraction column, scanned and detected by electrospray ionization source with positive and negative ion alternate scanning mode and multiple reaction monitoring mode. The results showed that the whitening active ingredients were separated within 3 minutes, with a good linear relationship (R>0.999), and the detection limit was 0.10mg/kg~0.75mg/kg. The recoveries (n=6) were 78.84%-104.85%, and the RSDs were 0.24%-11.35%. This method is suitable for the rapid determination of whitening active ingredients in cosmetics.

Quasi-continuous infrared matrix-assisted laser desorption electrospray ionization source coupled to a quadrupole time-of-flight mass spectrometer for direct analysis from well plates.

High-throughput screening (HTS) is a technique mostly used by pharmaceutical companies to rapidly screen multiple libraries of compounds to find drug hits with biological or pharmaceutical activity. Mass spectrometry (MS) has become a popular option for HTS given that it can simultaneously resolve hundreds to thousands of compounds without additional chemical derivatization. For this application, it is convenient to do direct analysis from well plates. Herein, we present the development of an infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) source coupled directly to an Agilent 6545 for direct analysis from well plates. The source is coupled to a quadrupole time-of-flight (Q-TOF) mass spectrometer to take advantage of the high acquisition rates without sacrificing resolving power as required with Orbitrap or Fourier-transform ion cyclotron resonance (FTICR) instruments. The laser used for this source operates at 100 Hz, firing 1 pulse-per-burst, and delivers around 0.7 mJ per pulse. Continuously firing this laser for an extended duration makes it a quasi-continuous ionization source. Additionally, a metal capillary was constructed to extend the inlet of the mass spectrometer, increase desolvation of electrospray charged droplets, improve ion transmission, and increase sensitivity. Its efficiency was compared with the conventional dielectric glass capillary by measured signal and demonstrated that the metal capillary increased ionization efficiency due to its more uniformly distributed temperature gradient. Finally, we present the functionality of the source by analyzing tune mix directly from well plates. This source is a proof of concept for HTS applications using IR-MALDESI coupled to a different MS platform.

Chemical constituents in different parts of Ixeris sonchifolia based on UPLC-LTQ-Orbitrap-MS~n

The chemical constituents in stem leaf, root, and flower of Ixeris sonchifolia were identified by the ultra performance li-quid chromatography coupled with linear ion trap quadrupole-orbitrap mass spectrometry(UPLC-LTQ-Orbitrap-MS~n). The separation was performed on an Acquity UPLC BEH C_(18) column(2.1 mm×100 mm, 1.7 μm) with a mobile phase of water(containing 0.1% formic acid, A)-acetonitrile(B) with gradient elution. With electrospray ionization source, the data of 70% methanol extract from stem leaf, root and flower of I. sonchifolia were collected by high-resolution full-scan Fourier transform spectroscopy, data dependent acquisition, precursor ion scan, and selected ion monitoring in the negative and positive ion modes. The compounds were identified based on accurate molecular weight, retention time, fragment ions, comparison with reference standard, Clog P and references. A total of 131 compounds were identified from the 70% methanol extract of I. sonchifolia, including nucleosides, flavonoids, organic acids, terpenoids, and phenylpropanoids, and 119, 110, and 126 compounds were identified from the stem leaf, root and flower of I. sonchifolia, respectively. In addition, isorhamnetin, isorhamnetin-7-O-sambubioside and caffeylshikimic acid were discovered from I. sonchifolia for the first time. This study comprehensively analyzed and compared the chemical constituents in different parts of I. sonchifolia, which facilitated the discovery of effective substances and the development and application of medicinal material resources of I. sonchifolia.

Diversity of organic components in airborne waste discharged from sewer pipe repairs.

Air-discharged waste from commonly used trenchless technologies of sewer pipe repairs is an emerging and poorly characterized source of urban pollution. This study reports on the molecular-level characterization of the atmospherically discharged aqueous-phase waste condensate samples collected at four field sites of the sewer pipe repairs. The molecular composition of organic species in these samples was investigated using reversed-phase liquid chromatography coupled with a photodiode array detector and a high-resolution mass spectrometer equipped with interchangeable atmospheric pressure photoionization and electrospray ionization sources. The waste condensate components comprise a complex mixture of organic species that can partition between gas-, aqueous-, and solid-phases when water evaporates from the air-discharged waste. Identified organic species have broad variability in molecular weight, molecular structures, and carbon oxidation state, which also varied between the waste samples. All condensates contained complex mixtures of oxidized organics, N- and S-containing organics, condensed aromatics, and their functionalized derivatives that are directly released to the atmospheric environment during installations. Furthermore, semi-volatile, low volatility, and extremely low volatility organic compounds comprise 75-85% of the total compounds identified in the waste condensates. Estimates of the component-specific viscosities suggest that upon evaporation of water waste material would form the semi-solid and solid phases. The low volatilities and high viscosities of chemical components in these waste condensates will contribute to the formation of atmospheric secondary organic aerosols and atmospheric solid nanoplastic particles. Lastly, selected components expected in the condensates were quantified and found to be present at high concentrations (1-20 mg L-1) that may exceed regulatory limits.

Two-dimensional isomer differentiation using liquid chromatography-tandem mass spectrometry with in-source, droplet-based derivatization.

Saccharides are increasingly used as biomarkers and for therapeutic purposes. Their characterization is challenging due to their low ionization efficiencies and inherent structural heterogeneity. Here, we illustrate how the coupling of online droplet-based reaction, in a form of contained electrospray (ES) ion source, with liquid chromatography (LC) tandem mass spectrometry (MS/MS) allows the comprehensive characterization of sucrose isomers. We used the reaction between phenylboronic acid and cis-diols for on-the-fly derivatization of saccharides eluting from the LC column followed by in situ MS/MS analysis, which afforded diagnostic fragment ions that enabled differentiation of species indistinguishable by chromatography or mass spectrometry alone. For example, chromatograms differing only by 2% in retention times were flagged to be different based on incompatible MS/MS fragmentation patterns. This orthogonal LC-contained-ES-MS/MS method was applied to confirm the presence of turanose, palatinose, maltulose, and maltose, which are structural isomers of sucrose, in three different honey samples. The reported workflow does not require modification to existing mass spectrometers, and the contained-ES platform itself acts both as the ion source and the reactor, all promising widespread application.

Analysis of volatile short-chain fatty acids in the gas phase using secondary electrospray ionization coupled to mass spectrometry.

Quantification of metabolites present within exhaled breath is a major challenge for on-line breath analysis. It is also important for gauging the analytical performance, accuracy, reproducibility, reliability, and stability of the measuring technology. Short-chain fatty acids (SCFAs) are of high interest for nutrition and health. Their quantification enables a deep mechanistic understanding of a wide range of biological processes and metabolic pathways, while their high volatility makes them an attractive target for breath analysis. This article reports, for the first time, the development and testing of a modular, dynamic vapor generator for the qualitative and quantitative analysis of volatile SCFAs in the gaseous phase using a secondary electrospray ionization (SESI) source coupled to a high-resolution mass spectrometer. Representative compounds tested included acetic acid, propionic acid, butyric acid, pentanoic acid and hexanoic acid. Gas-phase experiments were performed both in dry and humid (95% relative humidity) conditions from ppt to low ppb concentrations. The results obtained exhibited excellent linearity within the examined concentration range, low limits of detection and quantification down to the lower ppt area. Mixture effects were also investigated and are presented.

Use of mass spectrometry as a tool for the search or identification of flavonoids in Urticaceae

Abstract The Urticaceae family, circumscribed within the Rosales, was investigated in this study with an overview of the current literature about phytochemical studies using the Liquid Chromatography coupled to Mass Spectrometry (LC-MS) technique. The aim of this study was to review the secondary metabolites identified in the Urticaeae using LC-MS analysis. A systematic review was performed using Scifinder and ScienceDirect databases. Phenolic substances are the most abundant in the Urticaceae family, especially flavones, phenolic acids, and flavonols. We have shown that flavonoids are important chemotaxonomic markers of the chemical composition of the Urticaeae. Following chemical attributes, the C-glycosylated and O-glycosylated flavones stand out as the main skeletons. Our results revealed the chemical profile and structural variability of micromolecules from each genus of Urticaceae. This approach demonstrates a greater use of reversed-phase and liquid chromatography coupled to a mass spectrometer with a negative mode electrospray ionization (ESI) source. In addition, the mobile phase is usually composed of binary systems and eluted by gradient systems. Finally, this paper presents the identification of molecular ion patterns and fragmentation of chemical markers in Urticaceae, identified and isolated using LC-MS, which has been proven to be a valuable tool in several areas, such as phytochemistry, chemosystematics, and chemophenetics. In conclusion, this review is expected to help identify and separate phenolic compounds from the Urticaceae family.

Determination of organophosphate diesters in facility vegetable soils using ultra-high performance liquid chromatography-electrostatic field orbitrap high resolution mass spectrometry

Organophosphate diesters (Di-OPEs) are biotic or abiotic degradation products of organophosphate esters (OPEs). Current analytical methods focus on detecting Di-OPEs in human urine. Human exposure to Di-OPEs in environmental matrices has not been systematically studied. Soil plays an important role in the environmental migration and transformation of organic pollutants. Previous studies found that OPEs are ubiquitous in soil. However, few studies reported OPEs metabolite pollution in soil, especially in facility vegetable soil. In this study, an ultra-high performance liquid chromatography-electrostatic field orbitrap high resolution mass spectrometry (UPHLC-Orbitrap HRMS) method was developed for the determination of five Di-OPEs (bis(2-chloroethyl) phosphate (BCEP), bis(1,3-dichloro-2-propyl) phosphate (BDCP), di-n-butyl phosphate (DnBP), diphenyl phosphate (DPhP), and bis(2-ethylhexyl) phosphate (DEHP)) in the facility vegetable soil. The pretreatment process and chromatographic and mass spectrometric conditions were optimized in the present study. Comparative study of the purification effects of different solid-phase extraction columns showed that Oasis WAX cartridge had best purification efficiency for the five Di-OPEs. The cartridge was first activated using 3 mL methanol, 3 mL methanol containing 5% (v/v) ammonia, and 3 mL 0.1 mol/L sodium acetate-acetic acid buffer solution. Then, the cartridge was rinsed with 3 mL of 30% (v/v) methanol aqueous solution, and finally eluted using 8 mL methanol containing 5% (v/v) ammonia. The effects of mobile phase (with respect to solvent composition and flow rate) and column temperature on the shape and intensity of chromatographic peaks were studied. The optimized UHPLC conditions were as follows: chromatographic column, Thermo Accucore RP-MS; column temperature, 30 ℃; mobile phase, 0.2 mmol/L ammonium acetate aqueous solution and methanol; flow rate, 0.2 mL/min. In the UHPLC-Orbitrap HRMS experiment, the five Di-OPEs were analyzed in full MS mode with negative ionization. Instrumental parameters, such as sheath gas and auxiliary gas, were optimized to determine the MS conditions. The optimized Orbitrap HRMS conditions were as follows: heating electrospray ionization source (HESI), full MS mode with negative ionization; scan range, m/z 100-500; ion transfer tube temperature, 320 ℃; automatic gain control of target particle count, 1×106; sheath gas flow rate, 8.58 L/min; auxiliary gas flow rate, 17.40 L/min; spray voltage, 3.2 kV; and S-lens voltage, 50 V. The limits of detection and quantification were 0.001-0.047 ng/g and 0.004-0.156 ng/g, respectively. The correlation coefficients of the calibration curve were 0.9985-0.9999. At three spiked levels, 5.0, 25.0, and 50.0 ng/g, the recoveries of the five Di-OPEs ranged from 56.9% to 133.0% with relative standard deviations of 4.4%-18.9%. The established method was applied to the analysis of the five Di-OPEs in 16 facility vegetable soils. The detection frequencies of the five Di-OPEs exceeded 60% in all soil samples, indicating that the Di-OPEs were ubiquitous in the facility vegetable soil. The contents of the five Di-OPEs in the facility vegetable soil samples ranged from 2.53-6.94 ng/g. DnBP (1.37-3.20 ng/g) and DPhP (0.47-2.44 ng/g) were the predominant congeners in the facility vegetable soil samples, accounting for 23.4%-68.8% and 16.3%-35.9% of the five Di-OPEs, respectively. The developed method is simple, sensitive, and reproducible and can be used effectively for the determination of Di-OPEs in soil. The results of this study will be helpful for understanding the environmental behavior of Di-OPEs and their human exposure in facility vegetable soils.

Determination of five amanita peptide toxins in poisonous mushrooms by ultra performance liquid chromatography-quadrupole electrostatic field orbitrap high resolution mass spectrometry

Food poisoning by toxic mushrooms occurs frequently worldwide. It is one of the most common food poisoning events and the main cause of death. Amanita peptide toxins are the most common lethal toxins in poisonous mushrooms. Presently, a novel method based on ultra performance liquid chromatography-quadrupole electrostatic field orbitrap high resolution mass spectrometry (UPLC-Q/Orbitrap HRMS) was developed for the determination of five amanitapeptide toxins (α-amanitin, β-amanitin, γ-amanitin, phalloidin, and phallacidin). Because the isotope summit of α-amanitin affects the detection of β-amanitin, it cannot be distinguished by low resolution mass spectrometry. Therefore, experimental conditions including chromatography and mass spectrometry were explored in detail. The five peptide toxins were extracted from poisonous mushrooms with pure water and filtered through a 0.22 μm teflon microporous membrane. The procedure was rapid, simple, and environmentally friendly. Chromatographic separation was performed on a strong polarity HSS T3 column (100 mm×2.0 mm, 2.1 μm) with gradient elution using acetonitrile and 5 mmol/L ammonium acetate containing 0.1% (v/v) formic acid as mobile phases at a flow rate of 0.3 mL/min. The column temperature was set to 40 ℃. The analytes were ionized using a heating electrospray ionization source and collected in positive ion mode. Full scanning/data-dependent secondary mass spectrometry (Full mass-ddMS2) mode was used for qualitative analysis of the targets within 10 min. The target ion selective scan (Targeted-SIM) mode was used for quantification by external standard calibration. The measured and theoretical values of the exact mass and the MS2 fragment ions of the five compounds were within an error of 5×10-6. Method validation was performed according to the criteria recommended by the Chinese National Standard. All the compounds showed an excellent linear relationship in the range of 1.0-20.0 μg/L. The correlation coefficients (r) ranged from 0.9974 to 0.9989. The limit of detection was 0.006 mg/kg for all five compounds. Recoveries ranged from 81.8% to 102.4%. There was no matrix effect in the blank mushroom sample for the five compounds, and the relative standard deviations ranged from 3.2% to 8.3%. This method provides abundant compound characteristic mass information, such as retention time, exact mass, fragment ions, and other information. The data can be used to identify suspected compounds based on the extracted ion flow diagram and isotope distribution information. Comparison between the actual exact mass and the theoretical exact mass, combined with the fragment ions enables identification of the structures of unknown compounds and collision methods, which can be confirmed in the absence of standard materials. In this study, the isomer of γ-amanitin was identified as amaninamide. The novel method is simple, accurate, specific, and sensitive. The method permits the rapid qualitative and quantitative detection of compound in public health emergency settings and will provide reliable technical support for the rapid screening of such toxic compounds and the structural locking of unknown toxins in the future.

Correlation analysis of modern analytical data - a chemometric dissection of spectral and chromatographic variables.

The Standard Practices for Infrared Multivariate Quantitative Analysis (ASTM E1655) provide a guide for determining physicochemical properties of materials using multivariate calibration techniques applied to chemical sources that have high multicollinearity and correlated information. Partial least squares (PLS) is the most widely used multivariate regression method due to its excellent prediction capabilities and easy optimization. Initially applied to chromatographic data, PLS has also shown great results in near-infrared (NIR) and mid-infrared (MIR) spectroscopies. However, complex chemical matrices with low correlation may not be efficiently modeled using PLS or other multivariate analyses limited by grouping similar information (such as latent variables or principal components). Therefore, this study aims to evaluate the multicollinearity of different analytical techniques, such as high-temperature gas chromatography (HTGC), NIR, MIR, hydrogen nuclear magnetic resonance (1H NMR), carbon-13 nuclear magnetic resonance (13C NMR), and Fourier transform ion cyclotron resonance mass spectrometry coupled to the electrospray source in positive and negative ionization modes (ESI(±)FT-ICR). Descriptive statistics (coefficient of determination, R2) and principal component analysis (PCA) were used to identify the distribution of correlated information. Results showed that NIR and MIR spectroscopies exhibited a higher percentage of correlated variables, while 13C NMR and ESI(±)FT-ICR MS had more discrete profiles. Therefore, PLS development may be more effectively applied to NIR, MIR, and 1H NMR data, while 13C NMR and mass spectra may require other algorithms or variable selection methods in combination with PLS.

Assessment of Micro- and Nanoplastic Composition (Polymers and Additives) in the Gastrointestinal Tracts of Ebro River Fishes

One of the main routes of fish exposure to micro- and nanoplastics (MNPLs) is their ingestion. MNPLs can act as reservoirs of organic contaminants that are adsorbed onto their surfaces, or that can leach from their complex formulations, with potential impacts on biota and along the aquatic food chain. While MNPLs have been reported in fishes worldwide, complete information on MNPL compositions, polymers and additives continues to be scarce. In this work, the presence of MNPLs in the gastrointestinal tracts (GIT) of fish from the Ebro River (Spain) was investigated using a double suspected screening approach to assess and quantify polymers and additives. The sample-preparation procedure consisted of sequential alkaline and acidic digestions with KOH and HNO3, followed by ultrasonic-assisted extraction (USAE) with toluene. The analysis of polymers was carried out with size-exclusion chromatography followed by high-resolution mass spectrometry using an atmospheric pressure photoionization source, operating in negative and positive ionisation modes (SEC-(±)-APPI-HRMS) using full-scan acquisition (FS). Plastic additives were assessed using high-performance liquid chromatography with a C18 analytical column coupled to HRMS equipped with an electrospray ionisation source operating under positive and negative conditions (LC-(±ESI)-HRMS). The acquisition was performed in parallel with full-scan (FS) and data-dependent scan (ddMS2) modes, working under positive and negative ionisation modes. The polymers most frequently detected and quantified in fish GITs were polysiloxanes, polyethylene (PE), polypropylene (PP) and polystyrene (PS). PE was detected in 84% of the samples, with a concentration range from 0.55 to 3545 µg/g. On the other hand, plasticisers such as phthalates and stabilisers such as benzotriazoles were the most frequently identified plastic additives.

Phytochemical Investigation of Marker Compounds from Indigenous Korean Salix Species and Their Antimicrobial Effects.

Salix species, including willow trees, are distributed in the temperate regions of Asian countries, including South Korea. Willow trees are used to treat pain and inflammatory diseases. Due to the medicinal properties of willow trees, pharmacological studies of other Salix spp. have gained attention; however, only a few studies have investigated the phytochemicals of these species. As part of our ongoing natural product research to identify bioactive phytochemicals and elucidate their chemical structures from natural resources, we investigated the marker compounds from indigenous Korean Salix species, namely, Salix triandra, S. chaenomeloides, S. gracilistyla, S. koriyanagi, S. koreensis, S. pseudolasiogyne, S. caprea, and S. rorida. The ethanolic extract of each Salix sp. was investigated using high-performance liquid chromatography combined with thin-layer chromatography and liquid chromatography−mass spectrometry-based analysis, and marker compounds of each Salix sp. were isolated. The chemical structures of the marker compounds (1−8), 3-(4-hydroxyphenyl)propyl β-D-glucopyranoside (1), 2-O-acetylsalicin (2), 1-O-p-coumaroyl glucoside (3), picein (4), isograndidentatin B (5), 2′-O-acetylsalicortin (6), dihydromyricetin (7), and salicin (8) were elucidated via nuclear magnetic resonance spectroscopy and high-resolution liquid chromatography−mass spectrometry using ultrahigh-performance liquid chromatography coupled with a G6545B Q-TOF MS system with a dual electrospray ionization source. The identified marker compounds 1−8 were examined for their antimicrobial effects against plant pathogenic fungi and bacteria. Dihydromyricetin (7) exhibited antibacterial activity against Staphylococcus aureus, inducing 32.4% inhibition at a final concentration of 125 μg/mL with an MIC50 value of 250 μg/mL. Overall, this study isolated the marker compounds of S. triandra, S. chaenomeloides, S. gracilistyla, S. koriyanagi, S. koreensis, S. pseudolasiogyne, S. caprea, and S. rorida and identified the anti-Staphylococcus aureus bacterial compound dihydromyricetin.

Analysis of Chemical Constituents of Traditional Chinese Medicine Jianqu before and after Fermentation Based on LC-MS/MS

To detect the chemical constituents in Jianqu samples under different fermentated states by using UPLC-QTOF-MS/MS technology, to conduct preliminary analyses, and to establish an HPLC method for the simultaneous determination of hesperidin and naringenin in Jianqu, and the variation of the two components during fermentation were compared. Waters ACQUITYTM UPLC HSST3 column (2.1 mm × 100 mm, 1.8 μm) was used; the mobile phase was 0.1% formic acid aqueous solution (A)-0.1% formic acid acetonitrile (B); The flow rate was 0.4 mL·min-1 with gradient elution; the column temperature was 45 °C; injection volume was 5 μL. The mass spectra of the samples were collected by negative ion mode under the electrospray ion source, and the data were screened and matched by UNIFI software. Hypersil gold C18 column (100 mm × 2.1 mm, 1.9 μm) was used; the mobile phase was acetonitrile (A)-0.1% acetic acid (B);; the flow rate with gradient elution was 0.3 mL·min-1; the column temperature was 30 °C; the injection volume was 2 μL. The content changes of hesperetin and naringenin in Jianqu at different fermentation time were detected. A total of 54 compounds were identified, including flavonoids, amino acids, organic acids, terpenoids, coumarins, lignans, and other compounds. Under the selected HPLC conditions, the linear relationship between hesperidin and naringenin was discovered (r2 = 0.9996). The content of hesperidin and naringenin changed significantly in the whole fermentation process. The highest concentration of content was observed at 36 h of fermentation and then decreased to varying degrees. This experiment can effectively identify various chemical components in Jianqu during different fermentation periods, and determine the content of the characteristic components, so as to provide a scientific basis for further study of Jianqu fermentation processing technology as well as a sound pharmacodynamic material basis.

Characterization of Sulfides in Atmospheric Residue Fractions Using Selective Oxidation Followed by Fourier Transform Ion Cyclotron Resonance Mass Spectrometry

An improved analytical method was developed for identifying sulfide compounds from aromatic fractions in Lungu atmospheric residue (LGAR). Sulfides in residue aromatics were selectively oxidized into sulfoxides using tetrabutyl ammonium periodate (TBAP) and identified by positive-ion Electrospray Ionization Source (ESI) Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS). Results showed that sulfides with lower polarity in LGAR aromatics could effectively ionize in ESI using this approach. Additionally, the oxidized sulfides were mainly S1 and S2 class species. The most abundant oxidation-generated sulfoxides O1S1 and O1S2 in LGAR aromatics had DBE values of 3~10 and 8~12, respectively. The S2 class species, whose condensation degree was higher than that of S1, were likely in the form of containing both cyclic sulfides and thiophenic compounds.

Cytotoxicity assessment and suspected screening of PLASTIC ADDITIVES in bioplastics of single-use household items

Bioplastics made of renewable sources provide an excellent alternative to fossil-based materials. However, similar or greater quantities of plastic additives than fossil-based plastics are used in the formulations of bioplastics to improve their performance and barrier properties. Nowadays, there is an increasing concern about sources of chemical exposure. However, there is an important knowledge gap regarding complex additive mixtures, particularly in bio-based materials. In this study, we have characterised the presence of plastic additives in single-use materials (collected from retail shops in Spain), which are made of the most common bio-based biodegradable materials, poly(lactic acid) (PLA) and poly(hydroxybutyrate) (PHB), in contrast with a fossil-based plastic material that is extensively made from high-density polyethylene (HDPE). The approach consisted of the pulverization of material in the nano-micro range (100 nm-10 μm), with the materials being extracted using different solvents and ultrasonic-assisted solvent extraction (UASE). 100% of the additives in the material cannot be extracted, but since they were performed in the same condition for all materials can inform about the fingerprint of primary organics and the relative abundances between the different materials. The extracts were analysed by high-performance liquid chromatography coupled with high-resolution mass spectrometry equipped with a heated electrospray ionisation source operated in positive and negative ionisation conditions (HPLC-HESI(+/−)-HRMS), separately, using a suspect screening approach. A total number of 203 additives were tentatively identified (confidence level 2) in the bioplastics items of this study. An average of 123 plastic additives were found in PLA items and 121 in PHB items. Plasticisers were the most abundant additives; the phthalates group was the most commonly found, while 63 plastic additives were confirmed by standards and quantified. In parallel, the cytotoxicity of plastic particles in terms of cell viability and oxidative stress was studied using A549 alveolar basal epithelial cells, and the toxicity of the different extracts was also established using HepG2 adenocarcinoma cells. The main results of this study demonstrate that the plastic particles did not show a significant reduction in cell viability, but oxidative stress was significant, with PLA being the material that showed the highest effect. On the other hand, extracts of plastic particles did not show inhibition of cell viability except for HDPE extract, but the different extracts produced oxidative stress, with PLA showing the highest effect. Although the item showing the highest concentrations of additives was the extract of PLA material while also showing the most elevated oxidative stress, the low migration of toxicants from plastic materials ensures their safe use. However, this also supports the idea that bioplastics can contain many toxic substances in their formulations, some of which are unknown and should be studied in more depth.