Hydrophilic-lipophilic Balanced Research Articles

Evaluation of GC-EI&CI-TOFMS for Nontarget Analysis of Industrial Wastewater Using Hydrophilic-Lipophilic-Balanced SPME.

The evaluation of nontarget analysis (NTA) techniques for the monitoring of wastewater is important as wastewater is an anthropogenic pollution source for aquatic ecosystems and a threat to human and environmental health. This study presents the proof-of-concept NTA of industrial wastewater samples. A prototype hydrophilic-lipophilic-balanced (HLB) SPME and gas chromatography interfaced with time-of-flight high-resolution mass spectrometry (GC-TOFMS) with electron ionization (EI) and chemical ionization (CI) in parallel are employed. The HLB-SPME consists of a poly(divinylbenzene-co-N-vinylpyrrolidone) structure, allowing the extraction of hydrophilic as well as lipophilic substances. As the combination of parallel CI and EI data provides a comprehensive data set as a unique feature, this study is strongly focused on the compound identification procedure and confidence reporting of exemplary substances. Furthermore, the use of three different CI reagent ions, including [N2H]+/[N4H]+, [H3O]+, and [NH4]+, enables a broad range of analytes to be ionized in terms of selectivity and softness. The complementary information provided by EI and CI data allows a level 3 identification or higher in 69% of cases. The polarity coverage based on the physicochemical properties of the analytes (such as volatility, water solubility, hydrophilicity, and lipophilicity) was visualized by using Henry's law and octanol-water partitioning constants. In conclusion, the presented approach is shown to be valuable for water analysis and allows enhanced and accelerated compound identification compared to utilizing only one type of ionization.

Characterization of a hydrophilic-lipophilic balanced SPME material for enrichment of analytes with different polarities from aqueous samples

The choice of the extraction material is a critical step in solid-phase microextraction (SPME), especially the extraction of analytes which strongly differ in their polarity is challenging. This study is intended to give first insights into the headspace extraction of analytes with large differences in polarity by using a novel hydrophilic-lipophilic-balanced (HLB) SPME material with different particle sizes (5 µm, 5 + 30 µm, and 30 µm) in classical SPME, as well as SPME arrow format. The obtained results were compared to conventional and already established divinylbenzene carbon wide-range polydimethylsiloxane (DVB-CWR-PDMS) and PDMS SPME arrow fibers. The chosen model analytes toluene, indole, phenol, anisole, o-xylene, naphthalene, 2-heptanone, n-dodecane, and lindane are assigned to different substance classes with Kaw values ranging from -5.0–2.5 and Kow values from 1.5 to 7.5. The highest amount of the analytes was extracted by the HLB 5 + 30 µm arrow and the HLB 5 µm arrow, whereas the HLB SPME with a six times smaller phase volume compared to an SPME arrow, performed almost as well as the DVB-CWR-PDMS arrow. The 8-point calibration with the 5 + 30 µm HLB SPME arrow in the range of 5–70 µg L−1 of the analytes showed good linearity with R2 values ranging from 0.9765 to 0.9982, MDLs from 0.9 to 6.2 µg L−1 and RSDs from 2 to 31 %. For all of the observed analytes, the HLB material performed better than the conventional extraction materials and has great potential to replace the conventional extraction materials in many applications.

Performance Evaluation of Extraction Coatings with Different Sorbent Particles and Binder Composition.

Binders are critical components used in the preparation of a range of extraction devices, including solid-phase microextraction (SPME) devices. While the main role of a binder is to affix the sorbent particles to the selected support, it is critical to select the optimal binder to ensure that it does not negatively impact the coating's particle sorption capability. This work presents the first comprehensive investigation of the interactions between binders and solid sorbent particles as these interactions can significantly impact the performance of the coating. Specifically, the findings presented herein provide a better understanding of the extraction mechanisms of composite coatings and new rules for predicting the particle adhesion forces and binder distribution in the coating. The influence of binder chemistry on coating performance is investigated by examining a selection of the most used binders, namely, polydimethylsiloxane (PDMS), polyacrylonitrile (PAN), poly(vinylidene difluoride) (PVDF), polytetrafluoroethylene amorphous fluoroplastics (PTFE AF 2400), and polybenzimidazole (PBI). The solid particles (e.g., hydrophilic-lipophilic balanced (HLB) and C18) used in this work were selected for their ability to provide optimal extraction coverage for a broad range of analytes. The results show that PDMS does not change the properties of the solid particles and that the binder occupies a negligible volume due to shrinking after polymerization, resulting in the solid particles making up most of the coating volume. Hence, the coating sorption characteristics correspond closely to the properties of the selected solid particles. On the other hand, the results also showed that PTFE AF 2400 can interact with the active surface of the sorbent, leading to the deactivation of the sorbent particles. Therefore, the extraction performance and permeability coefficients decrease as the size of the penetrant increases, indicating a rigid porous structure. The results of this study can aid in the optimization of SPME devices as they provide reference values that can be used to determine the optimal binder and the sorbent affinity for the targeted compounds. Finally, the present work also provides the broader scientific community with a strategy for investigating the properties of sorbent particle/binder structures and defines the characteristics of a good coating/membrane by analyzing all parameters such as kinetics, thermodynamic equilibria, and morphology.

Sorption affinity and mechanisms of per-and polyfluoroalkyl substances (PFASs) with commercial sorbents: Implications for passive sampling

Effective monitoring tools, including passive samplers, are essential for the wide range of per- and polyfluoroalkyl substances (PFASs) in aquatic matrices. However, knowledge of the extent and mechanisms of PFASs sorption with sorbents in a passive sampling context is limited. To address this, sorption behavior of 45 anionic, neutral and zwitterionic PFASs ranging in perfluorocarbon chain length (C3-C16) and functional groups with 11 different commercial sorbents (cross-linked β-cyclodextrin polymers, activated carbon, anion exchange (AE), cation exchange, hydrophilic-lipophilic balanced (HLB) and non-polar) was investigated. A broad range of equilibrium sorbent-MilliQ water (MQ) distribution coefficients (Kd) were observed (10−1.95 to 108.30 mL g−1). Similar sorbent types (e.g., various AE and HLB sorbents) exhibited very different sorption behavior, likely due to their different polymeric structures and relative importance of sorbate/sorbent interactions other than coulombic interactions. HLB and AE with hydroxyl functionalities are most effective for sampling of the full suite of PFASs. Reduced sorptive affinity was observed in the presence of matrix co-constituents in wastewater influent for most PFASs. HLB had the smallest reduction in log Kd in wastewater suggesting that these sorbents are appropriate for applications in complex matrices. Sufficient sorbent capacity was observed for linear uptake of many target analytes which facilitates passive sampling.

Sequential Solid-Phase Microextraction with Biocompatible Coating Materials to Address Displacement Effects in the Quantitative Analysis

Solid-phase microextraction (SPME) is a simple and effective sample-preparation technique for the analysis of complex samples. However, sample matrices containing high concentrations of nonpolar substances or spiked analytes in free form can cause swelling, saturation, and/or competition phenomena in the coating material. This results in a displacement effect wherein polar analytes with low affinities for the solid coating material are displaced by nonpolar substances in the matrix or spiked analytes with a high affinity. Therefore, the quantitative analysis of polar analytes can be challenging, as the displacement effect causes non-linearity in the calibration curves. This paper presents a comprehensive investigation of the conditions under which the displacement effect occurs and how it influences the quantitative analysis of polar analytes. To remedy this issue, a sequential SPME strategy using two SPME blades with different selectivities is applied. SPME blades offer a large surface area and coating volume─and thus, greater extraction capacity─which may mitigate the displacement effect. In addition, the biocompatible coatings on the SPME blades are comprised of small amounts of sorbent particles embedded by a polyacrylonitrile (PAN) binder, which allows them to be directly immersed into complex matrixes such as biological and food samples, as the PAN acts as a barrier that prevents the adsorption of large macromolecules (e.g., cells and proteins). As such, a C18/PAN-coated blade was applied for the first extraction step, which significantly decreased the concentrations of nonpolar compounds in the sample. In the second step, a hydrophilic-lipophilic balanced (HLB)/PAN-coated blade was employed to extract the polar analytes and any remaining nonpolar analytes. The proposed sequential SPME strategy successfully enabled the quantitative determination of polar and nonpolar drugs of abuse with log P values ranging from 0.16 to 4.98 in biological matrices while also providing good linearities.

Investigation of Imidazolium-Based Ionic Liquids as Additives for the Separation of Urinary Biogenic Amines via Capillary Electrophoresis

Ionic liquids (ILs), such as imidazoles, can be used to prevent the sorption of analytes onto the walls of the capillary. Prior works have confirmed that coating the capillary wall with a cationic layer can increase its surface stability, thereby improving the repeatability of the separation process. In this study, micellar electrokinetic chromatography (MEKC) is employed to evaluate how two ILs with different anions—namely, 1-hexyl-3-methylimidazolium chloride [HMIM+Cl−] and 1-hexyl-3-methylimidazolium tetrafluoroborate [HMIM+BF4−]—affect the separation efficiency for biogenic amines (BAs) such as metanephrine (M), normetanephrine (NM), vanilmandelic acid (VMA), and homovanillic acid (HVA) in urine samples. To this end, solid-phase extraction (SPE) is employed using different sample pH values, with the results demonstrating that HVA and VMA is easily extracted at a sample pH of 5.5, while a sample pH of 9.0 facilitated the extraction of M and NM. In the applied SPE protocol, selected analytes were isolated from urine samples using hydrophilic–lipophilic-balanced (HLB) columns and eluted with methanol (MeOH). The validation data confirmed the method’s linearity (R2 > 0.996) for all analytes within the range of 0.25–10 µg/mL. The applicability of the optimized SPE-MEKC-UV method was confirmed by employing it to quantify clinically relevant BAs in real urine samples from pediatric neuroblastoma (NBL) patients.

Comparative Study of Various Procedures for Extracting Doxorubicin from Animal Tissue Samples

This article presents a comparative study of selected deproteinization-, liquid–liquid-extraction- (LLE), and solid-phase-extraction (SPE)-based procedures for the isolation of doxorubicin (DOX) and daunorubicin (DAU) as an internal standard (IS) from rat tissue samples. During the experiments, all samples were analyzed via liquid chromatography coupled with fluorescence detection (LC-FL), with analytes being monitored at excitation and emission wavelengths of 487 and 555 nm, respectively. The absolute recoveries of the sample-preparation procedure were then calculated and compared, and the advantages and disadvantages of each approach were considered in depth. Ultimately, SPE with hydrophilic–lipophilic balanced (HLB) sorbents was selected as the most effective extraction procedure as it enabled the absolute recovery of DOX from tissue samples at a level of 91.6 ± 5.1%. Next, the selected HLB-SPE protocol was coupled with LC-FL separation and the resultant method was validated according to FDA and ICH requirements. The validation data confirmed that the developed procedure met all required criteria for bioanalytical methods, with a limit of detection (LOD) and limit of quantification (LOQ) of 0.005 µg/g and 0.01 µg/g, respectively. Finally, the developed protocol was successfully tested on various rat tissues enriched with DOX, confirming its potential as an interesting alternative to previously reported protocols for pharmacokinetic studies and clinical investigations aimed at analysis of the level and biodistribution of DOX in tissue samples after systemic administration of this drug.

Leveraging multi-mode microextraction and liquid chromatography stationary phases for quantitative analysis of neurotoxin β-N-methylamino-L-alanine and other non-proteinogenic amino acids

Effective quantitative analysis of BMAA (β-N-methylamino-L-alanine) and its isomers without the need for derivatization has always been an analytical challenge due to their poor retention and separation on various liquid chromatography stationary phases. Previous studies that utilized conventional hydrophilic interaction chromatography (HILIC) demonstrate false negatives compared to reverse-phase workflows with derivatization. This work evaluates the chromatographic behavior of BMAA and its isomers, in their underivatized forms, on selected stationary phases, in particular fluorophenyl-based columns, to attain effective retention and separation. Detection and quantification were achieved with an ion-trap mass spectrometer. Extraction and preconcentration were achieved via solid phase microextraction (SPME) by assessing the effectiveness of multiple extraction phases, including hydrophilic-lipophilic balanced (HLB) and mixed-mode (MM). A MM extraction phase consisting of C8 and benzene sulfonic acid moieties provided ideal extraction performance for BMAA and its isomers (2,4-diaminobutyric acid, DABA; N-(2-aminoethyl) glycine, AEG). Chromatographic separation was achieved within 8 min on a fluorophenyl stationary phase, ensuring high throughput without derivatization, and showing exceptional improvement from conventional HILIC methods. Limits of quantification in water for BMAA and AEG were 2.5 µg L−1 and DABA was 5 µg L−1, with linear dynamic ranges from 2.5 µg L−1 - 200 µg L−1 for BMAA and AEG and 5 µg L−1 - 200 µg L−1 for DABA.

Evaluation, comparison and combination of molecularly imprinted polymer solid phase extraction and classical solid phase extraction for the preconcentration of endocrine disrupting chemicals from representative whole water samples

Estrogens are endocrine disrupting chemicals and of high concerns due to demonstrated harmful effects on the environment and low effect levels. For monitoring and risk assessment, several estrogens were included in the "watch list" of the EU Water Framework Directive which sets very low environmental quality standard (EQS) levels for Estrone (E1) and 17β-Estradiol (E2) of 0.4 ng L−1 and for 17α-Ethinylestradiol (EE2) of 0.035 ng L−1 requiring sensitive detection methods, as well as extensive sample preparation. A sensitive, derivatization-free, isotope dilution calibration HPLC-MS/MS method for a panel of 5 selected estrogens (including the 3 estrogens of the EU WFD watchlist), and a procedure for the reproducible preparation of a representative whole water matrix including mineral water, humic acids and solid particulate matter are presented. These are used in a diligent comparison of classical solid phase extraction (SPE) on hydrophilic-lipophilic balanced (HLB) phase to SPE on an estrogen-specific molecularly imprinted polymer phase (MISPE) for ultra-trace levels of the analytes (1–10 ng L−1). Additionally, a two-step procedure combining HLB SPE disks followed by MISPE is evaluated. The tow-step procedure provides superior enrichment, matrix removal and sample throughput while maintaining comparable recovery rates to simple cartridge SPE. Estimated method quantification limits (MQLs) range from 0.109–0.184 ng L−1 and thus meet EQS-levels for E1 and E2, but not EE2. The representative whole water matrix provides a reproducible comparison of sample preparation methods and lays the foundation for a certified reference material for estrogen analysis. The presented method will serve as the basis for an extended validation study to assess its use for estrogen monitoring in the environment.

[Determination and level investigation of 45 hormones in edible bird's nest by solid phase extraction-high performance liquid chromatography-tandem mass spectrometry].

建立了固相萃取-高效液相色谱-串联质谱(SPE-HPLC-MS/MS)同时测定食用燕窝中皮质激素、雌激素、雄激素及孕激素等5类45种激素的多残留分析方法。采用乙腈-乙酸乙酯(1∶1, v/v)超声辅助提取、亲水亲脂平衡固相萃取柱净化,甲醇洗脱。分别在正、负电喷雾电离源、多反应监测模式下检测45种激素。正离子模式下的流动相为乙腈-0.1%甲酸水溶液,负离子模式下的流动相为乙腈-水,色谱柱为Phenomenex Kinetex C18柱(100 mm×2.1 mm, 2.6 μm)。在优化条件下,45种激素在0.20~20.0 μg/L范围内线性关系良好,相关系数(R2)≥0.9990,方法的检出限(LOD)为0.04~0.70 μg/kg,定量限(LOQ)为0.16~2.00 μg/kg。按三水平(2.0、4.0、20.0 μg/kg)进行加标回收试验,氟米龙、布地奈德、醛固酮、醋酸氟轻松、炔雌醇的回收率为40.2%~63.6%,可对这5种激素进行定性分析,其余40种激素的平均加标回收率为72.2%~112.3%,相对标准偏差(RSD)为2.5%~11.6%,该方法可对40种激素进行准确定性定量,精密度好,灵敏度高,简便、快速。从2017~2021年,通过研究建立的方法对来自马来西亚、印度尼西亚、泰国和越南等国家的1021个燕窝样品进行监测,仅勃地酮、雄烯二酮、孕酮有检出(大于检出限),其他激素均小于检出限。孕酮检出率为100%,勃地酮、雄烯二酮检出率分别为79%和89%, 3种激素含量范围分别为0.097~3.58、0~0.096和0~1.77 μg/kg。与同为动物源性食品的鸡蛋、纯牛奶、乳制品相比,所有测定的鸡蛋样品中均检出雄烯二酮,含量比其他3类产品略高;勃地酮在4类产品中的含量差别不大,均为微量;孕酮含量在鸡蛋中最高,其次是纯牛奶,燕窝中含量最低。研究结果表明,食用燕窝带入的激素种类少,含量低,对健康影响小。

Profiling of Unsaturated Lipids by Raman Spectroscopy Directly on Solid-Phase Microextraction Probes.

Lipids play a critical role in cellular signaling, energy storage, and the construction of cellular membranes. In this paper, we propose a novel on-site approach for detecting and differentiating enriched unsaturated lipids based on the direct coupling of SPME probes with Raman spectroscopy. To this end, different SPME particles, namely, hydrophilic-lipophilic balanced (HLB), mixed-mode (C8-SCX), and C18, were embedded in polyacrylonitrile (PAN) and tested for their efficacy as biocompatible coatings. The C18/PAN coating showed less background interference compared to the other sorbent materials during the analysis of unsaturated lipids. In addition, different SPME parameters that influence extraction efficiency, such as extraction temperature, extraction time, and washing solvent, were also investigated. Our results indicate a clear dependence between the Raman band intensity related to the number of double bonds in fatty acids mixture and the number of double bonds in a fatty acid. Our findings further show that Raman spectroscopy is especially useful for the analysis of lipid unsaturation, which is calculated as the ratio of n(C═C)/n(CH2) using the intensities of the Raman bands at 1655/1445 cm-1. Furthermore, the developed protocol reveals great SPME activity and high detection ability for several unsaturated lipids in different complex matrixes, such as cod liver oil. Finally, the applicability of this technology was demonstrated via the characterization of cod liver oil and other vegetable oils. Thus, the proposed SPME-Raman spectroscopy approach has a great future potential in food, environmental, clinical, and biological applications.

Haloquinone Chloroimides as Toxic Disinfection Byproducts Identified in Drinking Water.

Haloquinone chloroimides (HQCs) are suspected to be highly toxic contaminants, and their production during drinking water disinfection is predicted. However, HQC disinfection byproducts (DBPs) have not been reported in drinking water to date because of analytical limitations. In this study, we developed an analytical method to detect five HQCs, including 2,6-dichloroquinone-4-chloroimide (2,6-DCQC), 2,6-dibromoquinone-4-chloroimide (2,6-DBQC), 2-chloroquinone-4-chloroimide (2-CQC), 3-chloroquinone-4-chloroimide (3-CQC), and 2,6-dichloroquinone-3-methyl-chloroimide (2,6-DCMQC). This method combined a derivatization reaction of HQCs with phenol in alkaline solutions to produce halogenated indophenols, a solid-phase extraction pretreatment using hydrophilic-lipophilic balanced (HLB) cartridges, and a multiple reaction monitoring (MRM) method for quantification. The method was demonstrated to be sensitive and accurate with recoveries of 71-85% and limits of detection of 0.1-0.2 ng/L for the five tested HQCs. Using this method, five tested HQCs were identified in drinking water samples from nine water treatment plants and water distribution systems as new DBPs at concentrations of up to 23.1 ng/L. The cytotoxicity of the five tested HQCs in HepG2 cells was higher than or comparable to that of 2,6-dichloro-1,4-benzoquinone (2,6-DCBQ), an emerging DBP that was hundreds to thousands of times more toxic than regulated DBPs. This study presents the first analytical method for HQC DBPs in drinking water and the first set of occurrence and cytotoxicity data of HQC DBPs.

Evaluation of the selectivity of molecularly imprinted polymer cartridges for nitroimidazoles. Application to the simultaneous extraction of nitroimidazoles and benzimidazoles from samples of animal origin

In this work the selectivity of commercial molecularly imprinted polymer (MIP) cartridges for nitroimidazoles has been evaluated. For this, a sample treatment based on molecularly imprinted solid phase extraction (MISPE) has been developed and applied to the simultaneous extraction of nitroimidazoles and benzimidazoles. These cartridges were compared with other sorbents commonly used for the solid phase extraction (SPE) of veterinary drugs such as C18, hydrophilic/lipophilic balanced (HLB), and strong cation exchange (SCX). Analysis of the extracts was carried out by HPLC-MS/MS. The use of MIP cartridges provided good results for all nitroimidazoles and for most of the benzimidazoles studied (except for the most apolar analytes), both in terms of recovery and precision and in the cleaning of the extracts. Thus, although MIPs have been designed to specifically bind to a target molecule, i.e., nitroimidazoles, they are able to extract other analytes, such as benzimidazoles, with high efficiency. The developed method was applied to the determination of four nitroimidazoles and ten benzimidazoles in samples of animal origin. All nitroimidazoles and eight of the benzimidazoles were successfully determined. Validation of the method afforded values in the range of 91–111% for the recovery studies. The analysis of intra-day and inter-day precision showed relative standard deviation values lower than 9.5 % and 12 % respectively. These results indicate that the proposed method is suitable for the determination of nitroimidazoles and benzimidazoles in samples of animal origin.

A modified method of on-line solid phase extraction and fluorometric detection for underway monitoring and onboard analysis of trace ammonium in seawater

A sensitive and automated method for underway and onboard analysis of trace ammonium in seawater was established using on-line solid phase extraction and fluorescence detection. Ammonium reacted with o-phthalaldehyde and sulfite to produce isoindole derivative that was concentrated on a hydrophilic lipophilic balanced (HLB) cartridge and then rapidly eluted with ethanol and measured using a fluorescence detector. The work was modified from our previous developed method, where 30 mL of a seawater sample was introduced into a reaction bottle with a 10 mL pipette. In the modified method a 10 mL sample loop was adopted for sample introduction to reduce possible contamination caused by manual operation. More importantly, the present method makes underway monitoring much more automatic. The detection limit was 3.5 nmol L−1, and the relative standard deviation was 3.52% (n = 12) for a sample spiked with 40 nmol L−1 ammonium standard. The method was free of the effects of salinity, carry-over and dissolved organic matter, with recoveries between 94.0% and 106.1%. An equation was introduced to correct matrix effects, and pure water and seawater blank were found to be equivalent to ammonium concentration of 8.1 ± 2.2 nmol L−1 and 6.0 ± 0.8 nmol L−1, respectively. The correction method can also be introduced to determination of other trace analytes where matrix effect cannot be neglected. A technique of stopped-flow was adopted and stopped-flow time could be adjusted depending on the concentration of ammonium in the seawater samples. The sample throughput was 7–9 h−1, and the method permitted the analysis of samples over a wide range of concentrations. The modified method has been successfully applied for shipboard underway determination of ammonium in a coastal area and shipboard determination the ammonium concentration of discrete samples in three vertical profiles during a cruise in the South China Sea.

A multi-family offline SPE LC-MS/MS analytical method for anionic, cationic and non-ionic surfactants quantification in surface water

Of the large number of emerging pollutants discharged from wastewaters into surface waters, surfactants are among those with the highest concentrations. However, few monitoring in river waters of these substances have already been performed and only on a few families, mostly anionic.This work aimed to develop a multi-family analytical strategy suitable for the quantification of low concentrations of surfactant in surface waters. Twelve families of surfactants, anionic, cationic and non-ionic were selected. Their quantification by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) and their extraction by SPE were optimized by comparing different retention mechanisms. The best performances were obtained with a C18 grafted silica LC column and a hydrophilic-lipophilic balanced (HLB) polymeric SPE cartridge.The final analytical method was validated and applied for the quantification of surfactants in 36 river water samples. Method limits of quantification (LQ), intra and inter days precision and trueness were evaluated. With LQ between 15 and 485 ng/L, and trueness over 80%, this method was suitable for monitoring surfactants in surface water. Application on French river water samples revealed the presence of anionic, cationic and non-ionic surfactants with median concentrations from 24 ng/L for octylphenol ethoxylates (OPEO), up to 4.6 μg/L regarding linear alkylbenzene sulfonates (LAS).

Development of Europium-Sensitized Fluorescence-Based Method for Sensitive Detection of Oxytetracycline in Citrus Tissues.

Antimicrobial compounds have been successfully used to control many plant and animal diseases. Recently, oxytetracycline (OTC) and streptomycin have been approved for the treatment of Huanglongbing in citrus. Since the application of OTC is under strict regulations, several methods have been developed to determine and monitor its levels in the environment including high-performance liquid chromatography, ELISA, colorimetric, and fluorometric assays. In this study, we developed a fluorometric method for the determination of OTC in plant tissues based on its complexation with europium. Our preliminary trials showed that phenols and flavonoids interfere with the europium assay by reacting with the sensitizing reagent, cetyltrimethylammonium chloride. Consequently, we used the 60 mg hydrophilic–lipophilic balanced (HLB) cartridges to purify the OTC from the plant matrix. The recovery of OTC from spiked leaf samples was 75 ± 7.6%. Using the 500 mg HLB, we were able to detect 0.3 ppm OTC in the final sample extract, which corresponds to 3 µg g−1 fresh weight (FWT). The developed method was successfully used to measure the level of OTC in leaves obtained from trunk-injected trees. The results obtained by the europium method were similar to those obtained using the ELISA assay. We also tested the cross-reactivity of OTC metabolites with the europium method. The 4-epi-OTC showed a high cross-reactivity (50.0 ± 3.6%) with europium assay, whereas α-apo-OTC and β-apo-OTC showed small cross-reactivity. We showed that the europium-sensitized fluorescence-based method can be successfully used to assess OTC in citrus plant tissues after a cleanup step. Our results showed that this method was sensitive, reproducible, and can be used to analyze many samples simultaneously.

Assessment of solid phase microextraction as a sample preparation tool for untargeted analysis of brain tissue using liquid chromatography-mass spectrometry

This work presents an evaluation of solid-phase microextraction (SPME) SPME in combination with liquid chromatography-high resolution mass spectrometry (LC-HRMS) as an analytical approach for untargeted brain analysis. The study included a characterization of the metabolite coverage provided by C18, mixed-mode (MM, with benzene sulfonic acid and C18 functionalities), and hydrophilic lipophilic balanced (HLB) particles as sorbents in SPME coatings after extraction from cow brain homogenate at static conditions. The effects of desorption solvent, extraction time, and chromatographic modes on the metabolite features detected were investigated. Method precision and absolute matrix effects were also assessed. Among the main findings of this work, it was observed that all three tested coating chemistries were able to provide comparable brain tissue information. HLB provided higher responses for polar metabolites; however, as these fibers were prepared in-house, higher inter-fiber relative standard deviations were also observed. C18 and HLB coatings offered similar responses with respect to lipid-related features, whereas MM and C18 provided the best results in terms of method precision. Our results also showed that the use of methanol is essential for effective desorption of non-polar metabolites. Using a reversed-phase chromatographic method, an average of 800 and 1200 brain metabolite features detected in positive and negative modes, respectively, met inter-fibre RSD values below 30% (n=4) after removal of fibre and solvent artefacts from the associated datasets. For features detected using a lipidomics method, a total of 900 and 1800 features detected using C18 fibers in positive and negative mode, respectively, met the same criteria. In terms of absolute matrix effects, the majority of the model metabolites tested showed values between 80 and 120%, which are within the acceptable range. Overall, the findings of this work lay the foundation for further optimization of parameters for SPME-LC-HRMS methods suitable for in vivo and ex vivo brain (and other tissue) untargeted studies, and support the applicability of this approach for non-destructive tissue metabolomics.

Method validation, persistence, and safety evaluation of 2,4-D in tea (Camellia sinensis) by LC-MS/MS.

An analytical method was developed by using LC-ESI(-ve)-MS/MS to investigate the residue dynamics of 2,4-D (2,4-dichlorophenoxyacetic acid) in green tea leaves, processed tea, tea liquor, and tea-cropped soil at Singhiajhora Tea Estate and Putinbari Tea Estate at Terai Region, Darjeeling District, West Bengal, India. In this method, an acidified methanol was used for extraction and subsequent clean-up was done by HLB (hydrophilic lipophilic balanced) cartridges. The method was validated as per SANTE guideline (SANTE/11813/2017). The limit of quantification (LOQ) of 2,4-D was 0.05 mgkg-1 and average % recoveries were in the range from 88.05 to 113.28 with relative standard deviation (RSD) 3.46 to 6.43. The dissipation of 2,4-D followed the 1st-order reaction kinetics with a half-life (T1/2) of 1.51-1.61 day at the recommended dose and 2.50-2.72day for doubled recommended dose in tea for both locations. This method can be applied successfully for the determination of 2,4 D residues in/on tea matrix and subsequent studies on safety evaluation showed that the use of 2,4-D in tea is safe.

Development and Validation of a Multi-detection Confirmatory Method for Antibiotics Determination in Piglet Kidneys by UHPLC-TOF-MS According Commission Decision 2002/657/EC

The combination of ultra-high-performance liquid chromatography (UHPLC) with time-of-flight mass spectrometry (ToF-MS) is one of the most powerful analytical tools in terms of selectivity, sensitivity and speed. However, due to the gap on regulation and guidelines concerning the validation criteria for the use of ToF-MS methodology not only for screening but also for confirmatory and quantification purposes, this study presents, for the first time, an approach on how to use the referred methodology for confirmatory and quantification purposes. Thus, an UHPLC-ToF-MS method was developed and validated for the simultaneous determination of 43 antibiotic residues from 7 different families (tetracyclines, sulfonamides, quinolones, penicillins, macrolides, cephalosporins and trimethoprim) in piglet kidneys as a confirmatory method. A simple sample preparation consisting of extraction with a mixture of acetonitrile and ethylenediaminetetraacetic acid (EDTA) followed by solid-phase extraction (SPE) with a hydrophilic-lipophilic balanced (HLB) cartridge and finalized with a defatting step with n-hexane was used. The methodology was validated according the Commission Decision 2002/657/EC, by defining suitable criteria for antibiotic identification, as well as by evaluating required parameters like selectivity, specificity and ruggedness that fulfil the required criteria. The calculated values of decision limit (CCα) and of detection capacity (CCβ) for antibiotics with established maximum residue limits (MRLs) fully complied with required criteria. For antibiotics for which no MRL was set, values ranging from 0.2 to 0.5 μg kg−1 and from 0.5 to 0.8 μg kg−1 were achieved for CCα and for CCβ, respectively. Linearity (all R2 > 0.95), recovery (81.4–110.0%) (lowest values of 4.6% and 7.1% were obtained for repeatability and for reproducibility, respectively), LODs (0.04–13.0 μg kg−1), LOQs (0.09–32.0 μg kg−1) and matrix effects (35.5–118.1 %) were also evaluated. After validation, the method was applied to analyse piglet kidney samples. Antibiotic residues were determined in 10 of the 15 analysed samples. However, all the antibiotics concentrations were below the MRLs.

A solid-phase extraction method that eliminates matrix effects of complex pulp mill effluents for the analysis of lipophilic wood extractives

Abstract The extraction of lipophilic wood extractives from pulp and paper process waters proves to be a challenging task, due to harsh and alternating process and sample conditions. This study has determined the potential use of polymeric sorbents for solid-phase extraction (SPE) and compared to classical silica-based reversed-phase packed columns, with polymeric hydrophilic-lipophilic balanced (HLB) cartridges being the sorbent with the most potential. Recovery functions were obtained with an internal standard mixture representative for the main lipophilic wood extractive groups, which are fatty acids and alcohols, sterols, sterol esters and triglycerides. The impact of pH, sample volume and sample matrix, expressed as TOC and cations, on the retention behavior of lipophilic extractives during SPE of industrial samples were determined with polymeric HLB sorbent. High variations in the composition of pulp mill matrices led to different optimal extraction conditions. Thus, a new SPE protocol was developed, which bypasses matrix interferences and omits the loss of analytes due to sample preparation. The method is applicable to different pulp mill effluents with large discrepancies in pH and sample matrices, resulting in recoveries >90 % with RSD <5 % for all lipophilic wood extractives.