Quantification of cresols in liquid smoke samples employing liquid-liquid extraction with low-temperature purification and analysis by gas chromatography-mass spectrometry.

Liquid smoke is a food additive and cresols are among its chemical constituents, potentially toxic to human health. Thus, the objective of this study was to develop a method to quantify cresols in liquid smoke. First, the liquid-liquid extraction with low temperature purification (LLE-LTP) was validated for cresols in water, as there are no cresol-free liquid smoke samples. Analyzes were performed by gas chromatography coupled to mass spectrometry in full scan mode. LLE-LTP was subsequently applied in five commercial samples of liquid smoke. Validation results showed that the proposed extraction method was selective for cresols, linear in the range of 0.5 to 35 mg L-1, limit of quantification of 0.5 mg L-1, recovery rate between 90% and 104% and relative standard deviation lower than 10%. The quantification of cresols in liquid smoke samples ranged from 3.0 to 38.3 mg L-1 and the concentration of these chemical contaminants in liquid smoke remained constant for at least 21 days at 25 °C.

Characterizing lipid constituents of B. moojeni snake venom: a comparative approach for chemical and biological investigations.

Snake venoms are complex mixtures majorly composed of proteins with well-studied biological effects. However, the exploration of non-protein components, especially lipids, remains limited despite their potential for discovering bioactive molecules. This study compares three liquid-liquid lipid extraction methods for both chemical and biological analyses of Bothrops moojeni snake venom. The methods evaluated include the Bligh and Dyer method (methanol, chloroform, water), considered standard; the Acunha method, a modification of the Bligh and Dyer protocol; and the Matyash method (MTBE/methanol/water), featuring an organic phase less dense than the aqueous phase. Lipidomic analysis using liquid chromatography with high-resolution mass spectrometry (LC-HRMS) system revealed comparable values of lipid constituents' peak intensity across different extraction methods. Our results show that all methods effectively extracted a similar quantity of lipid species, yielding approximately 17-18 subclasses per method. However, the Matyash and Acunha methods exhibited notably higher proportions of biologically active lipids compared to the Bligh and Dyer method, particularly in extracting lipid species crucial for cellular structure and function, such as sphingomyelins and phosphatidylinositol-phosphate. In conclusion, when selecting a lipid extraction method, it is essential to consider the study's objectives. For a biological approach, it is crucial to evaluate not only the total quantity of extracted lipids but also their quality and biological activity. The Matyash and Acunha methods show promise in this regard, potentially offering a superior option for extracting biologically active lipids compared to the Bligh and Dyer method.

Automated preparation of plasma lipids, metabolites, and proteins for LC/MS-based analysis of a high-fat diet in mice.

Blood plasma is one of the most commonly analyzed and easily accessible biological samples. Here, we describe an automated liquid-liquid extraction (LLE) platform that generates accurate, precise, and reproducible samples for metabolomic, lipidomic, and proteomic analyses from a single aliquot of plasma while minimizing hands-on time and avoiding contamination from plasticware. We applied mass spectrometry to examine the metabolome, lipidome, and proteome of 90 plasma samples to determine the effects of age, time of day, and a high-fat diet in mice. From 25 µL of mouse plasma, we identified 907 lipid species from 16 different lipid classes and subclasses, 233 polar metabolites, and 344 proteins. We found that the high-fat diet induced only mild changes in the polar metabolome, upregulated Apolipoproteins, and induced substantial shifts in the lipidome, including a significant increase in arachidonic acid (AA) and a decrease in eicosapentaenoic acid (EPA) content across all lipid classes.

Determining pyroxasulfone herbicide in honey samples using liquid-liquid extraction with low temperature purification (LLE-LTP).

Pyroxasulfone is a selective, systemic, pre-emergence herbicide which acts to inhibit weeds in potato, coffee, sugar cane, eucalyptus, and soybean plantations, among others. This active ingredient was classified by Brazilian legislation as a very dangerous product for the environment, and to date there are no studies involving the development of extraction methods for monitoring this compound in environmental matrices. Therefore, the objective of this study was to optimize and validate liquid-liquid extraction with low temperature purification followed by a gas chromatography coupled to mass spectrometry analysis to determine this herbicide in honey samples. The results showed that the best extractor phase was acetonitrile and ethyl acetate (6.5 mL:1.5 mL), with recovery rates close to 100% and relative standard deviations below 11%. The validation proved that the extraction method was selective, precise, accurate and linear in the range of 3-225 µg kg-1, reaching a limit of quantification of 3 µg kg-1, with a -25.95% matrix effect. Monitoring on real samples did not reveal episodes of environmental contamination with pyroxasulfone residue.

Rapid determination of 61 acid dyes in chili paste, hotpot seasoning, and bearnaise using double liquid-liquid extraction and UHPLC-Q-Orbitrap-MS.

The aims of this study were to establish a novel method for simultaneously determining 61 acid dyes in chili, hotpot seasoning, and bearnaise sauce using double liquid-liquid extraction (d-LLE) technology. A mixture of water, methanol, and dichloromethane (1:3:1, v/v/v) was used as the extraction solution, which was actively separated into aqueous and organic phases at a fixed ratio. The clean-up step was initially completed by discarding the organic phase layer, which contained abundant lipophilic compounds. Subsequently, the aqueous phase was further separated by salting out, which effectively removed interference from the highly hydrophilic compounds. As a result of these two purification steps, the matrix suppression effect was significantly reduced by a minimum of 16.9%. Finally, the extract was analyzed using an ultrahigh-performance liquid chromatography-quadrupole Orbitrap mass spectrometer (UHPLC-Q-Orbitrap-MS), and the characteristic ion fragments (SO3 - , m/z 79.9557) of the acid dyes were utilized for the preliminary qualitative analysis. The results showed that the 61 acid dyes showed a good linear relationship in the range of 0.01-0.2 µg/mL, and the limit of quantification (LOQ) was 0.01 mg/kg. The average recoveries were 74.3%-99.7%, with relative standard deviations (RSD) ≤10%. The proposed method can rapidly identify and quantify acid dyes in complex foods at a low cost, with high sensitivity and reliability.

Combining surface-enhanced Raman spectroscopy and paper spray mass spectrometry for the identification and confirmation of psychotropic substances in alcoholic beverages.

Criminal practices in which an individual becomes vulnerable and prone to sexual assault after ingesting drinks spiked with doping substances have become a social concern globally. As forensic protocols require a multi-tiered strategy for chemical evidentiary analysis, the backlog of evidence has become a significant problem in the community. Herein, a fast, sensible, and complementary dual analytical methodology was developed using a single commercial paper substrate for surface-enhanced Raman spectroscopy (SERS) and paper spray mass spectrometry (PS-MS) analysis to identify psychotropic substances added to alcoholic beverages irrefutably. To study and investigate this criminal practice, pharmaceutical formulations containing distinct psychotropic substances (zolpidem, clonazepam, diazepam, and ketamine) were added to drinks typically consumed at parties and festivals (Pilsen beer, açaí Catuaba®, gin tonic, and vodka mixed with Coca-Cola Zero®). A simple liquid-liquid extraction with a low-temperature partitioning (LLE-LTP) procedure was applied to the drinks and effectively minimized matrix effects. As a preliminary analysis, SERS spectra combined with Hierarchical Clustering Analysis (HCA) provided sufficient information to investigate the samples further. The presence of the protonated species for the psychotropic substances in the spiked drinks was readily verified in the mass spectra and confirmed by tandem mass spectrometry. Finally, the results demonstrate the potential of this methodology to be easily implemented into the routine of forensic laboratories and to be further employed at harm reduction tends at parties and festivals to detect contaminated beverages promptly and irrefutably as an efficient tool to prevent such crimes.

Using the Intrinsic Geometry of Binodal Curves to Simplify the Computation of Ternary Liquid-Liquid Phase Diagrams.

Phase diagrams are powerful tools to understand the multi-scale behaviour of complex systems. Yet, their determination requires in practice both experiments and computations, which quickly becomes a daunting task. Here, we propose a geometrical approach to simplify the numerical computation of liquid-liquid ternary phase diagrams. We show that using the intrinsic geometry of the binodal curve, it is possible to formulate the problem as a simple set of ordinary differential equations in an extended 4D space. Consequently, if the thermodynamic potential, such as Gibbs free energy, is known from an experimental data set, the whole phase diagram, including the spinodal curve, can be easily computed. We showcase this approach on four ternary liquid-liquid diagrams, with different topological properties, using a modified Flory-Huggins model. We demonstrate that our method leads to similar or better results comparing those obtained with other methods, but with a much simpler procedure. Acknowledging and using the intrinsic geometry of phase diagrams thus appears as a promising way to further develop the computation of multiphase diagrams.

Volatile Compound Release from Oak Chips in Model Wine Media: Combined Influence of Toasting Degree, Size, Time of Contact, and Ethanol Content.

The effects of size, toasting degree, and time of contact on the release of volatile compounds from Quercus alba (L.) chips during a simulated fermentation and post-fermentative process were studied. The results obtained indicated that the large-size chips favored the release of furfural and furfuryl alcohol, while the small ones increased the concentration of cyclotene and maltol. The interaction between chip size and time of contact showed that the small-size chips are more sensitive to the increase of ethanol concentration for the extraction rate of some compounds (furfural, vanillin, maltol, cyclotene, whiskey lactones, and eugenol) compared to the large-size ones, increasing their concentrations at the end of maceration. The toasting degree of oak chips had a different influence on the volatile compounds studied. Cyclotene and guaiacol concentrations increased with the toasting intensity, whereas the extracted concentration of all compounds increased from light to medium-toasted chips, except for eugenol, and then decreased by further increasing the toasting level for 5-methylfurfural, whiskey lactones, eugenol, and only using high-level toasted chips for furfuryl alcohol, maltol, and vanillin. A possible protection effect of the chip size toward the possible degradation or volatilization losses of furfural for high toasting degrees was observed.

GC vs. HPLC in quantitation of CBD, CBG, ∆9-THC and CBN in plasma using different sample preparation methods.

The sensitivity of complex analytical procedures depends not only on the sensitivity of the analytical instrument used, but also on the recovery degree of the examined analyte by the employed sample preparation method. The recovery degrees of individual cannabinoids reported in literature, estimated using the same sample preparation method, are unexpectedly divergent. Therefore, the aim of this study was a thorough assessment of the most commonly used sample preparation methods, such as protein precipitation, LLE, QuEChERS and SPE, in the context of the reliability of the obtained results. The presented report shows that the highest sensitivity, precision and reliability of the chromatographic analysis of CBG, CBD, ∆9-THC and CBN in human plasma can be obtained using SPE. The recovery degrees of these cannabinoids by SPE are highly repeatable and exceed 95 %, while they are significantly lower for such sample preparation methods as protein precipitation, LLE and QuEChERS (ca. 80, 65 and 87, respectively). Moreover, the supernatants obtained by the latter methods contain interferents evoking matrix-effect, which makes reliable quantification of the listed cannabinoids by GC difficult. To our knowledge, the paper is the first such extensive comparison of sample preparation procedures used for the determination of cannabinoids in plasma by GC-MS and HPLC-MS. The presented results and the discussion allow to understand why different recovery degrees for the same xenobiotic can be find in literature despite they have been estimated using the same or different sample preparation method or different chromatography types.

The Selective Separation of Carnosic Acid and Rosmarinic Acid by Solid-Phase Extraction and Liquid-Liquid Extraction: A Comparative Study.

Rosmarinus officinalis leaves (ROLs) are widely used in the food and cosmetics industries due to their high antioxidant activity and fascinating flavor properties. Carnosic acid (CA) and rosmarinic acid (RA) are regarded as the characteristic antioxidant components of ROLs, and the selective separation of CA and RA remains a significant challenge. In this work, the feasibility of achieving the selective separation of CA and RA from ROLs by solid-phase extraction (SPE) and liquid-liquid extraction (LLE) was studied and compared. The experiments suggested that SPE with CAD-40 macroporous resin as the adsorbent was a good choice for selectively isolating CA from the extracts of ROLs and could produce raw CA with purity levels as high as 76.5%. The LLE with ethyl acetate (EA) as the extraction solvent was more suitable for extracting RA from the diluted extracts of ROLs and could produce raw RA with a purity level of 56.3%. Compared with the reported column chromatography and LLE techniques, the developed SPE-LLE method not only exhibited higher extraction efficiency for CA and RA, but can also produce CA and RA with higher purity.

Extraction method for determining dinotefuran insecticide in water samples.

Dinotefuran is a compound belonging to the third generation of nicotinoid insecticides, and has been effective in combating pests that are resistant to conventional insecticides, such as organophosphates, carbamates, and pyrethroids. This molecule presents high-water solubility (39,830 mg L-1 at 25 °C) compared to other pesticides, which facilitates its drag and leaching to lower soil layers. Therefore, the present study aimed to optimize and validate liquid-liquid extraction with low temperature purification (LLE-LTP) to determine dinotefuran residues in water by high performance liquid chromatography with diode array detection (HPLC-DAD). The results revealed that the analyte recovery ranged from 85.44 to 89.72% with a relative standard deviation <5.8. LLE-LTP was selective, precise, accurate, and linear in the range from 10.0 to 210 µg L-1, and presented limits of detection and quantification of 5.00 and 10.00 µg L-1, respectively. The matrix effect was <14%. The stability study of dinotefuran in water revealed significant stability of this molecule in water in the absence of light (>130 days), and a half-life of 7 days in water with sunlight. LLE-LTP coupled to HPLC-DAD was a simple, easy, and efficient method for extracting and analyzing dinotefuran in water samples.

Pre-analytical sample handling standardization for reliable measurement of metabolites and lipids in LC-MS-based clinical research.

The emerging disciplines of lipidomics and metabolomics show great potential for the discovery of diagnostic biomarkers, but appropriate pre-analytical sample-handling procedures are critical because several analytes are prone to ex vivo distortions during sample collection. To test how the intermediate storage temperature and storage period of plasma samples from K3EDTA whole-blood collection tubes affect analyte concentrations, we assessed samples from non-fasting healthy volunteers (n = 9) for a broad spectrum of metabolites, including lipids and lipid mediators, using a well-established LC-MS-based platform. We used a fold change-based approach as a relative measure of analyte stability to evaluate 489 analytes, employing a combination of targeted LC-MS/MS and LC-HRMS screening. The concentrations of many analytes were found to be reliable, often justifying less strict sample handling; however, certain analytes were unstable, supporting the need for meticulous processing. We make four data-driven recommendations for sample-handling protocols with varying degrees of stringency, based on the maximum number of analytes and the feasibility of routine clinical implementation. These protocols also enable the simple evaluation of biomarker candidates based on their analyte-specific vulnerability to ex vivo distortions. In summary, pre-analytical sample handling has a major effect on the suitability of certain metabolites as biomarkers, including several lipids and lipid mediators. Our sample-handling recommendations will increase the reliability and quality of samples when such metabolites are necessary for routine clinical diagnosis.

A miniaturized liquid-liquid extraction method for further Na, K, Ca, and Mg determination in crude oil by FAAS.

In this study, a miniaturized liquid-liquid extraction (LLE) method for pre-concentration of Na, K, Ca, and Mg in crude oil was proposed. Analytes in crude oil were quantitatively extracted to the aqueous phase, followed by flame atomic absorption spectrometry (FAAS) determination. The following parameters were evaluated: type of extraction solution, sample mass, heating temperature and time, stirring time, centrifugation time, and the use of toluene and chemical demulsifier. Accuracy was evaluated by comparing the results obtained by the proposed method (LLE-FAAS) with those obtained after high-pressure microwave-assisted wet digestion and FAAS determination (reference values). No statistical difference was observed between the reference values and those using the optimized conditions for LLE-FAAS: 2.5 g of sample; 1000 µL of 2 mol L-1 HNO3, 50 mg L-1 of chemical demulsifier in 500 µL of toluene, 10 min of heating at 80 °C, 60 s of stirring, and 10 min of centrifugation. Relative standard deviations were lower than 6%. The limits of quantification (LOQ) were 1.2, 1.5, 5.0, and 0.50 µg g-1 for Na, K, Ca, and Mg, respectively. The proposed miniaturized LLE method presents several advantages, such as ease-of-use, high throughput (up 10 samples can be processed per 1 h), uses a high sample mass reaching low LOQs. In addition, the use of a diluted solution for extraction reduces the amount of reagents (around 40 times) and consequently laboratory residue generation, becoming an environmental friendly method. Suitable LOQs were achieved for analyte determination at low concentration even using a simple and low-cost sample preparation system (miniaturized LLE method) and a relatively low-cost determination technique (FAAS), avoiding the use of microwave ovens and more sensitivity techniques, which are required for routine analyses.

A novel fully-automated method to measure steroids in serum by liquid chromatography-tandem mass spectrometry.

Background: Steroids play a key role in numerous physiological processes. Steroid determination is a useful tool to explore various endocrine diseases. Because of its specificity, mass spectrometry is considered to be a reference method for the determination of steroids in serum compared to radioimmunoassay. This technology could progress towards more automation for the optimal organization of clinical laboratories and ultimately for the benefit of patients. Methods: A fully automated ultra-high-performance liquid chromatography-tandem mass spectrometry method was developed and fully validated to determine five steroids in serum. Sample preparation was based on protein precipitation with filtration followed by online solid phase extraction. Chromatographic separation was performed using a biphenyl stationary phase. Results: The method was successfully validated according to European Medicine Agency guidelines. Coefficients of variation did not exceed, respectively, 8.4% and 8.1% for intra- and inter-assay precision. Method comparison with radioimmunoassay showed a proportional bias for all compounds, except for testosterone in men. Comparison with another LC-MS/MS method demonstrated acceptable concordance for all steroids, although a small bias was observed for androstenedione. Conclusion: The novelty of this method is that it has been fully automated. Automation provides benefits in traceability and allows significant savings in cost and time.

Critical Evaluation of Analytical Methods for the Determination of Anthropogenic Organic Contaminants in Edible Oils: An Overview of the Last Five Years.

Anthropogenic contaminants, as pesticides, polycyclic aromatic hydrocarbons (PAHs) and monochloropropanediols (MCPDs), have become important to be controlled in edible oils, since their regular occurrence. In fact, alerts from the Rapid Alert System for Food and Feed (RASFF) in oils normally include these compounds. From a critical point of view, tools used to control these compounds in the last 5 years will be discussed, including sample preparation, analysis and current regulations. Extraction and analysis methods will be discussed next, being liquid-liquid extraction (LLE) and QuEChERS, with or without clean-up step, as well as chromatographic methods coupled to different analyzers (mainly mass spectrometry), the most commonly used for extraction and analysis respectively. Occurrence in samples will also be reviewed and compared with the legal maximum residue limits (MRLs), observing that 4%, 20% and 60% of the analyzed samples exceed the legal limits for pesticides, MCPDs and PAHs respectively.

Quantitative Analysis of Decoquinate Residues in Hen Eggs through Derivatization-Gas Chromatography Tandem Mass Spectrometry.

A novel precolumn derivatization-gas chromatography tandem mass spectrometry (GC-MS/MS) method was developed to detect and confirm the presence of decoquinate residues in eggs (whole egg, albumen and yolk). Liquid-liquid extraction (LLE) and solid phase extraction (SPE) were used to extract and purify samples. The derivatization reagents were pyridine and acetic anhydride, and the derivatives were subjected to GC-MS/MS detection. After the experimental conditions were optimized, satisfactory sensitivity was obtained. The limits of detection (LODs) and limits of quantification (LOQs) for the decoquinate in eggs (whole egg, albumen and yolk) were 1.4-2.4 µg/kg and 2.1-4.9 µg/kg, respectively. At four spiked concentration levels, the average recoveries were 74.3-89.8%, the intraday RSDs ranged from 1.22% to 4.78%, and the inter-day RSDs ranged from 1.61% to 7.54%. The feasibility and practicality of the method were confirmed by testing egg samples from a local supermarket.

Separation and Detection of Abamectin, Ivermectin, Albendazole and Three Metabolites in Eggs Using Reversed-Phase HPLC Coupled with a Photo Diode Array Detector.

An innovative and sensitive approach using high-performance liquid chromatography-photo diode array detection (HPLC-PDAD) was developed and optimized for the simultaneous determination of abamectin (ABM), ivermectin (IVM), albendazole (ABZ) and three metabolites in eggs. The samples were extracted with acetonitrile (MeCN)/water (90:10, v/v), and the extracts containing the targets were cleaned up and concentrated by a series of liquid-liquid extraction (LLE) steps. A reversed-phase C18 column and a mobile phase consisting of 0.1% trifluoroacetic acid (TFA) aqueous solution and methanol (MeOH) were utilized to perform optimal chromatographic separation. The developed method was validated on the basis of international guidelines. The limits of detection (LODs) and quantitation (LOQs) were 2.1-10.5 µg/kg and 7.8-28.4 µg/kg, respectively. Satisfactory linear relationships were observed for the targets in their corresponding concentration ranges. The mean recoveries ranged from 85.7% to 97.21% at 4 addition levels, with intraday and interday relative standard deviations (RSDs) in the ranges of 1.68-4.77% and 1.74-5.31%, respectively. The presented protocol was demonstrated to be applicable and reliable by being applied for the detection of target residues in locally sourced egg samples.

GC-MS/MS Determination of Steroid Hormones in Urine Using Solid-Phase Derivatization as an Alternative to Conventional Methods.

Solid-phase analytical derivatization (SPAD) is a promising hybrid sample preparation technique combining the clean-up and preconcentration of the sample in a single step. In this work, a novel SPAD method based on the preparation of trimethylsilyl (TMS) derivatives of steroid hormones (testosterone, estrone, DHT, estriol, estradiol, and progesterone) in Phenomenex Strata C18-E (100 mg, 1 mL) cartridges has been developed and applied for their GC-MS/MS determination in human urine samples. The proposed procedure allows the detection and quantification of steroids with limits of 1.0-2.5 and 2.5-5 ng/mL, respectively. These characteristics are comparable with those obtained with a conventional liquid-liquid extraction, while the recovery of analytes in the proposed SPAD procedure is higher. The major advantages of SPAD are a short derivatization time, high efficiency, and the possibility to automatize the procedure. However, its cost-effectiveness in routine practice is still questionable.

Optimization and validation of LLE-LTP to determine florpyrauxifen-benzyl herbicide in water samples by HPLC-DAD.

Florpyrauxifen-benzyl is a systemic herbicide which acts on weeds of common occurrence in rice cultivation areas using flooded or rainfed systems. The Brazilian Health Regulatory Agency (ANVISA) authorized the commercialization of this pesticide, but did not establish guidelines with an extraction and quantification method of residues of this compound as a form of environmental monitoring. Therefore, the present study aimed to optimize and validate the liquid-liquid extraction with low temperature purification (LLE-LTP) to determine florpyrauxifen-benzyl content in water samples by high performance liquid chromatography with diode array detection (HPLC-DAD). The recovery ranged from 95.84 to 105.4% with a relative standard deviation less than 1.5. LLE-LTP was selective, precise, accurate, linear in the range from 4.00 to 150 µg L-1, and the limit of quantification was 4.00 µg L-1. The stability study of the compound in water revealed its degradation in 25 days and DT50 in approximately 5 days. LLE-LTP coupled to HPLC-DAD presented itself as a simple, easy and efficient method of extracting and analyzing it in water samples.

Separation of Benzene and Cyclohexane Using Eutectic Solvents with Aromatic Structure.

The separation of benzene and cyclohexane is a challenging process in the petrochemical industry, mainly because of their close boiling points. Extractive separation of the benzene-cyclohexane mixture has been shown to be feasible, but it is important to find solvents with good extractive performance. In this work, 23 eutectic solvents (ESs) containing aromatic components were screened using the predictive COSMO-RS and their respective performance was compared with other solvents. The screening results were validated with experimental work in which the liquid-liquid equilibria of the three preselected ESs were studied with benzene and cyclohexane at 298.5 K and 101.325 kPa, with benzene concentrations in the feed ranging from 10 to 60 wt%. The performance of the ESs studied was compared with organic solvents, ionic liquids, and other ESs reported in the literature. This work demonstrates the potential for improved extractive separation of the benzene-cyclohexane mixture by using ESs with aromatic moieties.