Sample Preparation Parameters Research Articles

A Critical Review on the Factors Affecting the Bond Strength of Direct Restorative Material Alternatives to Amalgam.

This article comprehensively reviews the performance of simplified direct restorative materials that have the potential to be an alternative to amalgam. Following an understanding of the material structures and clinical performances, this review provides an analysis of the bonding mechanisms and influential factors on the bond strength. These factors include substrate-related variations, involving primary vs. permanent dentin, sound- vs. caries-affected/demineralized dentin comparisons and surface-related factors and pretreatments. Special attention is given to the factors changing the substrate surface, such as different contaminants, remedy methods after contamination and different conditioning methods related to the materials. Variations in sample preparation and bond strength test parameters are also evaluated for the analysis of the outcomes. This review aims to provide an overview of the factors involved in the application procedure of direct restorations together with in vitro testing variations to guide the selection of suitable materials by understanding strengths and shortcomings.

Optimized high-throughput protocols for comprehensive metabolomic and lipidomic profiling of brain sample

Establishing direct causal and functional links between genotype and phenotype requires thoroughly analyzing metabolites and lipids in systems biology. Tissue samples, which provide localized and direct information and contain unique compounds, play a significant role in objectively classifying diseases, predicting prognosis, and deciding personalized therapeutic strategies. Comprehensive metabolomic and lipidomic analyses in tissue samples need efficient sample preparation steps, optimized analysis conditions, and the integration of orthogonal analytical platforms because of the physicochemical diversities of biomolecules. Here, we propose simple, rapid, and robust high-throughput analytical protocols based on the design of experiment (DoE) strategies, with the various parameters systematically tested for comprehensively analyzing the heterogeneous brain samples. The suggested protocols present a systematically DoE-based strategy for performing the most comprehensive analysis for integrated GC-MS and LC-qTOF-MS from brain samples. The five different DoE models, including D-optimal, full factorial, fractional, and Box-Behnken, were applied to increase extraction efficiency for metabolites and lipids and optimize instrumental parameters, including sample preparation and chromatographic parameters. The superior simultaneous extraction of metabolites and lipids from brain samples was achieved by the methanol-water-dichloromethane (2:1:3, v/v/v) mixture. For GC-MS based metabolomics analysis, incubation time, temperature, and methoxyamine concentration (10 mg/mL) affected metabolite coverage significantly. For LC-qTOF-MS based metabolomics analysis, the extraction solvent (methanol-water; 2:1, v/v) and the reconstitution solvent (%0.1 FA in acetonitrile) were superior on the metabolite coverage. On the other hand, the ionic strength and column temperature were critical and significant parameters for high throughput metabolomics and lipidomics studies using LC-qTOF-MS. In conclusion, DoE-based optimization strategies for a three-in-one single-step extraction enabled rapid, comprehensive, high-throughput, and simultaneous analysis of metabolites, lipids, and even proteins from a 10 mg brain sample. Under optimized conditions, 475 lipids and 158 metabolites were identified in brain samples.

Optimization of an analytical method based on miniaturized matrix solid-phase dispersion combined with gas chromatography-mass spectrometry for the determination of bisphenols in mussel samples

In this study, a new method has been optimized to determinate nine bisphenols (BPs) in mussel samples. This approach is based on miniaturized matrix solid-phase dispersion combined with gas chromatography-mass spectrometry determination after trimethylsilylation with 99% N,O-Bis-(trimethylsilyl)trifluoroacetamide + 1 % trimethylchlorosilane. The optimization process investigated the derivatization (derivatizing agent volume, time and temperature) and sample preparation (C18 co-column packed and C18 dispersant agent amounts and elution volume) parameters. A second clean-up step was performed using EMR-Lipid cartridges in order to efficiently remove lipids from mussel, the optimized parameters were Captiva EMR-Lipid sorbent amount and elution volume. The whole method was validated and very good linearity (r2 > 0.99) was obtained. The recoveries (accuracy) at two concentration levels ranged from 51.2 to 109 % and the coefficients of variation (precision) were between 0.55 and 13 %. The limits of quantification were between 0.014 μg/Kg dry weight for BPM and 4.0 μg/Kg dry weight for BPS. The analytical procedure was applied to four mussel samples from Galician Rías. Except for one of four samples whose BPAF level was below the limit of quantification, all BPs were found in concentrations that ranged from 0.39 to 301 μg/Kg dry weight.

Use of turbidimetry for determination of heat treatment intensity applied at pasteurization of milk

Express methods for estimating the intensity of heat treatment of milk are necessary in industry and in research work. For this reason, there are many ways to measure this parameter, which are based on different physical principles, including turbidimetric methods. The Harland & Ashworth turbidimetric method has been developed for a long time, however, due to its high reliability and ease of implementation, it is still used in practice. However, this method has a drawback: it takes a long time to perform the analysis. In this regard, the aim of the work is to develop an express method for evaluating the thermal class of milk based on the principle of measuring the concentration of soluble whey proteins. The result is achieved through the use of a turbidimetric measurement method with optimized sample preparation parameters and parameters for measuring the optical density of a suspension of protein aggregates. The method is implemented as follows. The milk sample is mixed with 0.1 N acetate buffer (pH 4.6) in a ratio that allows to obtain a concentration of soluble milk whey proteins from 0.05% to 0.1%. Recommended dilution coefficients: 1:3 for samples of ultra-pasteurized milk and pasteurized milk with high heat treatment intensity; 1:7 for samples of pasteurized milk with low heat treatment intensity and 1:14 for raw milk samples. The solution is filtered on a membrane filter with a pore size of 0.45 microns. The resulting filtrate is mixed with 24% trichloroacetic acid (TCA) in a 1:1 ratio to coagulate soluble whey proteins and form protein aggregates. After holding for 5–10 minutes, the optical density of the suspension of protein aggregates is measured at a wavelength of 650 nm. The content of water-soluble whey proteins in the sample can be calculated according to the calibration curve. The developed method allows to obtain the measurement result in less time than the Harland & Ashworth turbidimetric method.

FEATURES OF STRUCTURE, SUBSTRUCTURE, AND RESIDUAL STRESS OF Ti41.5Zr41.5Ni17 QUASICRYSTALS QUENCHED FROM LIQUID STATE

The structural state, stress, and substructure parameters of ribbons obtained by rapid solidification from the liquid state of the Ti41.5Zr41.5Ni17 alloy and powders obtained by deformation were investigated using X-ray diffraction and SEM microscopy methods. The influence of sample preparation parameters such as quenching rate and intensity of deformation, on the content of the Ti41.5Zr41.5Ni17 quasicrystalline phase and the crystalline phase of the approximant (W-phase), is considered. It is noted that the volume content of the W-phase increases with higher deformation intensity and decreases with higher quenching rate. Accumulation of specific defects in the quasicrystalline structure, known as phason defects, occurring during quenching, is observed in ribbon samples. The concentration of phason defects is nonuniform over the cross section of the samples, increasing from the side of the ribbon in contact with the quenching wheel to the free surface. This non-uniformity may be associated with the participation of phason defects at the initial stage of the phase transformation during the formation of the W-phase. The concentration of phason defects significantly increases during deformation of ribbons.

Creation and experience of using a multi-method for controlling pesticides in grain on the territory of the Russian Federation

This article is devoted to the development and testing of a multimethod for determining pesticides in grain, legume and oil crops using gas chromatography with mass spectrometric detection. Optimal parameters of sample preparation and chromatograph-mass spectrometric determination for the simultaneous analysis of 430 substances were established. Metrological characteristics such as uncertainty, repeatability and precision were calculated. The method was tested at the Moscow branch of the All-Russian Research Institute of Animal Husbandry on 120 samples of grain, legume and oil crops; these data are also presented in the article.

Successful 3D imaging of cleared biological samples with light sheet fluorescence microscopy.

In parallel with the development of tissue-clearing methods, over the last decade, light sheet fluorescence microscopy has contributed to major advances in various fields, such as cell and developmental biology and neuroscience. While biologists are increasingly integrating three-dimensional imaging into their research projects, their experience with the technique is not always up to their expectations. In response to a survey of specific challenges associated with sample clearing and labeling, image acquisition, and data analysis, we have critically assessed the recent literature to characterize the difficulties inherent to light sheet fluorescence microscopy applied to cleared biological samples and to propose solutions to overcome them. This review aims to provide biologists interested in light sheet fluorescence microscopy with a primer for the development of their imaging pipeline, from sample preparation to image analysis. Importantly, we believe that issues could be avoided with better anticipation of image analysis requirements, which should be kept in mind while optimizing sample preparation and acquisition parameters.

A tool to assess analytical sample preparation procedures: Sample preparation metric of sustainability

Sample preparation is a key step in most analytical methods, generally regarded as the least green step of the entire procedure. The existing green metrics assess the greenness of sample preparation techniques through the evaluation of the whole analytical procedure: including sampling, sample preparation, and the final detection/quantitation. Such inclusion of the entire method makes assessing the sustainability of a newly developed sample preparation technique quite challenging, as many aspects not solely linked to the sample preparation step are unavoidably considered. Thus, an alternative metric that can explicitly and exclusively evaluate the sample preparation is proposed. The metric is simple; it reports the result with a clock-like diagram, displaying the greenness outcome of main sample preparation parameters and a total score. This new metric can differentiate closely related microextraction approaches in terms of sustainability. The metric is also open-source and can be used by downloading the Excel sheet provided.

Biological function of Extracellular Vesicles (EVs): a review of the field.

Extracellular vesicles (EVs) theranostic potential is under intense investigation. There is a wealth of information highlighting the role that EVs and the secretome play in disease and how these are being utilized for clinical trials and novel therapeutic possibilities. However, understanding of the physiological and pathological roles of EVs remain incomplete. The challenge lies in reaching a consensus concerning standardized quality-controlled isolation, storage, and sample preparation parameters. Interest in circulating EV cargo as diagnostic and prognostic biomarkers is steadily growing. Though promising, various limitations need to be addressed before there can be successful, full-scale therapeutic use of approved EVs. These limitations include obtaining or manufacturing from the appropriate medium (e.g., from bodily fluid or cell culture), loading and isolating EVs, stability, and storage, standardization of processing, and determining potency. This review highlights specific topics, including circulation of abnormal EVs contribute to human disease and the theranostic potential of EVs. Theranostics is defined as a combination of the word's therapeutics and diagnostics and describes how a specific medicine or technique can function as both. Key findings include, (1) EVs and the secretome are future theranostics which will be utilized as both biomarkers for diagnosis and as therapeutics, (2) basic and translational research supports clinical trials utilizing EVs/secretome, and (3) additional investigation is required to fully unmask the theranostic potential of EVs/secretome in specific diseases and injuries.

Miniaturized affinity chromatography: A powerful technique for the isolation of high affinity GAGs sequences prior to their identification by MALDI-TOF MS

Glycosaminoglycans (GAGS) are involved in many biological processes through interactions with a variety of proteins, including proteases, growth factors, cytokines, chemokines and adhesion molecules. Identifying druggable GAG-protein interactions for therapeutic purposes is a challenge for the analytical community. In this context, this work investigates the use of a new miniaturized monolithic affinity column (poly(GMA-co-MBA) grafted with antithrombin III (AT III)) to specifically capture and elute high affinity sequences contained in low molecular weight heparin (enoxaparin) for further on-line characterization. This miniaturized, high binding capacity affinity column allows the specific capture of high-affinity oligosaccharide chains from Enoxaparin, even at low concentrations and with a minimal consumption of AT III. In addition to purification, this elution process enables preconcentration for direct analysis by capillary zone electrophoresis. It was found that many of oligosaccharide chains in enoxaparin were eliminated and that certain chain sequences were retained and enriched. Direct coupling with MALDI-TOF MS was successfully used to further characterize the specifically retained oligosaccharides where nano-ESI-TOF MS failed. After optimization of the sample preparation and ionization parameters, direct on-line analysis was performed by applying the elution volume released from the miniaturized affinity column (≤1 μL) directly to the MALDI plate. Finally, this original miniaturized analytical workflow coupling miniaturized AT III-affinity chromatography to MALDI-TOF MS detection is able to select, enrich and detect and identify high affinity sequences (mainly DP4 in size length with a high degree of sulfation) from low molecular weight heparin samples. A more specific selection of GAG sequences can be achieved by increasing the ionic strength during the washing step of affinity chromatography. This is consistent with the known binding pattern between heparin and AT III.

A Denoising Autoencoder for Improved Kikuchi Pattern Quality and Indexing in Electron Backscatter Diffraction

The rapid collection and indexing of electron diffraction patterns as produced via electron backscatter diffraction (EBSD) has enabled crystallographic orientation and structural determination, as well as additional property-determining strain and dislocation density information with increasing speed, resolution, and efficiency. Pattern indexing quality is reliant on the noise of the collected electron diffraction patterns, which is often convoluted by sample preparation and data collection parameters. EBSD acquisition is sensitive to many factors and thus can result in low confidence index (CI), poor image quality (IQ), and improper minimization of fit, which can result in noisy datasets and misrepresent the microstructure. In an attempt to enable both higher speed EBSD data collection and enable greater orientation fit accuracy with noisy datasets, an image denoising autoencoder was implemented to improve pattern quality. We show that EBSD data processed through the autoencoder results in a higher CI, IQ, and a more accurate degree of fit. In addition, using denoised datasets in HR-EBSD cross correlative strain analysis can result in reduced phantom strain from erroneous calculations due to the increased indexing accuracy and improved correspondence between collected and simulated patterns.

The Mechanical Properties Improvement of Polyurethane Rubbers by Using Titanium Oxide and Calcium Carbonate Modifiers (Additives)

The structure of polyurethane and the physico-chemical properties of its components are considered. The most promising modifiers have been selected: TiO2, CaCO3. The features of the preparation of the studied polyurethane mixture, its composition, manufacturing technology and equipment are described in detail. The geometric properties, manufacturing processes and testing methods of samples are presented. The graphical dependencies of the influence of the parameters of sample preparation on their mechanical properties such as hardness, tensile stress at break, Jung s modulus are given. The rational parameters of the technology for the production of samples modified with fillers to give them stable mechanical properties have been set. The maximum proportions of these fillers in polyurethane rubber are limited to 30-35%. It is proved that the addition of TiО2 and CaCО3 to the composition of polyurethane rubbers gives them special properties that allow them to expand the range of their industrial usage.

Ultrasound assisted dispersive solid phase microextraction using polystyrene-polyoleic acid graft copolymer for determination of Sb(III) in various bottled beverages by HGAAS

A new polyoleic acid-polystyrene (PoleS) block/graft copolymer was synthesized and applied as adsorbent for ultrasound assisted dispersive solid phase microextraction (UA-DSPME) of Sb(III) in different bottled beverages and analysis using hydride generation atomic absorption spectrometry (HGAAS). Adsorption capacity of the PoleS was 150 mg g−1. Several sample preparation parameters such as sorbent amount, solvent type, pH, sample volume and shaking time were optimized (based on central composite design (CCD) approach) and evaluated in respect to the recovery of Sb(III). The method revealed a high tolerance limit of matrix ions presence. Under optimized conditions, linearity range, the limit of detection, the limit of quantitation, extraction recovery, enhancement factor, preconcentration factor were 5–800 ng L−1, 1.5 ng L−1, 5.0 ng L−1, 96%, 82, 90, respectively. Accuracy of the UA-DSPME method was confirmed based on different certified reference materials and standard addition method. Factorial design was utilized to estimate the influences of variables of recovery of Sb(III).

Preparation of High-Quality Samples for MEMS-Based In-Situ (S)TEM Experiments.

A novel focused ion beam (FIB)-based methodology for the preparation of clean and artifact-free specimens on micro-electro-mechanical-system (MEMS)-based chips for in-situ electrical and electro-thermal experiments in a (scanning) transmission electron microscope ((S)TEM) is introduced. Owing to an alternative geometry, the lamellae are attached to a MEMS-based chip directly after the lift-out procedure and afterward further treated or thinned to electron transparency. The quality of produced lamellae on a chip resembles the quality of a classical FIB-prepared sample that is here demonstrated by high-resolution STEM imaging and analytical techniques. Various sample preparation parameters and the performance of in-situ prepared samples have been evaluated through electrical-biasing experiments.

Development, validation, and implementation of a robust and quality control-friendly focused peptide mapping method for monitoring oxidation of co-formulated monoclonal antibodies.

Monoclonal antibody (mAb) coformulation containing two therapeutic proteins provides benefits of improved therapeutic efficacy and better patient compliance. Monitoring of the individual mAb stability in the coformulation is critical to ensure its quality and safety. Among post-translational modifications (PTMs), oxidation is often considered as one of the critical quality attributes (CQAs) as it potentially affects the structure and potency. Although hydrophobic interaction chromatography (HIC) and reversed phase liquid chromatography (RPLC) have been used to monitor overall protein oxidation, mass spectrometry of peptide digests resolved by LC methods can afford superior selectivity and sensitivity for specific PTMs. With the advent of the Quadrupole Dalton (QDa) mass spectrometer as an affordable add-on detector, implementation of targeted oxidation assays in development and quality control (QC) laboratories is now feasible. In this study, as the first effort to implement MS-based methods for antibody coformulation in QC laboratories, we developed and validated a high-throughput and robust focused peptide mapping method using QDa for simultaneous site-specific monitoring of oxidation of methionine and tryptophan residues in heavy-chain (HC) complementary determining regions (CDRs) of two co-formulated mAbs. The method was validated in terms of accuracy, precision, linearity, range, quantitation limit (QL), specificity, and solution stability per recommendations in ICH Q2. The method robustness was systematically assessed involving multiple sample preparation and instrument method parameters. The method met the validation criteria in GMP laboratories with excellent robustness and was implemented in both GMP and development environments.

Evaluation of a Raman Chemometric Method for Detecting Protein Structural Conformational Changes in Solution

Raman scattering shows promise as a powerful routine tool, to determine both secondary and the smaller tertiary structural changes that precede aggregation in both solutions and solids. A method was developed utilizing principal component analysis (PCA) of Raman spectra for detection of small, but meaningful, pH induced changes in tertiary protein structure linked to aggregate formation using α-lactalbumin solutions as a model. The sample preparation and spectral parameters, were optimized for a bulk Raman probe. Analysis of large regions (600–1850 cm−1) yielded principal component (PC) scores useful for semi-quantitative comparison of protein conformation between formulations. PC loadings corresponded to specific structural peaks known to change with solution pH. PCA of circular dichroism (CD) spectra of dilute solutions yielded similar results. Sucrose is a common formulation excipient with a Raman spectrum that overlaps many protein peaks. With sucrose in the protein solution, the ability of PCA to discern protein structural changes from the Raman spectra was somewhat reduced. Analysis of a more limited spectral region (1530–1780 cm−1) with negligible sucrose spectral contribution improved the discrimination of protein conformational states. The new Raman method accurately distinguished differences in protein structure in concentrated solutions. The long-term goal is to explore Raman characterization as a routine monitoring tool of protein stability in both solution and solid states.

Expanding the use of cork as biosorbent through the determination of new psychoactive substances in postmortem blood samples

This study aimed to investigate the applicability of cork in a solid phase-based extraction method coupled to a 96-deepwell plate for LC-MS/MS determination of new psychoactive substances and other drugs of abuse in blood. Nowadays, there is a concern in developing new analytical methods that follows the principles of “green analytical chemistry”. These include the miniaturization of methods, utilization of minimal sample and solvent volumes, development of greener solvents and sorbents, among others. Regarding sorbents, natural compounds such as cork can be a great choice to be employed as extraction phase due to its large availability and low cost. Nevertheless, these new materials can help overcome analytical challenges related to postmortem analysis. Regarding sample preparation, blood samples were primarily submitted to protein precipitation by mixing 200 μL of blood, 250 μL of water, 250 μL of zinc sulfate and 50 μL of NaOH, followed by agitation and centrifugation. An aliquot of 500 μL of the supernatant was added to a 96-deepwell plate containing 30 mg of cork powder with particle size ≤ 250 μm. Extraction was performed by agitation for 15 minutes in an orbital shaker at 300 rpm, with consecutive removal of the aqueous phase. Desorption was performed by the addition of 1150 μL of acetonitrile with further agitation at 300 rpm for 2 minutes. The organic layers were recovered and dried under nitrogen stream, reconstituted in 30 μL of methanol and 2 μL was injected in the analytical system. Sample preparation parameters were optimized through univariate and multivariate strategies with the software Statistica 8.0 (Statsoft, Tulsa, USA). A Nexera UFLC system coupled to a LCMS-8045 triple quadrupole mass spectrometer (Shimadzu, Japan) was used for the analysis. Separation was performed on an Acquity UPLC® BEH C18 (1.7 μm, 2.1 × 50 mm) column (Waters, USA) using water (A) and acetonitrile (B) both containing 0.1% formic acid as mobile phases. Total run time was 7.5 minutes. The method was validated according to ANSI/ASB Standard 036 for the following parameters: lower limit of quantification (LLOQ), linearity, carryover, selectivity, precision, bias and dilution integrity. To verify the applicability of the method, three real postmortem blood samples were analyzed. The optimization of extraction parameters was carried out for the following items: choice of cork conformation (powder or sheet), cork particle size, cork mass, extraction time, desorption time, solvent type and solvent volume. The best parameters values are described in “Methods” session. LLOQs ranged between 0.5 and 5 ng/mL. Calibration curves were linear through the ranges of 0.5–85 ng/mL for ethylone, MDA, MDMA, N-ethylpentylone, 25B-NBOMe, 25C-NBOMe, 25I-NBOMe, fentanyl, pentylone and LSD; 1–85 ng/mL for cathinone, mephedrone and PCP; and 5–100 ng/mL for 25-NBOH, with determination coefficient (r2) > 0.99 and absence of carryover effects. Bias, within-run and between-run precision and dilution integrity results were within the acceptable values. The three samples were positive for the presence of MDA and MDMA with concentrations ranging between 23.11–26.59 ng/mL and 79.21–198.13 ng/mL, respectively. Sample preparation optimization was found to be relevant for the achievement of better analytical responses. Additionally, the use of the 96-deepwell plate allows simultaneous processing of multiple samples, with further possibility of automation. This study demonstrate that cork can be employed as a biosorbent for the determination of NPS and other drugs of abuse in postmortem blood through proper optimization, validation and application of the methodology. Therefore, it can be an alternative extraction phase to be considered in method development for toxicological purposes.

Optimising confocal Raman microscopy for spectral mapping of cement-based materials

Raman spectroscopy combined with confocal imaging, i.e. confocal Raman microscopy (CRM) is a relatively new technique with huge potential for high-resolution chemical mapping of phase composition and spatial distribution in cement-based materials. However, the effects of sample preparation and various operating parameters on mapping quality has not been systematically studied. This paper optimises CRM for spectral mapping of carbonated and non-carbonated cement-based materials. The effects of sample preparation and scanning parameters on the detection of four main phases (calcite, portlandite, ettringite and unreacted cement) were investigated. Results show that although freshly cut cementitious samples can be analysed as-is, the Raman signals improve with short gentle drying and surface grinding/polishing prior to analysis. Increasing laser power, exposure time and scan accumulation, and short laser wavelength yields higher signal-to-noise (SNR) ratio in the obtained spectrum. The use of a 4.15 mW laser power, 2 s exposure time and scan accumulation of 2 with 532 nm laser represents a good operating condition for Raman analysis of cement-based materials. This produces SNR > 10 for all investigated phases at short testing time and low risk of laser-induced damage. Microcracking caused by localised heating during closely-spaced mapping can be limited by impregnating the sample with epoxy to protect the microstructure. We show for the first time that CRM can be used to quantify the volume fraction of calcium carbonate and portlandite at high resolution when combined with SEM. The advantages and limitations of CRM for mapping cement-based materials are discussed.

High-Throughput Single-Step plasma sample extraction optimization strategies with experimental design for LC-MS and GC–MS integrated metabolomics and lipidomics analysis

To acquire deeper insights into the underlying molecular mechanisms in biological systems, a comprehensive and in-depth examination of a wide diversity of molecular species is necessary. Especially, it is very crucial to examine intermediary molecular levels such as protein, metabolite, and lipid to demonstrate direct causative and functional linkages between genotype and phenotype. Therefore, multiple sample sets/aliquots and analytical methods are required to cover different intermediary molecular levels due to the dissimilarity of physicochemical characteristics of biomolecules. However, existing methods used for simultaneous analysis of metabolites and lipids have not been thoroughly tested for reproducibility and wide applicability. Here, we have developed and optimized high-throughput and robust metabolomics and lipidomics analysis using experimental design strategies for single-step extraction protocols and also method parameters. Due to the high number of variables, an approach based on a desirability function was applied to optimize sample preparation and chromatographic parameters of metabolomics and lipidomics analysis. In these experiments, metabolite extraction efficiency was tested with acetone, acetonitrile, ethanol, methanol, water, and their combination, while lipids with hexane, chloroform, dichloromethane, and methyl tert-butyl ether. In the presented study, it is found that methanol:water:chloroform (3:1:3, v/v/v) mixture was superior to the other extraction solvent combinations in all performance measures. Moreover, the derivatization steps in GC–MS and chromatographic parameters in LC-qTOF-MS were tested. The derivatization efficiency increased with a higher concentration of methoxyamine hydrochloride and incubation time with the derivatization reagent. For LC-qTOF-MS, reconstitution solution, and the column temperature was found critical for high throughput metabolomics and lipidomics analysis, respectively. This systematic optimization for sample preparation and method parameters for integrated GC–MS and LC-qTOF-MS allows simultaneous and high-throughput analysis of metabolites as well as lipids from a single sample, making it a simple and practical strategy for different sample types.

Solvent and Flow Rate Effects on the Observed Compositional Profiles and the Relative Intensities of Radical and Protonated Species in Atmospheric Pressure Photoionization Mass Spectrometry.

Sample preparation and instrument parameters have regularly been demonstrated to impact upon the observed results in atmospheric pressure photoionization, mass spectrometry (MS), and analytical techniques in general but may be overlooked when such methods are applied to the characterization of real-world samples. An initial investigation into different solvent systems demonstrated that the inclusion of ethyl acetate inverted the ratio of relative intensities of radical and protonated species (R/P). Design of experiments was performed and indicated that the injection flow rate is also a significant factor. The impact of the solvent system and flow rate on signal intensity, the observed compositional profile, and R/P of selected molecular groups is demonstrated further. An inversion of R/P is observed at higher flow rates in solvent systems commonly used in petroleomics studies, effecting a loss of molecular speciation. The findings presented reiterate the critical importance in considering experimental parameters when interpreting the results of analytical procedures.